Copyright 0 1984 by the Genetics Society of America

MORPHOGENETICALLY SPECIFIC MUTABILITY IN DROSOPHILA ANANASSAE

CLAUDE W. HINTON

Department of Biology, The College of Wooster, Wooster, Ohio 44691

Manuscript received September 5, 1983 Revised copy accepted December 24, 1983

ABSTRACT

A stock exhibiting hypermutability with respect to visible mutants (Om) affect- ing optic morphology was subjected to genetic analysis. The production of Om mutants, independently recovered with a frequency of two per IO4, is restricted to females and depends primarily on homozygosity of their X chromosomes; in heterozygotes, Om mutability is stimulated by the presence of either one of two extrachromosomally replicating elements previously identified in other stocks having cryptic mutability systems. The semidominant and nonpleiotropic Om mutants are not associated with gross rearrangements and they map to at least 15 loci. Most of the loci defined by mapping are represented by two or more Om mutants which, despite considerable interlocus mimicry, sometimes display locus- specific phenotypes. O m mutants are moderately unstable, and they are subject to dominant suppressors that arise spontaneously at either of two X-linked loci. An interpretation of these observations invokes an X-linked transposable element (tom) that specifically inserts into control sequences shared by a set of structural genes involved in eye morphogenesis.

OR many years, rarity and randomness have persisted as fundamental prop- F erties of spontaneous mutation despite the identification of mutable genes and mutator genes (e.g., MCCLINTOCK 1956; GREEN 1976); such phenomena were generally regarded as special cases or experimental aberrations of the normal mutation process. But recent molecular analyses of mobile middle repet- itive DNA sequences in eukaryotes show that transposable elements are common to many of the formerly disparate and apparently special cases (see SHAPIRO 1983 for a timely collection of reviews in this area). Rules for the distribution of transposable elements in the genome remain elusive, but they are clearly involved in high frequency and nonrandom mutational events.

As a matter of historical interest, SPENCER (1935) reported from this depart- ment his observations on several species of Drosophila suggesting that mutability is temporally nonrandom. Recent analyses of a small number of D . ananassue stocks have exposed and partially defined one overt and two cryptic hypermu- tability systems (HINTON 1979, 198 1, 1983) that confirm KIKKAWA’S (1 938) impression of high mutability in this species. In this report, a fourth system is described in which an entirely novel form of nonrandomness occurs: the visible mutants are limited to those disrupting morphogenesis of the eye.

Genetics 106 631-653 April, 1984.

632 C. W. HINTON

MATERIALS AND METHODS

Analysis of the ca (claret eye color, 2L) stock revealed a cryptic mutability system composed of an extrachromosomally transmitted mutator element responsible for chromosome damage in sperm and a chromosomally specified suppressor function (cytotype) responsible for repair, at fertilization, of the mutator-induced lesions. Thus, neither intrastock matings nor outcrosses of ca females to males from a standard reference stock ( p x , plexus wing venation, 3R) indicate any predisposition for unusual mutability, but the reciprocal outcross of ca stock males to p x females produces high frequencies of chromosome rearrangements and visible mutants, especially dominant Minutes, among the F1 progeny (HINTON 1981). To assign the putative Minute mutants to linkage groups, a ca; p x tester stock was constructed in 1979 by selecting Fn double mutant homozygotes from a mass mating of p x stock females to ca stock males. Subsequent use of the ca; p x stock for this purpose was marked by the recovery, among relatively small testcross progenies, of seven independent semidominant mutants with phenotypes resembling the familiar Bar eye mutant of D. melanogaster. Prior to this episode, only three dominant eye morphology mutants (Puffed, Lobe and Star) were known in D. ananassae.

Recovery of additional optic morphology mutants (symbolized Om) from ca; p x intrastock matings identified this stock as their source. The genetic basis for Om mutability was examined in two separate mating schemes consisting of outcrosses and backcrosses between the ca; p x stock and its antecedents, the ca and p x stocks. The ca and ca; p x stocks carry only standard (+) chromosome sequences, whereas the p x stock is homozygous for In(2L)A. A derivative of the ca stock, denoted ca Xg, was produced (HINTON 1981) by recurrently backcrossing F1 +/ca; fix/+ females and their daughters to males from the ca stock for nine generations to reconstitute the ca genome without the extrachromosomal mutator.

Om mutability was also examined with respect to another cryptic mutability system described (HINTON 1983) in a stock marked with pc (peacock wings, 3R). The pc system consists of a 3-linked dominant mutator that causes lesions in sperm chromosomes provided that an extrachromosomally inherited suppressor element, also present in this stock, does not intervene to prevent or repair the lesions. In addition to the mutator, the third chromosome of the pc stock bears a recombinationally separable, dominant male crossing over enhancer whose effect is also suppressed by an extrachro- mosomal factor which could be the Same as that suppressing mutability. The pc stock carries In(2L)A as does its derivative p c Xlo constructed, analogously to ca Xg, to provide the pc stock chromosomal genome without the extrachromosomal suppressor factor@).

Putative Om mutants were routinely diagnosed for transmission, and linkage groups were assigned by their segregation patterns with respect to sex and the heterozygous ca and/or p x markers. X - linked Om mutants were mapped using a stock having the scSs (scute, missing scutellar bristles), m6’ (miniature wings), and wl (white-tinged, eye color) markers widely spaced in the X chromosome. Another stock bearing the more closely linked markers cop (copper eye color), f” (forked bristles), Bx’ (Beadex, excised wing margins) and w g (white-golden eye color) was selected to monitor recombination in their vicinity. Om mutants assigned to the second chromosome were mapped using a stock carrying the + sequence and the markers e65 (ebony body color) and pea (peach eye color), and those showing linkage with the third chromosome p x marker were subsequently mapped using a stock marked with sfw (straw body color), p c and p r (purple eye pigmentation). Several stocks with other markers (HINTON 1980) were occasionally used for supplementary mapping. Because the Om mutants map to several loci within each of the three major chromosomes and most loci are represented by more than one mutant, each mutant is specifically symbolized as, for example, Om(ID)9, wheie the chromosome number and locus are designated in parentheses followed by an acquisition number. Some new Om mutants, recovered from parents having a previously identified Om mutant, are distinguished by a lower case letter following the acquisition number of the original mutant: e.g., Om(3C)9a.

Stocks and experimental matings were cultured in 25 X 95-mm shell vials containing 10 ml of a medium composed of corn meal, molasses, brewer’s yeast, agar and propionic acid and seeded with live baker’s yeast. Experimental crosses usually involved three or four, 3- to 4-day-old virgin adults of each sex per mating vial; after 7 days, the parents were transferred to a vial containing fresh medium for a second week’s progeny sample. Most matings of a specified type were replicated at different times, often with different lines of the ca; p x stock. In addition to eye morphology variants, the progenies were scored for Minute (M) mutants whose frequencies were used previously in analyses

MORPHOGENETICALLY SPECIFIC MUTABILITY 633

of the ca and pc mutator systems to quantitate variations in mutability. Rare visible mutants of other types were also observed, but neither these nor the Minutes were systematically tested for transmis- sion. Chi-squared contingency tests or tables prepared by KASTENBAUM and BOWMAN (1970) were used to evaluate the statistical significance of differences between numbers of mutants observed in different matings. Scanning electron micrographs of selected Om mutants were prepared in the Electron Microscopy Laboratory, Pathology Department, University of North Carolina.

RESULTS

The results of intrastock matings, pooled from intermittent assays of three separate but homogeneous lines of the c a ; p x stock, show (Table 1) 18 Om mutants in a total of 93,111 progeny, more conveniently expressed as 19.3 mutants/l O5 progeny. A fourth line of the ca; p x stock produced no Om mutants among a total of 27,472 offspring scored; matings involving this apparently stable line were excluded from all tabulations.

Outcrosses of ca; p x females to p x stock males produced almost as many Om mutants (14. 1/105) as intrastock matings, whereas the reciprocal outcross re- sulted in only one possible Om, a sterile male having one normal and one Bar- like eye. Thus, Om mutability appears to be restricted, entirely or largely, to ca; p x females. But F1 females from either reciprocal outcross, tested in matings to either p x or ca; p x males, produced, among 73,875 progeny, only one Om mutant whose parentage is ambiguous. The explanation that Om mutability is a function of a recessive gene was tested by assaying XI, X2 and XS females, randomly selected from recurrent backcrosses of F1, XI and XZ females to males from the ca; p x stock and expected to be Vz, Y 4 and '/8 homozygotes, respectively. These fractions of the rate of Om mutability observed for ca; p x homozygotes are reasonably approximated by the observed frequencies of 5.2 (XI), 8.6 (Xz) and 15.9 (X,) Om mutants per lo5 progeny after combining the output of females

TABLE 1

Visible mutants recovered in progenies from the ca; px and px stocks, their reciprocal outcrosses and their hybrids

~

Parents Progeny Parents Progeny

Mutants Mutants Total Total

Female Male scored Om Others Female Male scored Om Others

ca; px ca; px 93,111 18 36 M, 1 N , px px 23,212" 0 13 M

ca; px px 42,439 6 64 M, 1 D1, px ca;px

FI PX 29,760 0 9 M, 2 D1 FI PX 26,497 0 3 M FI ca; px 8,928 0 1 M F1 ca; px 8,690 1 0 X I ca; px 29,898 2 17 M XI ca;px 27,889 1 19 M, 1 Dl Xn ca; px 29,267 3 15 M , 1 D1 Xn ca; px 29,162 2 13 M, 2 D1 x3 px 12,250 3 6 M X3 ca; px 12,983 1 3 M P X FI 9,745 0 5 M Px FI 8,263 0 2 M ca; px FI 1,439 0 2 M ca; px FI 1,245 0 2 M XI px 17,540 7 1 1 M, 1 Cy X I PX 14,590 2 1 4 M

1 Dl 39,560 1 26 M, 1 Dl

l w

a From HINTON (198 1).

634 C . W. HINTON

derived from the reciprocal outcrosses. Among the 12 Om mutants recovered from X I , Xp and Xs females, eight descended from outcrossed ea; px females and four from daughters of outcrossed ea; px males; the absence of a significant difference suggests the absence of a major cytotypic or extrachromosomal factor in Om mutability. One other comparison, not shown in the tabulated results, is based on segregation of the ca marker from Zn(2L)A, +/+, ea heterozygotes in each generation (the inverted sequence includes ea+ and about V4 of 2L); +/ea and ca/ca daughters were assayed separately and produced seven and five, respectively, Om mutants among nearly equal progenies.

Except for the probable exclusion of 2L, the foregoing results provide no indication of the location of the postulated recessive gene responsible for Om mutability. The possibility of X linkage was examined by exploiting the different sources of X chromosomes in Fl males from reciprocal outcrosses. Neither group of F1 males produced any Om progeny (Table l ) , but they nevertheless differed as shown by the performance of their X1 daughters derived from backcrosses to ea; px stock females. The X 1 daughters with both X chromosomes from the ea; px stock produced Om progeny at the rate of 39.9/105 as compared with the yield of 13.7/105 from their heterozygous X 1 counterparts. Although the rates are not statistically distinguishable, the difference indicates the involvement of an X-linked factor, or factors, in Om mutability.

Given that the major difference with respect to Om mutability between the ea; px stock and its px antecedent is X linked, one may inquire whether the same difference exists between the ea; px and ca stocks; for this purpose, outcrosses between the ea; px and ea stocks were made (Table 2). It is consistent with previous experience that no Om and very few M mutants were observed among offspring from outcrossed ea; px males. From the reciprocal outcross of ea; px females, one Om mutant was recovered, and about 0.5% of the progeny were Minutes as expected if the ea males carry the extrachromosomal mutator and if

TABLE 2

Visible mutants recovered in progenies from the ca; px and ca stocks, their reciprocal outcrosses and their hybrids

Parents Progeny Parents Progeny

Mutants Mutants Total Total

Female Male scored Om Others Female Male scored Om Others

ca; px ca; px 93,111 18 36 M, 1 N , ca ca

ca; px cu 7,499 1 41 M ca ca; px P X FI 13,978 0 9 M P X FI ca; px FI 6,035 4 0 ca; px F1

1 Dl

XI P X 39,835 11 3 6 M , 1 dy X1 P X

FI Px 21,094 0 1 3 M FI p x ca XS cu; px FI P X

12,557"

4,653 10,370

1,217 20,560

21,359 2,442

12,782

0 11 M

0 2 M 0 65 M 0 0 2 23 M, 1 cx,

6 20 M 0 1 M 1 6 M

1 Pn

'From HINTON (1981).

MORPHOGENETICALLY SPECIFIC MUTABILITY 635

the ca; p x females lack the suppressor cytotype characteristic of ca stock females. That the extrachromosomal mutator was transmitted by ca stock females is shown by the high frequency of Minutes (0.6%) recovered from their F1 sons in assays to p x females. Backcrosses of F1 males to ca; p x females produced XI females of two types with respect to their X chromosomes. In parallel with the p x backcross results, X I females homozygous for the X chromosome from the ca; px stock produced more Om progeny (27.6/105) than the X1 heterozygotes (9.7,' lo5). Thus, the ca; p x stock appears to differ from both ancestral stocks in the same way with respect to Om mutability. Evidently, the X-linked factor responsible for Om mutability is not strictly recessive.

Tests of F1 females from outcrosses between the ca; p x and ca stocks (Table 2) revealed an unanticipated reciprocal cross difference: whereas the daughters of ca; px females produced, as expected, no Om mutants, the chromosomally equivalent daughters of ca females produced Om progeny with a frequency (28.1/105) comparable to that of ca; p x homozygotes. This indication that the extrachromosomal mutator, transmitted from ca females to their daughters but not from ca males to their daughters, stimulates Om mutability in heterozygotes was checked with F1 females from the parallel outcross of ca XS females to ca; p x males. These F1 females, lacking the mutator element but otherwise comparable to the daughters of ca females, produced only one Om mutant among 12,782 progeny

The paradox that the ca mutator stimulated Om mutability in heterozygous females without significantly increasing Minute frequencies could be resolved if the relevant property of this element is its extrachromosomal replication rather than its mutator activity. This possibility prompted examination of the p c stock whose extrachromosomally replicating element(s) exhibits suppressor rather than mutator functions. The yield of mutants (Table 3) among relatively small progenies from reciprocal outcrosses between the ca; p x and pc stocks was consistent with previously established properties of these stocks. F1 males, with

TABLE 3

Visible mutants recovered in progenies from the ca; px and pc stocks, their reciprocal outcrosses and their hybrids

Parents Progeny Parents Progeny

Mutants Mutants Total Total

Female Male scored Om Others Female Male scored Om Others

ca; p x ca; p x 93,111 18 36 M , 1 N , pc P C 1 Dl

ca; p x pc 4,737 1 9 M P C ca; p x PX F1 10,101 0 11 M, 4D1, p x FI

FI PX 20,151 0 6 M FI P X pc XIO ca; p x FI Px

1 M D1, 1 N

12,480" 0 4 M , 1 Pu, 1 Exc, 2 ct, 2 sc, 1 rst

4,123 0 8 M 9,192 0 3 M , 1 DL

23,512 7 1 7 M 1,180 0 0

16,879 0 4 M

a From HINTON (1 983).

636 C . W. HINTON

or without the suppressor(s), produced no Om mutants in assays to px females, but they demonstrated the eapected reciprocal cross difference with respect to other visible mutants caused by the dominant p c mutator. Among F1 females from the reciprocal outcrosses, those carrying the extrachromosomal suppressor from their pc mothers produced seven Om mutants (29.8/105) as compared with none from those having ca; p x mothers. This result was controlled by tests of F1 daughters of p c Xlo mothers having no extrachromosomal element: no Om mutants were found among 16,879 progeny examined. Therefore, regardless of their function as mutator or suppressor, either one of the two self-replicating extrachromosomal elements is capable of stimulating Om mutability in hetero- zygous females to at least, if not significantly more than, the level observed in ca; p x homozygotes.

Most of the comparisons of Om mutant frequencies in the preceding analyses were based on numbers so small as to be of little or no statistical significance. Because of the parallelism between matings described separately (Tables 1 -3), it seems reasonable to pool their results to provide more reliable estimates of Om frequencies and to increase confidence in the observed differences (Table 4). All outcrossed ca; p x males produced a total of 5 1,958 offspring of which only one, the sterile mosiac, was possibly an Om mutant. All F1 males assayed in matings to p x females collectively had 61,649 offspring, none of which was an Om mutant. The conclusion that males do not contribute to Om mutants among their progeny permits aggregation of all progeny from females of a given genotype regardless of their spouses. Together, all ca; p x stock females used in outcrosses (or as the recurrent parent in backcrosses to F1 males) produced 12 Om mutants among 64,6 1 1 progeny; the frequency of 18.6/1 O5 is virtually the same as that of 19.3/ lo5 for ca; p x intrastock matings, and this identity reinforces the conclusion that

TABLE 4

Summaq compilation of data on visible mutant yields by comparable genotypes listed in Tables 1-3

Assayed parents

Total P'ogenY scored

No. of mu- tants

recovered Yield of mutants per IO5 ptogeny

Om Others Om Others

+; px intrastock matings ca; + intrastock matings +; pc intrastock matings ca; p x intrastock matings ca; p x males, outcrosses ca; p x females, outcrosses FI males, backcrosses to px females F1 females, without extrachromosomal element F1 females, with extrachromosomal element XI females, from backcrossed FI females Xs females, from backcrossed XI females Xs females, from backcrossed X2 females XI females, heterozygous for ca; px X chromosome XI females, homozygous for cu; px X chromosome

23,212 12,557 12,480 93,111 5 1,958 64,611 6 1,649

144,781 44,871 57,787 58,429 25,233 35,150 57,375

0 13 0 11 0 11

18 38 1 38

12 120 0 102 2 44

13 37 3 37 5 31 4 9 4 39

18 49

0 56.0 0 87.6 0 88.1

19.3 40.8 1.9 73.1

18.6 185.7 0 165.4 1.4 30.4

29.0 82.4 5.2 64.0 8.6 53.0

15.9 35.7 11.4 111.0 31.4 85.4

MORPHOGENETICALLY SPECIFIC MUTABILITY 63 7

Om mutability is restricted to females. All F1 females from ca; p x outcrosses can be assigned to one or the other of two groups, those with and those without an extrachromosomally replicating element. Thirteen Om mutants were recovered among 44,871 offspring of F1 females carrying either the ca mutator or the p c suppressor; this frequency of 29.0/1 O5 is not significantly different from the frequency for ca; p x stock females. However, F1 females lacking either of the extrachromosomal elements exhibited significantly lower frequencies of Om mutants (two among 144,781, or 1.4/105) than either ca; p x stock females or F1 females carrying an extrachromosomal element. The inference that Om mutabil- ity in ca; p x stock females depends primarily on homozygosity for some compo- nent of their X chromosomes is strengthened by comparing the performance of homozygous X1 daughters of backcrossed F1 males carrying the ca; p x stock X chromosome (18 Om mutants among 57,375 offspring, 31.4/105) with that of heterozygous X1 daughters of F1 males bearing X chromosomes from either the ca or p x stocks (four Om among 35,150 progeny, 1 1.4/105), but the difference is not quite significant at the 5% level of confidence. If, however, the yield of X1 homozygotes is compared with the combined yield of six Om among 179,931 progeny of all heterozygous females (the X1 plus F1 females without any extra- chromosomally replicating element), the difference becomes highly significant.

With respect to Minutes and other visible mutants detected in these assays (Table 4), the yield from ca; p x intrastock matings was lower than the background levels recorded for the p x reference stock or for the ca and pc stocks carrying cryptic mutability systems. These cryptic mutators were responsible for the inflated frequencies of non-Om mutants found in the progenies of outcrossed ca; p x females and F1 males: some of the ca; p x females were outcrossed to mutable ca males (Table 2), and some of the F1 males expressed either the ca (Table 2) or p c (Table 3) mutator. Females of the F1 and subsequent generations produced no substantial deviations from the background frequencies of Minutes. For visible mutants, then, the ca; p x stock presents no unusual proclivity other than Om mutability.

Among the total of 106 putative Om mutants identified in this study, ten were sterile and four others, excluded from the tabulated data because they were initially considered dubious Om candidates, failed to transmit their eye abnor- mality. Only one possible Om mutant, the sterile son of an outcrossed ca; p x male, was mosaic. Although most of the Om mutants occurred as single specimens, four cultures contained two mutants which, in all four cases, proved to be located in different linkage groups and, thus, originated in separate events if not in separate parents. The overall absence of clustering and mosaicism indicates that Om mutants arise in oocytic rather than oogonial or somatic cells.

Most of the 92 transmitted Om mutants are semidominant alleles, expressed in heterozygotes but more extremely in homozygotes. Of course, the detection of autosomal Om mutants in outcross progenies requires expression in heterozy- gotes, but X-linked recessive mutants could have been detected among the sons of mutable females, and their occurrence is suggested by the recovery sex ratio of 24 males to 12 females among X-linked Om mutants as compared with 26 males to 30 females among the autosomal mutants. Nevertheless, semidominance

638 C. W. HINTON

appears to be the rule among X-linked Om mutants because all are expressed to some extent in heterozygous females; the degree of expression, however, is so weak for some mutants that their detection as females would have been unlikely. It is also the rule that X-linked Om mutants are more strongly expressed in homozygous females than in hemizygous males; there is no sex differential in expression of autosomal mutants.

All 92 transmitted Om mutants were assigned to linkage groups with 36 in X, 40 in 2, 16 in 3, and none in the genetically small, but cytologically large, heterochromatic fourth chromosome. Genetic maps of the three major chro- mosomes each measure about 1 10 map units based on currently available markers (C. W. HINTON, unpublished results), and the relative polytene chromosome lengths (HINTON and DOWNS 1975) are 17 (X), 44 (2) and 39 (3). A dispropor- tionate number of X-linked Om mutants could be expected to occur through biased recovery of weakly semidominant alleles in hemizygous males, but the significant excess of 2-linked over 3-linked Om mutants cannot be ascribed to this source; it seems rather that the mutants are not randomly distributed between the chromosomes.

Sixty-three of the first 75 Om mutants to be detected were mapped. For the X chromosome (Figure 1 and APPENDIX Table Al) , the 26 mutants fall into six groups representing six rather widely spaced loci. Within groups represented by more than one independently mapped allele, the map values are reasonably homogeneous. A more direct test of the assumption that mutants within a group represent recombinational alleles was provided by females heterozygous for f Om(lD)9 Bx w and either Om2, Om5, Om16 or Om30 also assigned to group 1D; no recombinant Om+ progeny were found among the combined total of 6431 scored. Om(lE)59a was not mapped independently, but Om53/0m59a females

n Bx w I 1 1 1 .... sc CO? m f

I I II x .... I 'i ..........

A B c 15 23 3 2 53 35 24 46 I 1 5,9 59a

E F

25 52 9b,9e 37 42 47 50

a e pea R I 1 I I I .... 2 ....

11 I I I 1 I I A

13 17 I 8 28 33 49

B C D 14 I IOa 22 4 21,63

7,8 9 c . 9 f

i o , i K i . r a 19,26,27

31,32,40,44

E 9h

P 270

b r i Tr stw PC P* 3 .... I I 1 I I I I I ....

A B J 1 43 57

20 29 55

6 9.2 xa

FIGURE 1 .-Consensus maps of the X , second and third chromosomes showing the distribution of O m mutants with respect to standard markers (see APPENDIX Tables Al-A3 for data). Dotted termidi indicate genetically unknown regions, and the dotted interstitial segment of X represents about 30 map units.

MORPHOGENETICALLY SPECIFIC MUTABILITY 639

produced no Om+ recombinants among 301 sons scored. Six Om loci are also defined by the 2-linked mutants (APPENDIX Table A2), but four of the six loci, represented by 27 of the 29 mutants, lie within 16 map units of each other. Recombinational allelism within group 2C was further supported by testing females heterozygous for e Om8 pea and either Oml , Om9c, Om26, Om27, Om31 or Om32; among 6579 progeny, only one Om+ fly was scored, and it was apparently a revertant of Om8 because it carried both the e and pea parental markers. On the other hand, the separability of Om(2B) and Om(2C) was confirmed by four e Om+ pea recombinants among 630 offspring from testcrossed + Om14 pea/e Om7 + females. Only three loci were identified by the eight 3-linked Om mutants (AFPENDIX Table A3) and these were widely spread along the map (Figure 1) . Variations in the number of alleles per locus suggest that the 15 Om loci are not equally mutable.

Inspection of the mapping results (APPENDIX Tables A 1 -A3) shows that none of the mapped mutants is associated with a rearrangement sufficiently large to suppress recombination in heterozygotes. As mentioned before, the map value for the same intervals in independent tests of different alleles at the same locus are reasonably consistent. Furthermore, the values for the same marker interval with and without an intervening Om mutant are also consistent; compare, for example, the e-pea region of 6.5 map units for summed 2A mutants with those of 6.9 and 6.6 for 2B and 2C mutants, respectively. No indications of pseudo- linkage, indicative of translocation, were found in the course of assigning Om mutants to linkage groups. The conclusion that Om mutants are not accompanied by gross rearrangements is supported by the failure to detect any aberrations in the polytene chromosomes of larvae heterozygous for one or another of ten Om mutants.

Three mutants (Om39, 5 4 and 56), recovered from matings of F1 Zn(2L)A, +/ +, ca; p c / p x females to p x males (Table 3), failed to transmit the ca marker in backcrosses to the ca; p x stock and, in subsequent testcrosses of Om/e pea females, they were found to lie to the left of e with about 0.3% recombination. Several different markers located within Zn(2L)A exhibit the same recombination fre- quency with the e marker in structural heterozygotes (C. W. HINTON, unpub- lished results). The probable location of these three Om mutants within Zn(2L)A implies that Om mutability is not restricted to chromosome segments from the ca; p x stock.

In addition to the rule of semidominance among Om mutants, they also share certain other phenotypic properties. Most of them were easily established and maintained in homozygous stocks (the viability and fertility of Om homozygotes discounts their origin as deficiencies). Phenotypic observations on Om flies from homozygous stocks demonstrated their effects to be limited to eye tissue; in particular, no consistent abnormalities of antennae, ocelli or of bristles bordering the eye were seen. Most of the mutants share a reduction in eye size, particularly on the anterior side, and this is accompanied by various irregularities in omma- tidial arrangement, size and pigmentation; however, the eye field, as defined by the area within boundaries formed by bristles normally contiguous to the eye, is not obviously smaller than normal. In contrast to common experience with

640 C. W. HINTON

Drosophila mutants, the Om mutants exhibit not only remarkably little pleiotropy but also relatively little variation between individuals within a stock. These generalizations are not without exceptions, and, within the range of common effects, there is considerable phenotypic variation between independent Om mutants. No critical morphometric or developmental analyses of Om mutants have been

conducted, but superficial examination suffices to show that alleles sharing the same locus often share phenotypic peculiarities. This assertion is supported by the following phenotypic descriptions of homozygous females from stocks of the mapped mutants, supplemented by scanning electron micrographs of represent- ative specimens (Figure 2).

Om(IA)15, 24, 37, 42, 47, 50. The circumferential reduction of differentiated eye tissue in these mutants is conspicuously marked by an anterior indentation varying between stocks from a distinct V to a rectangle and the posterior margin is merely irregularly shaped, sometimes with a slight indentation. The anterior indentation frequently includes flecks of red pigment and supernumerary inter- ommatidial bristles but no isolated ommatidia. Rounded ommatidia are irregu- larly arranged adjacent to the indentations and tend to be orthogonally arranged elsewhere.

Om(lB)23, 46. These two mutants share a small reduction in the size of the eyes whose anterior margin, rather than being indented, is somewhat ragged. Irregular departures from the normal hexagonal array of ommatidia contribute to patchy roughening of the eye surface.

FIGURE 2.-Scanning electron micropphs showing typical eye phenotypes of homozygous fe- males from the m; p x and selected Om niutant stocks. See text for descriptions.

MORPHOGENETICALLY SPECIFIC MUTABILITY 64 1

Om(lC)3, 11, 52. All three of these mutants are semilethal and present distinc- tive depressions of the normally contoured eye surface in addition to circumfer- ential reduction of eye tissue. Om3 and Om1 1 are quite similar with respect to their extended clefts between the dorsal and ventral halves of the eyes, and the tissue within the cleft contains numerous free interommatidial bristles. However, Om52 shows only a small anterior indentation, and its surface is marked by scattered ommatidial fusions which may be either darkly pigmented or colorless. The suggested possibility that Om52 represents a separable locus has not been examined genetically.

Om(lD)2, 5, 9, 9b, 9e, 9g, 16, 30, 36, 45, 48. There is considerable quantitative variation between the 11 mutants of this set, but their eye tissue reductions are accompanied by more or less prominent anterior indentations and by jumbled ommatidia around the indentations; orthogonal arrays of ommatidia are com- monly seen beyond the indentation. In most stocks, there are supernumerary interommatidial bristles located adjacent to the indentation which, as it extends posteriorly, becomes a glazed, pigmented trough. However, in the case of Om9g, the mouth of the indentation is itself occupied by glazed tissue bearing irregular flecks of pigment and several bristles about twice the length of interommatidials; and, in the case of Om9e, the mouth of the indentation contains a textured swelling which bears bristles similar to those of Om9g. The reduction of eye tissue is more severe in Om9e and Om9g than in the other ID mutants.

Om(lE)53, 59a. These two mutants are uniquely characterized by an excess of differentiated eye tissue leading to the appearance of one eye superimposed on another; the posterodorsal position of the superimposed tissue also contrasts with the preferential anteroventral effects of most Om mutants. The ommatidia of Om53 and Om59a are normal and hexagonally disposed over large areas. Heter- ozygous females are identifiable by their enlarged but otherwise normal eyes.

Om(lF)35. The perimeter of Om35 eyes is only slightly reduced, largely at the jagged anterior margin, but the ommatidia in the anterocentral region tend to be enlarged and their irregular packing defaces the normal surface contour.

Om(2A)13, 17, 18, 28, 33, 49. Quantitative variation within this group is associated with eye shape distortions ranging from straight anterior margins (Om13 and Om28), through a narrow anterior indentation (Om18), to broad anterior concavities (Om1 7, Om33, Om49); posterior margins are also irregular in the moderate and extreme alleles. The anterior indentations are more or less filled with glazed tissue which is pigmented but otherwise undifferentiated. Both the anterior and posterior margins contain irregularly arranged ommatidia and interommatidial bristles.

Om(2B)14, 22. The presence of numerous free interommatidial bristles in the anterior indentation is the major feature distinguishing these two strong mutants from those of the preceding group 2A mutants.

Om(2C)l, 4, 7, 8, 9c, 95 10, 12, 14a, 19, 26, 27, 31, 32, 40, 44. Fifteen of the 16 mutants in this series are hardly distinguishable qualitatively, and they vary little quantitatively within the moderate to strong range. These 15 mutants typically have a reduced dorsal mass of ommatidia from which a narrow sinuous band of ommatidia extends ventrally; the ventral tip of the band tends to point anteriorly. Much of the extensive anterior concavity contains glazed tissue that

642 C. W. HINTON

is pigmented throughout. Free interommatidial bristles are seldom seen in the glazed tissue, but a small black scab-like structure is commonly found just anterior to the glazed region. Except for the dorsal mass, there is little regularity in ommatidial arrangement, and interommatidial bristles are scarce or absent. Were it not for its failure to recombine with Om8 (see mapping results; N = 788), there would be little to justify assignment of Om9c to this locus because its phenotype differs radically from the other 15 Om mutants at 2C or any other of the identified loci. The entire surface of the moderately reduced eye is glazed with only occasional suggestions of individual ommatidia; several dispersed interom- matidial bristles project from the glazed surface. The eye is slightly constricted along the equator, and the central areas of both the dorsal and ventral halves usually lack pigment.

Om(2D)lOa, 21, 63. The eyes of these mutants tend to be lozenge shaped or sometimes broadly concave at the anterior margin. The most conspicuous effect of these mutants extends across the middle half of the eye: this roughly grained area contains no distinct ommatidia or interommatidial bristles and very little pigment, but it does include on the order of a dozen irregularly disposed bristles that are about three times the size of interommatidials.

Om(2E)9h. There is only a small reduction of eye size, seen as a slightly concave anterior margin, in this mutant. The ommatidia and their associated bristles tend to be disarrayed at this margin but more so in the central region of the eye. Om9h was discovered as a suppressor of Om(lD)9.

Om(2F)27a. Recessive lethality associated with this mutant has prevented ob- servations of homozygotes. The eyes of heterozygotes are of normal size and shape, but the ommatidia of the central region are disarrayed similarly to Om(2E)9h homozygotes.

Om(3A)43, 57. The primary phene of these mutants is a broad concavity of the anterior margin of the eye with an isthmus of rounded ommatidia extending to a smaller indentation of the posterior border. The anterior concavity is glazed and pigmented, but it contains no free interommatidial bristles which are also scarce in the isthmus.

Om(3B)20, 29, 55. Om 20 and Om29 eyes are shaped like commas with as few as 100 ommatidia in the dorsal mass. The more severely reduced, glazed and pigmented, ventral region contains very few or no distinct ommatidia. Interom- matidial bristles are restricted to the dorsal patch of ommatidia. Om55 is a very weak mutant displaying a slight anterior concavity bordered by irregularly arranged ommatidia.

Om(3C)6, 9a, 15a. Om6 and Om9a present a protruding mass of tissue located within a large unpigmented cleft which is extended posteriorly through the equator as a glazed and pigmented scar. The finely textured surface of the protruding tissue is marked by several bristles that vary in size between inter- ommatidials and smaller vibrissae. In the more extreme Om15a homozygotes, the ommatidia are restricted to the dorsoposterior quadrant of the eye field whose ventral boundary includes a more or less completely duplicated, but disordered, set of vibrissae. The extensive area between the ommatidia and the duplicated vibrissae is finely textured as in the protrusions of the less extreme mutants at this locus.

MORPHOGENETICALLY SPECIFIC MUTABILITY 643

Among the remaining 29 transmitted Om mutants that were assigned to linkage groups but not mapped, there are none with phenotypes markedly different from those just described for the mapped mutants. However, the specificity of their phenotypes, especially those of weaker mutants, is often not sufficient to assign an unmapped mutant to one or another of the identified loci with assurance; such ambiguity is expected on the basis of the extensive mimicry between mapped Om mutants assigned to different loci. Superficial mimicry may obscure locus-specific effects whose existence is patently demonstrated by the phenotypes of mutants representing the IC, IE, 20, 3B and 3C loci.

Casual inspection of homozygous Om mutant stocks at irregular intervals shows that they are generally true breeding, but certain incidental observations indicate that stability of Om mutants is not absolute. Within two generations of its homozygosis, the Om(1 D)9 stock produced at least four exceptional flies; two of these bred true as wild type in one case and in the other as Om(lD)9b (a consistently weaker allele than Om9) and apparently represented isolocus changes, whereas two others, identified by their extreme Om phenotypes, assorted the original Om9 and either one or the other of two new unlinked Om mutants, 0m(3C)9a and Om(2C)9c. Subsequently, another more extreme isolocus derivative, Om(lD)9e, was isolated from the Om9 stock, and Om(3C)15a was detected as a group of four females with extreme phenotypes in the Om(lA)15 stock. The extensive testcrosses of heterozygous females to map new Om mutants produced very few exceptional progeny, but they included a revertant of Om(2C)S (previ- ously described) and Om(2F)27a recovered as a single weak Om female which proved to be genotypically + pea Om/e pea +. Om(lE)59a was recovered as a single son among the progeny of an Om(2)59/+ female. These results prompted a more systematic examination of Om mutant stability.

Selected X-linked Om mutant stocks were assayed for stability in outcrosses to the cop f Bx w stock whose markers permitted identification and exclusion of nondisjunctional exceptions; only the daughters of outcrossed Om males were scored. Autosomal Om mutants were outcrossed to the px stock. The results of these assays (Table 5) show that females from the first 14 stocks listed produced five exceptions of which the wild-type derivative of Om5 and the weak derivatives of Om13 and Om1 7 all bred true as expected of complete or partial reversions of the parental mutants. On the other hand, tests of the two exceptions having more extreme Om phenotypes than their regular heterozygous siblings revealed the presence of the parental mutant and a new linked mutant, Om(2C)14a in one case and Om(2D)lOa in the other. The overall frequency of detected exceptions, 23.6/105, from mutant females is of the same magnitude as the frequency of original Om mutants recovered from the ca; px stock and, just as ca; px stock males exhibit no Om mutability, the progeny of assayed mutant males included no exceptions. Because the original Om mutants occurred in or were immediately backcrossed to the ca; px stock, the mutant stocks must share their genetic background and could be expected to exhibit recurrent Om mutability.

The data from stability assays of the Om(lD)9 stock are separated from the other results of Table 5 because this stock, as indicated by the casual observations already described, presents properties that are not readily explained by recurrent Om mutability. Many of the exceptional progeny of outcrossed Om(ID)9 parents

644 C. W. HINTON

TABLE 5

Stability of Om mutants assayed in FI outcross progenies scored for wild-type revertants (R) and weakly (W) or extremely (E) expressed exceptions

Progeny from tested females Progeny from tested males

stock Regular Exceptions Regular Exceptions Tested mutant

Om(lA)15 1,580 0 170 0 Om(lD)2 1,104 0 416 0 Om(ID)5 1,584 1 R 393 0 Om(1 D)I 6 955 0 1,027 0 Om( I D)36 1,231 0 177 0 Om(2A)I 3 780 1 w 764 0 Om(2a)I 7 1,330 1 w 2,882 0 Om(2B)I 4 2,376 1 E 2,522 0 Om(2C)l 927 0 900 0 Om(2C)7 842 0 1,029 0 Om(2C)S 2,818 0 2,087 0 Om(2C)lO 717 1 E 1,183 0 Om(2C)I 2 1,287 0 1,912 0 Om(3C)6 3,680 0 2,853 0 Sum 21,211 1 R, 2 W, 2 E 18,315 0 Om(ID)9 3,371 1 1 R, 3 W, 2 E 3,965 6 R , 4 W

occurred as apparent or, in some cases, obvious clusters; consequently, all phenotypically equivalent exceptions found in the same mating vial were assumed to have arisen from a single event, and the minimum number of events is recorded in Table 5 . Om9 males produced nearly as many exceptions as Om9 females, and the minimum frequency of exceptions is significantly higher than for the other assayed mutants. Analyses of the two extreme exceptions led to the identification of Om(2C)9f and Om(ID)9g (accompanied by a linked singed mu- tant), and two complete revertants apparently involved changes at the I D locus because they bred true. On the other hand, tests of 15 independent partial or complete revertants resulted in recovery of the original Om9 mutant and diag- nosis of dominant suppressor mutants. In one case, the suppressor is itself an Om mutant, Om(2E)9h, which is expressed in double heterozygotes to the exclusion of Om9 expression. No phenotypes, other than effects on Om expression, have been detected among the other 14 suppressors all of which are X linked. Mapping these suppressors in Su Om9/cop f Bx w females identified at least two loci: one locus, represented by one partial and 11 complete suppressor alleles, is about 2.5 map units to the left of Om9 and the other locus, represented by one partial and one complete suppressor allele, is more than 35 map units to the left of cop. From preliminary observations, it is clear that the suppressors are effective against Om mutants at loci other than ID.

The generation of suppressor mutants in the Om(lD)9 stock is not intrinsic to the 1D locus because four other assayed 1D mutants produced between them only one isolocus revertant (Table 5 ) and because the previously described test to resolve the locus by recombination in heteroallelic Om'/f Om9 Bx w females also resulted in no Om+ exceptions. Failure of the marked f Om9 Bx w stock (constructed for use in the foregoing test) to produce revertants among more

MORPHOGENETICALLY SPECIFIC MUTABILITY 645

than 3000 progeny examined shows that homozygosity for Om9 itself is not a sufficient condition for suppressor mutation. Nor is it a necessary condition because two more dominant X-linked suppressors have been isolated recently, one from the Om(2A)18 stock and one from a testcrossed Om(2C)26/e pea female.

SPECULATIONS

A challenging interpretation of Om mutability relies on two major assumptions. Following BRITTEN and DAVIDSON (1969), the Om mutants are assumed to represent a set of structural genes whose temporally coordinated expression in prospective eye tissue depends upon shared control sequences. Second, the shared control sequences of these structural genes present specific targets for a transposable element whose insertion impairs their function.

The superficial phenotypic observations on Om mutants support only the most rudimentary inferences about their mode of action. The rule of semidominance is compatible with cis-acting control sequences, but the available information does not discriminate between hypomorphic and hypermorphic functions; if the control sequences include a receptor site for an activator protein, then impair- ment of that association would most likely result in reduced output by the structural gene. Presumably, the dorsoposterior excess of ommatidia seen in mutants at the Om(1E) locus reflects an altered activity that is in some way the reverse of the functions exposed by Om mutants at the other loci. The minimal pleiotropy and minimal reduction of optic field size observed among Om mutants suggest that the genes are expressed only in tissue that has already been deter- mined as prospective eye. The prevalent anterior-posterior polarity among Om mutant phenotypes could be related to the determination wave (POODRY, HALL and SUZUKI 1973) that passes vertically, from posterior to anterior, across the imaginal eye disc, and the commonly seen equatorial defects coincide with the dorsal and ventral eye compartments already established in the blastoderm (BAKER 1978). Duplicated structures, such as the vibrissae of Om(3C) mutants, may reflect developmental regulations similar to those elicited by various pertur- bations of the eye disc (BRYANT 1978).

If each site identified by Om mutants is occupied by a different structural gene, the extensive phenotypic mimicry between mutants at different sites would be the consequence of their disruption of the same developmental program. More positive support for the involvement of a set of structural genes in Om mutability is provided by the unique or special phenotypes associated with several of the loci. The existence of a set of structural genes implies that its members should be represented by mutants from sources independent of the Om mutability system. Unfortunately, the extant non-Om eye morphology mutants in D. anan- assue are too few and unsuited for validating this expectation. For example, Om(3B) mutants map to the same region as Star, but this recessive lethal mutant was lost several years ago. Puffed and Lobe, also recessive lethals, could be candidates for positions at Om(2A) through Om(2D), but their association with rearrangements precludes recombination tests. Amusing but uncritical compar- isons of Om mutants with the dominant eye morphology mutants representing 16 loci in D. melanogaster (LINDSLEY and GRELL 1968) reveal some probable

646 C. W. HINTON

homologies; for a most striking example, Om(1D) mutant phenotypes are reason- able facsimiles of the D. melanogaster Bar eye phenotype and, in both species, the loci fall within 0.4 map unit off; on its Bx side.

The 15 loci identified by recombination tests (Figure 1) provide a minimum estimate of the number liable to Om mutability. The independent mapping procedure would not have exposed closely linked loci such as those suggested by the nonconforming phenotypes of Om(lC)52 and Om(2C)9c. On the other hand, the substantial but hardly exhaustive attempts to resolve the I D and 2C loci through recombination in heteroallelic females were negative. It is also possible that some loci were never represented by mutants, but this possibility is minimized by the failure to find novel phenotypes among the 29 unmapped mutants. The distribution of the 63 mapped mutants gives the impression of near-saturation because 12 of the 15 loci are represented by more than one mutant; however, use of the Poisson expectation for the number of undetected loci is inappropriate because of the apparent “hot spots” at lA, lD, 2A and 2C. Taken together, these considerations are consistent with the conclusion that the number of loci subject to Om mutability is finite and small, perhaps no more than 20.

Instead of a set of some 20 structural genes sharing common control sequences, one might imagine that the information for eye morphogenesis is included within the transposable element itself as in the transport of the w and rst genes by TE of D. melanogaster (ISING and BLOCK 1981). Although the w allele in TE is mutable, mutation is not concomitant with transposition, and the insertion sites of TE, although nonrandom, are neither recurrent nor apparently limited in number. To account for Om mutability in terms of such a carrier element would require that the information for eye development be expressed differently in its native and new sites, that locus-specific Om phenotypes be attributed to special chromosomal environments adjacent to the insertions and that insertion site number restriction be based on some entirely undefined device.

In addition to TE, several other unstable, as well as stable, spontaneous mutants of D. melanogaster have been recently associated with mobile genetic elements at the molecular level (RUBIN 1983). Although most of the mutants so analyzed were selected because of their special phenotypic or genetic properties, they and the role of transposable P factors as mediators of wholesale mutability in hybrid dysgenesis (BRECLIANO and KIDWELL 1983) lead to the suspicion that most spontaneous mutation in Drosophila involves insertion of mobile middle repetitive DNA sequences. Thus, participation of a transposable element (symbolized tom) in Om mutability requires no extraordinary assumptions, although some of its inferred properties distinguish tom from the transposable elements described so far in D. melanogaster.

Om mutability depends primarily, if not exclusively, on homozygosity for the X chromosome from the ca; px stock. This observation could mean that i X- linked recessive “mobilizer” is required for transposition of tom, i t ‘t is sii der to suppose that tom itself is X linked and, as surmised for the 1 L t P fa4 tors (RUBIN, KIDWELL and BINGHAM 1982), codes its own “tran 3osase” which reaches an effective concentration only in oocytes of homozygote. The sufficient mini- mum of one tom element per X chromosome permits d. ect estimates of its

MORPHOGENETICALLY SPECIFIC MUTABILITY 64 7

transposition rate from the frequencies of Om mutants recovered from various matings. Such estimates are not immediately comparable to those for other eukaryotic transposable elements because the latter usually exist in multiple copies dispersed throughout the genome. However, the approximate transposi- tion rate of for TE present as a single copy in heterozygotes is nearly an order of magnitude higher than for tom homozygotes. Similarly, the L factor, a putative X-linked transposable element, induces very high frequencies of X-linked recessive lethals and rearrangements (LIM 1981). T o some extent such rate differences could reflect the larger number of targets available to the D. melan- ogaster transposable elements as compared to the limited number for tom, but the comparisons are also confounded by the typically clustered recovery of D. melanogaster transposition mutants.

There appears to be no precedent among eukaryotic transposable elements that control sequences associated with structural genes present specific targets for insertion, as proposed for Om mutability. The most suggestive evidence in support of this idea comes from yeast in which selected regulatory mutants at three loci all contain intact T y elements in the 5’-noncoding regions of these genes (ROEDER and FINK 1983). In D. melanogaster, molecular evidence from three independent mutants with P factor insertions at equivalent points suggests preferential location within the complex w gene (RUBIN 1983), and the distribu- tion of such preferred sites among loci could be the basis for observed locus- specific responses to P element mutagenesis (GOLUBOVSKY, IVANOV and GREEN, 1977; ENCELS and PRESTON 1981) or to tom insertions at Om loci.

The observations that either of two different extrachromosomally replicating elements stimulates Om mutability in tom heterozygotes to the level of homozy- gotes imply more about the nature of the extrachromosomal elements than about tom. This effect could be analogous to the support provided replication-deficient vertebrate retroviruses by replication-competent relatives (VARMUS 1983) or the mobilization of deleted P factors by intact P factors in the same genome (RUBIN, KIDWELL and BINCHAM 1982). Unlike retroviruses and P factors, the extrachro- mosomally replicating elements are not known to integrate into chromosomes, but an affinity for chromosomes is suggested by their functions as a suppressor of mutability and male crossing over (the pc element) or as a clastogenic mutator (the ca element). It may not stress credulity too much more to suggest that tom arose in the ca stock as a product of the mutator and persisted as a rare variant until it was accidentally fixed during construction of the ca; px stock; this would rationalize the shared transposase functions of tom and the Ca mutator. Coexisting intrachromosomal and extrachromosomal forms of the same mobile repetitive sequences have been demonstrated (FLAVELL and ISH-HOROWICZ 198 1) for members of the copia family in D. melanogaster. Copia sequences have been detected as insertions in several spontaneous mutants, and their molecular organization resembles that of retroviruses.

Why is Om mutability restricted to females? The observation that phenotypes of X-linked Om mutants are more extreme in homozygous females than in hemizygous males suggests that they do not respond effectively to the dosage compensation mechanism (LUCCHESI 1978), and this might apply to tom itself if

648 C. W. HINTON

dosage compensation occurs in the male germ line. It seems quite as likely that tom does not transpose in primary spermatocytes because it, along with the rest of the X chromosome in these cells, is sequestered in the heterochromatic state (LIFSCHYTZ and LINDSLEY 1972). If homozygous autosomal Om mutants are stable in males (see next paragraph), both explanations based on hemizygosity of tom are invalid. Another possibly relevant sexual difference is crossing over which occurs regularly in females and, in D. ananassae, at lower levels in males of certain genotypes. The lack of appropriate linked markers prevented measure- ment of crossing over in ca; px or F1 males carrying tom and assayed for Om mutability. However, F1 ca/+; px/pc males, known to carry a dominant enhancer of male crossing over, produced no Om mutants among 10,101 progeny (Table 3) and, similarly in a supplementary test, no Om mutants were detected among 17,071 progeny of F1 tom males in which 15% recombination was measured between third chromosome markers separated by at least 90 map units in females. These samples sizes are not large enough to exclude a proportionately low level of Om mutability in these males, but crossing over per se does not support Om mutability in males at the level seen in females. In the 2-R hybrid dysgenesis system of D. melanogaster (BREGLIANO and KIDWELL 1983), mutational events are restricted to females and, for the most part, to meiosis.

Altogether, nine examples of Om mutant instability were encountered in this study; five were ostensibly wild-type revertants, three expressed weaker Om phenotypes and one had a more extreme Om phenotype. These products of Om instability can be interpreted, following correlated genetic and molecular analyses of unstable spontaneous mutants in D. melanogaster (COLLINS and RUBIN 1982; LEVIS, COLLINS and RUBIN 1982; RUBIN, KIDWELL and BINGHAM 1982), as complete excisions or in situ rearrangements of tom insertions at the mutant locus. At least six, and possibly all nine, cases were derived from the maternal Om mutant, and five of the nine were recovered among the 24,603 progeny of outcrossed Om females (Table 5) . From the meager numbers indicating restric- tion to females and a frequency of 20.3/105, it appears that Om mutant instability is subject to the same factors that govern production of Om mutants in the ca; px stock and of recurrent Om mutants in Om mutant stocks.

Finally, when the matter of dominant suppressors of Om mutants is considered, it is of interest that all 19 spontaneous mutants, representing ten D. melanogaster loci and selected for sensitivity to a recessive suppressor (su-Hw), carry insertions of the mobile gypsy element (MODOLELL, BENDER and MESELSON 1983). In yeast, three unlinked recessive suppressors of T y insertion mutants also increase exci- sion rates of the insertions (ROEDER and FINK 1983), and similar behavior may underly the formally analyzed properties of the Spm system in corn (FEDOROFF 1983). Although they are not diagnostic, the existence of suppressors of Om mutants is consistent with the claim that Om mutation involves the insertion of a transposable element. The preliminary observations on Om suppressors are particularly enigmatic with respect to their premeiotic origin, at two loci, in high frequencies and in both sexes. This pattern of mutability resembles that of several mutable genes in D. melanogaster (GREEN 1976), and it is quite different from the pattern of Om mutability; on the other hand, the possibility that suppressor mutability is entirely independent of Om mutability seems remote.

MORPHOGENETICALLY SPECIFIC MUTABILITY 649

This study, supported by research grant GM-16536 from the National Institutes of Health, was conducted largely in the stimulating laboratory ofJOHN LUCCHFSI whose gracious toleration of a fly- pushing guest is sincerely appreciated.

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ENGELS, W. L. and C. R. PRESTON, 1981 IdentifyingPfactors inDrosophila by means of chromosome breakage hotspots. Cell 26: 421-428.

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HINTON, C. W. and J. E. DOWNS, 1975 The mitotic, polytene and meiotic chromosomes ofDrosophila

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MORPHOGENETICALLY SPECIFIC MUTABILITY

42.6 30.4 41.2 36.6 41.1 39.7 43.7 40.2

65 1

25.8 13.9 24.6 23.0 26.3 22.5 27.3 24.5

APPENDIX

TABLE AI

Mapping results for X-linked Om mutants

Om23 263 Om46 402 Sum, B 665

Markers scored Mutant Progeny

no. scored Om sn Om sc Om cob m f Om Bx Om w Om

7.2 44.9 18.2 10.4 46.8 23.1 9.2 46.0 21.2

Om24 on225 Sum, A Om15 Om24 Om25 Om37 Om42 Om47 Om50

Om3 408 Om11 389 O n 5 2 679 Sum, C 1476

416 229 645 516 194 585 686 733 365 655

14.5 37.5 26.0 13.9 35.7 26.2 16.3 30.2 25.2 15.2 33.7 25.7

:::: 20.3

24.4 13.4 24.1 17.8 21.6 15.1 29.2

2.3 2.8 2.9 2.3 2.1 5.7 3.3 4.4

Sum, A 3734 I 21.9 I

16.8 14.9 21.0 20.8 18.3 23.0 20.4 16.1

Om9 4456 6.8 0.4 7.9 O d e 1282 5.9 0.4 7.7 Om9g 647 6.8 0.1 7.4 Sum, D 6385 6.6 0.4 7.8

7.7 10.1 7.6 8.2

Om2 571 Om5 509 Oin9 918 Om96 48 1 Om16 43 1 Om30 804 Om36 660 Om45 706

Om53,E 671

38.0 36.3 35.5 42.2 39.0 42.3 41.2 40.6

42.9 I 21.8

Om48 533 I Sum, D 5613 39.8

Boxed values (map units) used for consensus map (Figure 1).

65 2 C . W. HINTON

Om13 435 Om 17 781 Om 18 727 Om28 452 Om33 429 Om49 832 Sum, A 3656

TABLE A2

Mapping results for 2-linked Om mutants

9.4 8.5 6.1 7.8 7.0 2.9 7.5 9.1 5.1 2.1 7.2 8.2 7.0 6.5

Markers scored Mutant Progeny

no. scored Om ca e Om Om k a Om Om Pr Om

Om 14 582 0.9 Om22 442 1.8 Sum, B 1,024 , 1.3

Om 13

Om28 515 Sum, A 1,422

3.8 7.9 5.6

8.1 4.5 5.4 5.9 5.2 4.7 4.2 3.9 4.3 4.2 5.0 5.4 7.7 4.2 4.7 3.8 4.8

1.5 2.1 1.2 0.6 3.3 2.9 0.6 2.8 1.5 2.5 3.1 2.3 1.4 1.4 2.9 1.2 1.8

Om I 456 Om4 468 O s 7 576 Om8 758 Om9c 668 Oin9f 1,538 Om10 504 Om12 385 Om14a 3,242 Om 19 405 Om26 475 Om27 351 Om31 361 Om32 573 Om40 560 Om44 1,112 Sum, C 12,432

Om 1 Oa 1,244 Om21 377 Om63 650 I 6.3 I 2.0 Sum, D 2,271 6.2 [18

Boxed values (map units) used for consensus map (Figure 1).

MORPHOGENETICALLY SPECIFIC MUTABILITY

Om43 51 1 28.2 25.8 Om57 906 29.7 34.1 Sum, A 1417 29.1 31.1

Om20 697 18.2 7.6 Om29 776 14.2 9.7 O n 5 5 1145 21.3 7.2 Sum, B 2618 18.4 8.0

Om6 53 1 34.1 Om9a 443 31.6 Om 15a 628 30.1 Sum, C 1602 31.9

653

17.2

18.4

20.1

20.0 19.2 5.2 14.6 4.6 17.7 4.9