Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale
description
Transcript of Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale
![Page 1: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/1.jpg)
Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale
DeRisi, Iyer, and Brown (1997) Science 278, 680-686
![Page 2: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/2.jpg)
Introduction to Yeast
Free-living fungus; generally single-celled Eukaryotic; possesses a nucleus and other
intracellular organelles Grows by budding; size of bud reflects
progression through the cell cycle Can exist either as a haploid (1N) or diploid
(2N); haploids can be mated to form new diploids; diploids can be sporulated to make new haploids
First eukaryotic genome to be completely sequenced (1996)
![Page 3: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/3.jpg)
Yeast Genetic Nomenclature
All named genes have a three-letter abbreviation followed by a number
Wild type genes are denoted as italicized capital letters (i.e. ACT1)
Mutant alleles (which are usually recessive) are denoted by small-case letters, followed by an allele number (i.e. ura3-52); dominant alleles are denoted by capital letters
Frank deletions are usually followed by the symbol (i.e. his3)
![Page 4: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/4.jpg)
More Yeast Genetic Nomenclature
Not all genes in the genome have been named Such loci are therefore referred to by their chromosome location
The first space is always a Y The second space is a letter referring to the chromosome
number (A = chromosome I, B = II, etc.) The third space is either R or L, and refers to whether the locus is
to the Right or Left of the centromere as the chromosome is conventionally drawn (long arm is Left arm)
A three digit number follows; this is the locus number on that arm The last space is either a W or a C; referring to either the Watson
or Crick strand (top or bottom respectively as conventionally drawn)
![Page 5: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/5.jpg)
1. Have all the genes required for a particular process (in this case, global carbon utilization pathways) been identified?
2. How do such genes change in expression over time?
3. Are the genes involved in a particular process coordinately regulated?
Questions
![Page 6: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/6.jpg)
DeRisi, Iyer, and Brown Experiment: Diauxic shift
Metabolism of glucose v. ethanolInoculate culture; soon thereafter
isolate cells and make RNA, from this create cDNA labeled with Cy3-dUTP
At ~9h post-inoculation and every 2h thereafter, isolate cells and make RNA, from these create cDNA labeled with Cy5-dUTP
![Page 7: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/7.jpg)
Diauxic Shift: Experiment Parameters
Figure 5
![Page 8: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/8.jpg)
Figure
One microarray, this one compares the first two time points, after initial inoculation (green), then 9.5 h later (red)
![Page 9: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/9.jpg)
Figure 2: Repeated views of box outlined in Figure 1a
Results: As time proceeds, more and more differences are observed
By the last time point:710 genes induced at least 2x (183 genes at least 4x)
1030 genes repressed at least 2x (203 genes at least 4x)
>870 of these previously unknown to be associated with this process
![Page 10: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/10.jpg)
Figure 3
Changes in metabolism as diauxic shift proceeds; red genes are the ones turned on; green ones are the ones turned off
![Page 11: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/11.jpg)
Figure 4
Group behavior: Genes that work coordinately are regulated coordinately
![Page 12: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/12.jpg)
Figure 5
Genes whose expression increases markedly, but only in the last time point; most have a glucose-repressible carbon source response element (CSRE) in their promoters
![Page 13: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/13.jpg)
Figure 5
Ribosomal protein genes, all decline b/c of loss of Rap1 mRNA. Rap1 encodes a transcription factor required for synthesis of these genes
![Page 14: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/14.jpg)
Other uses of microarrays explored here
Determine effects of loss of a common transcription factorTUP1 encodes a transcriptional co-
repressor that works with Mig1pCompare TUP1 cells to tup1 cellsTUP1 cDNA = green; tup1 cDNA =
red; red spots therefore genes suppressed
by Tup1p
![Page 15: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/15.jpg)
Figure 2 (bottom center panel)
Many (10%) of the same genes induced by diauxic shift are also induced by the absence of Tup1p; suggesting Tup1p is important for diauxic shift
![Page 16: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/16.jpg)
Logic of the Yeast Metabolic Cycle: Temporal Compartmentalization of Cellular Processes
Tu et al., (2005) Science 310, 1152
![Page 17: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/17.jpg)
Figure 1
Oxygen consumption varies in a periodic manner: What genes are expressed in a periodic manner?
![Page 18: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/18.jpg)
Figure 2
Correlation of gene expression with O2 consumption
![Page 19: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/19.jpg)
Figure 2
Mitochondrial large ribosomal subunit protein
Fatty acyl CoA oxidase
Adenine deaminase
![Page 20: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/20.jpg)
Expression of Genes
Table 1
Note that energy and metabolism protein genes are more likely to be periodic than others
![Page 21: Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale](https://reader035.fdocuments.us/reader035/viewer/2022062518/56813fc3550346895daaa312/html5/thumbnails/21.jpg)
Genes can be arranged in superclusters
Figure 3D