1 / 21

Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale

Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale. DeRisi, Iyer, and Brown (1997) Science 278 , 680-686. Introduction to Yeast. Free-living fungus; generally single-celled Eukaryotic; possesses a nucleus and other intracellular organelles

lester-shaw
Download Presentation

Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. Exploring the Metabolic and Genetic Control of Gene Expression on a Genomic Scale DeRisi, Iyer, and Brown (1997) Science278, 680-686

  2. 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)

  3. 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 D symbol (i.e. his3D)

  4. 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)

  5. Questions 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?

  6. DeRisi, Iyer, and Brown Experiment: Diauxic shift • Metabolism of glucose v. ethanol • Inoculate 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

  7. Diauxic Shift: Experiment Parameters Figure 5

  8. One microarray, this one compares the first two time points, after initial inoculation (green), then 9.5 h later (red) Figure 1

  9. 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 Figure 2: Repeated views of box outlined in Figure 1a

  10. Changes in metabolism as diauxic shift proceeds; red genes are the ones turned on; green ones are the ones turned off Figure 3

  11. Group behavior: Genes that work coordinately are regulated coordinately Figure 4

  12. 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 Figure 5

  13. Ribosomal protein genes, all decline b/c of loss of Rap1 mRNA. Rap1 encodes a transcription factor required for synthesis of these genes Figure 5

  14. Other uses of microarrays explored here • Determine effects of loss of a common transcription factor • TUP1 encodes a transcriptional co-repressor that works with Mig1p • Compare TUP1 cells to tup1D cells • TUP1 cDNA = green; tup1D cDNA = red; • red spots therefore genes suppressed by Tup1p

  15. 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 Figure 2 (bottom center panel)

  16. Logic of the Yeast Metabolic Cycle: Temporal Compartmentalization of Cellular Processes Tu et al., (2005) Science 310, 1152

  17. Oxygen consumption varies in a periodic manner: What genes are expressed in a periodic manner? Figure 1

  18. Correlation of gene expression with O2 consumption Figure 2

  19. Mitochondrial large ribosomal subunit protein Fatty acyl CoA oxidase Adenine deaminase Figure 2

  20. Expression of Genes Note that energy and metabolism protein genes are more likely to be periodic than others Table 1

  21. Genes can be arranged in superclusters Figure 3D

More Related