Structure

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OB Fold Contributes to Telomere Maintenance

Marita Cohn1,*1Department of Biology, Genetics Group, Lund University, Solvegatan 35, SE-223 62 Lund, Sweden*Correspondence: marita.cohn@biol.lu.sehttp://dx.doi.org/10.1016/j.str.2012.12.005

The essential Cdc13 protein is part of the trimeric CST complex that confers genome stability by binding toand protecting yeast telomeres. In this issue of Structure, Mason and colleagues characterize an OB folddomain of Cdc13 (named OB2) and propose that homo-dimerization of OB2 is required for proper assemblyof the CST complex and telomere maintenance.

DNA ends are recognized by recombina-

tion proteins and elicit a signal for activa-

tion of the DNA damage repair pathway.

Therefore, maintenance of chromosomal

integrity in eukaryotes is dependent on

the assembly of specialized nucleoprotein

structures, telomeres at the ends of the

linear chromosomes. The telomere-asso-

ciated Cdc13 protein is essential for the

maintenance of telomeres in budding

yeast and plays a crucial role in the regu-

lation of the elongation of the telomeres

by telomerase. However, the molecular

mechanisms underlying the multiple func-

tions of Cdc13 have not yet been fully

elucidated.

The Cdc13 protein binds to the single-

stranded telomeric 30 overhang and asso-

ciates with the Stn1 and Ten1 proteins

to form the trimeric Cdc13, Stn1, Ten1

(CST) complex that is critical for proper

telomere capping and also negatively

regulates telomerase access to the DNA

30-end. Accordingly, a CST-like complex

has been shown to be widely conserved

in evolution and is implicated in the regu-

lation of theC strand synthesis in the repli-

cation of telomeres. However, when not

associated to Stn1 and Ten1, Cdc13

performs an apparently opposing func-

tion in the recruitment of telomerase to

facilitate telomere elongation (Pennock

et al., 2001). This is crucial for maintained

cell division, because if unextended,

telomeres will shorten in every cell cycle,

eventually leading to senescence and

cell death.

Cdc13 has some unusual structural

characteristics, as it contains multiple

OB-fold domains, a feature that is shared

by other telomere proteins binding single-

stranded DNA. OB folds are involved

in multiple functions, including nucleic

acid, oligosaccharide, and oligopeptide

binding, and were recently shown to

mediate Cdc13 dimerization (Sun et al.,

2011). The secondary structure of a

typical telomere protein OB fold is charac-

terized by a b-b-b-a-b-b pattern, which

forms a partially closed barrel-like struc-

ture with two orthogonally packed anti-

parallel b sheets, a so-called b barrel.

However, OB proteins seem especially

prone to divergent evolution, and the

plasticity in the structure has been a

challenge for their discovery. Recent

research indicates an unexpected and

highly interesting functional versatility of

the OB fold. Thus, solving structures of

additional OB-fold domains and pinpoint-

ing separate molecular functions of the

respective fold is of high relevance to the

telomere biology field and to science in

general.

In this issue of Structure, Mason

et al. (2013) describe the structural and

functional analysis of a region in the

S. cerevisiae Cdc13 protein that was

not previously characterized, localized in

between the recruitment domain and

the DNA-binding domain (Figure 1). The

protein crystal structure revealed an OB

fold (named OB2), which they demon-

strate is involved in homo-dimerization.

This is the fourth OB fold to be discovered

in S. cerevisiae Cdc13, and the previously

determined OB folds have been shown to

function in homo-dimerization (OB1), DNA

binding (OB1 and OB3), and protein

binding (OB1 and OB4) (Hughes et al.,

2000; Qi and Zakian, 2000; Mitchell

et al., 2010; Sun et al., 2011). Mason

et al. (2013) make site-directed mutagen-

esis of residues, indicated by the struc-

ture to be of importance for the establish-

ment of the OB2 protein-protein contacts,

and report the finding that some double

mutants lead to increased telomere length

and decreased viability at elevated tem-

peratures. Moreover, they cause a drastic

Structure 21, January 8, 20

decrease in the ability of the isolated OB2

to dimerize. This leads to the conclusion

that Cdc13 contains two separate OB

folds that are involved in the homo-dimer-

ization (OB1 and OB2). However, in con-

trast to previous findings on OB1, where

mutations in contacting residues abol-

ished the dimerization of the full-length

protein, disruption of the dimerization

interactions in OB2 does not lead to loss

of the dimerization capacity of the full-

length Cdc13 protein (Sun et al., 2011).

On the other hand, using isothermal titra-

tion calorimetry assays, they show that

the interaction of Cdc13 with Stn1 is

abolished when residues important for

the OB2 dimerization are mutated. Thus,

even though the isolated OB2 peptide

does not bind to Stn1 on its own, the

dimerization of the OB2 domains of two

full-length proteins is necessary for the

establishment of the interaction between

Cdc13 and Stn1. Since Stn1 interaction

has previously been shown to involve

two quite distantly located regions within

Cdc13 (the RD and the CTD), the authors

postulate that the function of OB2 homo-

dimerization is to bring these distant

domains together to form an Stn1 inter-

acting surface.

These data contribute with essential

knowledge on the structure and func-

tion of a Cdc13 domain that was previ-

ously unknown and further our under-

standing of the molecular mechanisms

for telomere regulation. However, since

the separate OB folds contribute with

both distinct and apparently overlapping

functions, solving the structure of the

full-length Cdc13 protein will be neces-

sary for further clarification of the domain

organization and full understanding of

the detailed intricate mechanisms. Sig-

nificantly, these new findings bring up

questions regarding the oligomericity of

13 ª2013 Elsevier Ltd All rights reserved 3

OB1 OB4OB3OB2RDN C1 190 340 497 693 924

Pol1(weak ssDNA binding)

Est1

Homo-dimeriza�on

ssDNA BindingDomain

Stn1

Homo-dimeriza�on

Figure 1. The Domain Organization of S. cerevisiae Cdc13The multiple and differential functions contributed by the OB fold structures(OB1–4) are indicated. Both OB1 and OB2 are involved in homo-dimerization,but in addition, OB1 interacts with the Pol1 subunit of the DNA polymerase a.OB3 confines the DNA binding domain that mediates the major high affinityinteraction to the single-stranded telomeric 30 overhang, whereas OB1displays a weak DNA interaction. The recruitment domain (RD) interactswith Est1 to recruit the telomerase enzyme, while OB4 interacts with Stn1 toform the CST complex that is essential for telomere end protection and alsonegatively regulates telomerase access to the DNA 30 end.

Structure

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the other subcomponents of

the CST complex. Further-

more, recent data indicate

that Cdc13 contains many

residues that are phos-

phorylated, and significantly,

SUMOylation has been

shown to promote its interac-

tion with Stn1 (Li et al., 2009;

Tseng et al., 2009; Hang

et al., 2011; Wu et al.,

2012). Thus, future studies of

Cdc13 should inevitably in-

clude the investigation of

whether such posttransla-

tional modifications are in-

volved in the regulation of

the dimerization of this spe-

cific OB2 domain, thereby providing

a way to regulate the assembly of the

CST complex. Distinct modifications

may provide the cue for conformational

changes, orchestrating the separate roles

of Cdc13 in the essential balancing act

needed for maintaining telomere length

homeostasis.

4 Structure 21, January 8, 2013 ª2013 Elsevi

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Hughes, T.R., Weilbaecher, R.G., Walterscheid,M., and Lundblad, V. (2000). Proc. Natl. Acad.Sci. USA 97, 6457–6462.

er Ltd All rights reserved

Li, S., Makovets, S., Matsuguchi, T.,Blethrow, J.D., Shokat, K.M., andBlackburn, E.H. (2009). Cell 136,50–61.

Mason, M., Wanat, J.J., Harper, S.,Schultz, D.C., Speicher, D.W., John-son, F.B., and Skordalakes, E.(2013). Structure 21, this issue,109–120.

Mitchell, M.T., Smith, J.S., Mason,M., Harper, S., Speicher, D.W.,Johnson, F.B., and Skordalakes, E.(2010). Mol. Cell. Biol. 30, 5325–5334.

Pennock, E., Buckley, K., and Lund-blad, V. (2001). Cell 104, 387–396.

Qi, H., and Zakian, V.A. (2000).Genes Dev. 14, 1777–1788.

Sun, J., Yang, Y., Wan, K., Mao, N.,Yu, T.Y., Lin, Y.C., DeZwaan, D.C.,

Freeman, B.C., Lin, J.J., Lue, N.F., and Lei, M.(2011). Cell Res. 21, 258–274.

Tseng, S.F., Shen, Z.J., Tsai, H.J., Lin, Y.H., andTeng, S.C. (2009). Nucleic Acids Res. 37, 3602–3611.

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Single-Stranded Nucleic Acid Recognition: Is Therea Code after All?

Antoine Clery,1 Julien Boudet,1 and Frederic H.-T. Allain1,*1Institute of Molecular Biology and Biophysics, ETH Zurich, Switzerland*Correspondence: allain@mol.biol.ethz.chhttp://dx.doi.org/10.1016/j.str.2012.12.006

Proteins that bind single-stranded nucleic acids have crucial roles in cells, and structural analyses havecontributed to a better understanding of their functions. In this issue of Structure, Dickey and colleaguesdescribe several high resolution structures of a single OB-fold bound to different single-stranded DNA(ssDNA) sequences and reveal a spectacular co-adaptability of the protein/ssDNA interactions.

Proteins that bind single-stranded (ss) nu-

cleic acid molecules (ssDNA or ssRNA)

have crucial roles in the three domains

of life: bacteria, archaea, and eukarya.

ssRNA binding proteins are involved at

each step of the mRNA journey in cells.

Some remain bound to the transcribed

RNA until it is degraded, whereas others

transiently bind to RNA at different stages

of specific processes such as processing

(e.g., splicing), nuclear export, and trans-

lation (Dreyfuss et al., 2002). Some RNA

binding proteins also function as RNA

chaperones by helping the RNA, which

is initially single-stranded, to form various

secondary or tertiary structures. ssDNA

binding proteins are involved in DNA

metabolism steps that require manipula-

tion of DNA in its single-stranded form.

They are involved in telomere-ends

maintenance, DNA replication, and DNA

recombination and repair (Broderick

et al., 2010; Richard et al., 2009). In

most cases, these proteins act by pro-

tecting ssDNA from nucleases, prevent-

ing the formation of secondary structures

and/or promoting the recruitment of addi-

tional factors on targeted ssDNAs.

Several of these factors do not have

redundant functions, and therefore they

need to act on a specific target at a certain

time. In addition to their tightly regulated

production in cells, their specificity of