Biology:Gal4 transcription factor

From HandWiki
Regulatory protein GAL4
Identifiers
OrganismSaccharomyces cerevisiae
SymbolGAL4
Entrez855828
UniProtP04386

The Gal4 transcription factor is a positive regulator of gene expression of galactose-induced genes.[1] This protein represents a large fungal family of transcription factors, Gal4 family, which includes over 50 members in the yeast Saccharomyces cerevisiae e.g. Oaf1, Pip2, Pdr1, Pdr3, Leu3.[2]

Gal4 recognizes genes with UASG, an upstream activating sequence, and activates them. In yeast cells, the principal targets are GAL1 (galactokinase), GAL10 (UDP-glucose 4-epimerase), and GAL7 (galactose-1-phosphate uridylyltransferase), three enzymes required for galactose metabolism. This binding has also proven useful in constructing the GAL4/UAS system, a technique for controlling expression in insects.[3] In yeast, Gal4 is by default repressed by Gal80, and activated in the presence of galactose as Gal3 binds away Gal80.[4]

Domains

Two executive domains, DNA binding and activation domains, provide key function of the Gal4 protein conforming to most of the transcription factors.

Gal4 domains and regulation

DNA binding

Gal4 N-terminus is a zinc finger and belongs to the Zn(2)-C6 fungal family. It forms a Zn – cysteines thiolate cluster,[5][6] and specifically recognizes UASG in GAL1 promoter. [7][8]

Gal4 transactivation

Localised to the C-terminus, belongs to the nine amino acids transactivation domain family, 9aaTAD, together with Oaf1, Pip2, Pdr1, Pdr3, but also p53, E2A, MLL.[9][10]

Regulation

Galactose induces Gal4 mediated transcription albeit Glucose causes severe repression.[11][12]

As a part of the Gal4 regulation, inhibitory protein Gal80 recognises and binds to the Gal4 region (853-874 aa).[13][14][15]

The inhibitory protein Gal80 is sequestered by regulatory protein Gal3 in Galactose dependent manner. This allows for Gal4 to work when there is galactose.[16][4][17][18]

Mutants

The Gal4 loss-of-function mutant gal4-64 (1-852 aa, deletion of the Gal4 C-terminal 29 aa) lost both interaction with Gal80 and activation function.[19][20][21]

In the Gal4 reverted mutant Gal4C-62 mutant,[22] a sequence (QTAY N AFMN) with the 9aaTAD pattern emerged and restored activation function of the Gal4 protein.

Inactive constructs

The activation domain Gal4 is inhibited by C-terminal domain in some Gal4 constructs.[23][24]

Function

Target

Transcription

The Gal4 activation function is mediated by MED15 (Gal11).[25][26][27][28][29][30][31]

The Gal4 protein interacts also with other mediators of transcription as are Tra1,[32][33][34] TAF9,[35] and SAGA/MED15 complex.[36][37]

Proteosome

A subunit of the 26 S proteasome Sug2 regulatory protein has a molecular and functional interaction with Gal4 function.[38][39] Proteolytic turnover of the Gal4 transcription factor is not required for function in vivo.[40] The native Gal4 monoubiquitination protects from 19S-mediated destabilizing under inducing conditions.[41]

Application

The broad use of the Gal4 is in yeast two-hybrid screening to screen or to assay protein-protein interactions in eukaryotic cells from yeast to human.

In the GAL4/UAS system, the Gal4 protein and Gal4 upstream activating region (UAS) are used to study the gene expression and function in organisms such as the fruit fly.[3]

The Gal4 and inhibitory protein Gal80 have found application in a genetics technique for creating individually labeled homozygous cells called MARCM (Mosaic analysis with a repressible cell marker).

See also

References

  1. ↑ "Studies on the positive regulatory gene, GAL4, in regulation of galactose catabolic enzymes in Saccharomyces cerevisiae". Molecular & General Genetics 135 (3): 203–12. 1974. doi:10.1007/BF00268616. PMID 4376212. 
  2. ↑ "Comparative amino acid sequence analysis of the C6 zinc cluster family of transcriptional regulators". Nucleic Acids Research 24 (23): 4599–607. December 1996. doi:10.1093/nar/24.23.4599. PMID 8967907. 
  3. ↑ 3.0 3.1 "GAL4 system in Drosophila: a fly geneticist's Swiss army knife". Genesis 34 (1–2): 1–15. 2002. doi:10.1002/gene.10150. PMID 12324939. 
  4. ↑ 4.0 4.1 "Gene activation by dissociation of an inhibitor from a transcriptional activation domain". Molecular and Cellular Biology 29 (20): 5604–10. October 2009. doi:10.1128/MCB.00632-09. PMID 19651897. 
  5. ↑ "DNA recognition by GAL4: structure of a protein-DNA complex". Nature 356 (6368): 408–14. April 1992. doi:10.1038/356408a0. PMID 1557122. Bibcode: 1992Natur.356..408M. 
  6. ↑ "The DNA binding domain of GAL4 forms a binuclear metal ion complex". Biochemistry 29 (12): 2023–9. March 1990. doi:10.1021/bi00464a019. PMID 2186803. 
  7. ↑ "Separation of DNA binding from the transcription-activating function of a eukaryotic regulatory protein". Science 231 (4739): 699–704. February 1986. doi:10.1126/science.3080805. PMID 3080805. Bibcode: 1986Sci...231..699K. 
  8. ↑ "Specific DNA binding of GAL4, a positive regulatory protein of yeast". Cell 40 (4): 767–74. April 1985. doi:10.1016/0092-8674(85)90336-8. PMID 3886158. 
  9. ↑ "The DNA binding and activation domains of Gal4p are sufficient for conveying its regulatory signals". Molecular and Cellular Biology 17 (5): 2538–49. May 1997. doi:10.1128/MCB.17.5.2538. PMID 9111323. 
  10. ↑ "GAL4 interacts with TATA-binding protein and coactivators". Molecular and Cellular Biology 15 (5): 2839–48. May 1995. doi:10.1128/MCB.15.5.2839. PMID 7739564. 
  11. ↑ "Studies on the positive regulatory gene, GAL4, in regulation of galactose catabolic enzymes in Saccharomyces cerevisiae". Molecular & General Genetics 135 (3): 203–12. 1974. doi:10.1007/BF00268616. PMID 4376212. 
  12. ↑ "Regulated expression of the GAL4 activator gene in yeast provides a sensitive genetic switch for glucose repression". Proceedings of the National Academy of Sciences of the United States of America 88 (19): 8597–601. October 1991. doi:10.1073/pnas.88.19.8597. PMID 1924319. Bibcode: 1991PNAS...88.8597G. 
  13. ↑ "NADP regulates the yeast GAL induction system". Science 319 (5866): 1090–2. February 2008. doi:10.1126/science.1151903. PMID 18292341. Bibcode: 2008Sci...319.1090K. 
  14. ↑ "The interaction between an acidic transcriptional activator and its inhibitor. The molecular basis of Gal4p recognition by Gal80p". The Journal of Biological Chemistry 283 (44): 30266–72. October 2008. doi:10.1074/jbc.M805200200. PMID 18701455. 
  15. ↑ "Interaction of positive and negative regulatory proteins in the galactose regulon of yeast". Cell 50 (1): 143–6. July 1987. doi:10.1016/0092-8674(87)90671-4. PMID 3297350. 
  16. ↑ "Rapid GAL gene switch of Saccharomyces cerevisiae depends on nuclear Gal3, not nucleocytoplasmic trafficking of Gal3 and Gal80". Genetics 189 (3): 825–36. November 2011. doi:10.1534/genetics.111.131839. PMID 21890741. 
  17. ↑ "Gene activation by interaction of an inhibitor with a cytoplasmic signaling protein". Proceedings of the National Academy of Sciences of the United States of America 99 (13): 8548–53. June 2002. doi:10.1073/pnas.142100099. PMID 12084916. Bibcode: 2002PNAS...99.8548P. 
  18. ↑ "Dilution kinetic studies of yeast populations: in vivo aggregation of galactose utilizing enzymes and positive regulator molecules". Genetics 77 (3): 491–505. July 1974. doi:10.1093/genetics/77.3.491. PMID 4369925. 
  19. ↑ "The genetic control of galactose utilization in Saccharomyces". Journal of Bacteriology 68 (6): 662–70. December 1954. doi:10.1128/jb.68.6.662-670.1954. PMID 13221541. 
  20. ↑ "Enzymatic Expression and Genetic Linkage of Genes Controlling Galactose Utilization in Saccharomyces". Genetics 49 (5): 837–44. May 1964. doi:10.1093/genetics/49.5.837. PMID 14158615. 
  21. ↑ "Function of positive regulatory gene gal4 in the synthesis of galactose pathway enzymes in Saccharomyces cerevisiae: evidence that the GAL81 region codes for part of the gal4 protein". Journal of Bacteriology 141 (2): 508–27. February 1980. doi:10.1128/JB.141.2.508-527.1980. PMID 6988385. 
  22. ↑ "Interaction of positive and negative regulatory proteins in the galactose regulon of yeast". Cell 50 (1): 143–6. July 1987. doi:10.1016/0092-8674(87)90671-4. PMID 3297350. 
  23. ↑ "Deletion analysis of GAL4 defines two transcriptional activating segments". Cell 48 (5): 847–53. March 1987. doi:10.1016/0092-8674(87)90081-X. PMID 3028647. 
  24. ↑ "A sequence-specific transcription activator motif and powerful synthetic variants that bind Mediator using a fuzzy protein interface". Proceedings of the National Academy of Sciences of the United States of America 111 (34): E3506-13. August 2014. doi:10.1073/pnas.1412088111. PMID 25122681. Bibcode: 2014PNAS..111E3506W. 
  25. ↑ "The Saccharomyces cerevisiae SPT13/GAL11 gene has both positive and negative regulatory roles in transcription". Molecular and Cellular Biology 9 (12): 5602–9. December 1989. doi:10.1128/MCB.9.12.5602. PMID 2685570. 
  26. ↑ "Peptides selected to bind the Gal80 repressor are potent transcriptional activation domains in yeast". The Journal of Biological Chemistry 275 (20): 14979–84. May 2000. doi:10.1074/jbc.275.20.14979. PMID 10809742. 
  27. ↑ "Regulation of expression of the galactose gene cluster in Saccharomyces cerevisiae. Isolation and characterization of the regulatory gene GAL4". Molecular & General Genetics 191 (1): 31–8. 1983. doi:10.1007/BF00330886. PMID 6350827. 
  28. ↑ "GAL11 (SPT13), a transcriptional regulator of diverse yeast genes, affects the phosphorylation state of GAL4, a highly specific transcriptional activator". Molecular and Cellular Biology 11 (4): 2311–4. April 1991. doi:10.1128/MCB.11.4.2311. PMID 2005915. 
  29. ↑ "A novel mutation that affects utilization of galactose in Saccharomyces cerevisiae". Current Genetics 2 (2): 115–20. October 1980. doi:10.1007/BF00420623. PMID 24189802. 
  30. ↑ "Yeast GAL11 protein is a distinctive type transcription factor that enhances basal transcription in vitro". Proceedings of the National Academy of Sciences of the United States of America 90 (18): 8382–6. September 1993. doi:10.1073/pnas.90.18.8382. PMID 8378310. Bibcode: 1993PNAS...90.8382S. 
  31. ↑ "GAL11 protein, an auxiliary transcription activator for genes encoding galactose-metabolizing enzymes in Saccharomyces cerevisiae". Molecular and Cellular Biology 12 (10): 4806. October 1992. doi:10.1128/MCB.12.10.4806. PMID 1406662. 
  32. ↑ "Analysis of Gal4-directed transcription activation using Tra1 mutants selectively defective for interaction with Gal4". Proceedings of the National Academy of Sciences of the United States of America 109 (6): 1997–2002. February 2012. doi:10.1073/pnas.1116340109. PMID 22308403. PMC 3277556. Bibcode: 2012PNAS..109.1997L. http://escholarship.umassmed.edu/cgi/viewcontent.cgi?article=1180&context=pgfe_pp. 
  33. ↑ "Tra1 as a screening target for transcriptional activation domain discovery". Bioorganic & Medicinal Chemistry Letters 19 (14): 3733–5. July 2009. doi:10.1016/j.bmcl.2009.05.045. PMID 19497740. 
  34. ↑ "Targets of the Gal4 transcription activator in functional transcription complexes". Molecular and Cellular Biology 25 (20): 9092–102. October 2005. doi:10.1128/MCB.25.20.9092-9102.2005. PMID 16199885. 
  35. ↑ "Use of a genetically introduced cross-linker to identify interaction sites of acidic activators within native transcription factor IID and SAGA". The Journal of Biological Chemistry 278 (9): 6779–86. February 2003. doi:10.1074/jbc.M212514200. PMID 12501245. 
  36. ↑ "The Saccharomyces cerevisiae Srb8-Srb11 complex functions with the SAGA complex during Gal4-activated transcription". Molecular and Cellular Biology 25 (1): 114–23. January 2005. doi:10.1128/MCB.25.1.114-123.2005. PMID 15601835.  (http://mcb.asm.org/content/25/1/114/F8.large.jpg)
  37. ↑ "Functional studies of the yeast med5, med15 and med16 mediator tail subunits". PLOS ONE 8 (8): e73137. 2013. doi:10.1371/journal.pone.0073137. PMID 23991176. Bibcode: 2013PLoSO...873137L. 
  38. ↑ "The Gal4 activation domain binds Sug2 protein, a proteasome component, in vivo and in vitro". The Journal of Biological Chemistry 276 (33): 30956–63. August 2001. doi:10.1074/jbc.M102254200. PMID 11418596. 
  39. ↑ "Evidence that proteolysis of Gal4 cannot explain the transcriptional effects of proteasome ATPase mutations". The Journal of Biological Chemistry 276 (13): 9825–31. March 2001. doi:10.1074/jbc.M010889200. PMID 11152478. 
  40. ↑ "Proteolytic turnover of the Gal4 transcription factor is not required for function in vivo". Nature 442 (7106): 1054–7. August 2006. doi:10.1038/nature05067. PMID 16929306. Bibcode: 2006Natur.442.1054N. 
  41. ↑ "The role of the proteasomal ATPases and activator monoubiquitylation in regulating Gal4 binding to promoters". Genes & Development 21 (1): 112–23. January 2007. doi:10.1101/gad.1493207. PMID 17167105. 

Further reading



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