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Elizabeth Grayhack

TitleAssociate Professor
InstitutionSchool of Medicine and Dentistry
DepartmentBiochemistry and Biophysics
AddressUniversity of Rochester Medical Center
School of Medicine and Dentistry
601 Elmwood Ave, Box 712
Rochester NY 14642
Other Positions
TitleAssociate Professor
InstitutionUniversity of Rochester Medical Center
DepartmentCancer Center

TitleAssociate Professor
InstitutionSchool of Medicine and Dentistry
DepartmentPediatrics

 
 Overview
Functional Proteomics: The wealth of sequenced genomes has resulted in the development of powerful genomic approaches to define the functions of the individual genes, to deduce the networks of interactions between genes and to mine additional information encoded in the genome. For example, a collection of 5,854 sequence-verified yeast expression plasmids (constructed in collaboration with Eric Phizicky, Mark Dumont and Mike Snyder) has opened the door to study signals within genes that regulate their expression. We observe 1,000 fold differences in expression of genes in this collection, despite the fact that all ORFs are expressed using the same set of extragenic regulatory signals. Thus, the ORFs themselves or their protein products are esponsible for these large differences in expression. Analysis of this collection has led to the two projects described below.

The roles of codons in gene expression and regulation: Although accurate and efficient translation of mRNA into proteins is crucially important for the cell, not all of the 61 codons that specify insertion of amino acids into nascent polypeptides behave equally in this process. In fact, almost all synonymous codons, i.e. those that specify the same amino acid, are used at very different frequencies within most organisms, including the yeast S. cerevisiae Although codon use has been implicated as a determinant of expression levels, neither the identity, nor the defining characteristics of codons that impair expression are known. Studies in the Grayhack laboratory are designed to define the rules and mechanisms by which specific codons, and arrangements of codons, impair gene expression, and to identify regulatory models dictated by codon usage in the yeast genome.

Developing yeast to express protein complexes for structural analysis: The structures of many eukaryotic proteins and protein complexes are still unknown because many eukaryotic proteins are either not soluble in E. coli or bear post-translational modifications that are not added in E. coli. To this end we developed a general solution to incorporate selenomethionine into proteins expressed in this yeast, for use in MAD phasing. In addition, we have developed vectors for simultaneous high level expression and affinity purification of protein complexes from S. cerevisiae. Our current goal is to explore the limits of protein expression, solubility and folding to learn in part how yeast differs from E. coli, and what factors contribute most to gene expression.

 
 Selected Publications
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  1. Letzring DP, Wolf AS, Brule CE, Grayhack EJ. Translation of CGA codon repeats in yeast involves quality control components and ribosomal protein L1. RNA. 2013 Sep; 19(9):1208-17.
    View in: PubMed
  2. Grant TD, Luft JR, Wolfley JR, Snell ME, Tsuruta H, Corretore S, Quartley E, Phizicky EM, Grayhack EJ, Snell EH. The Structure of Yeast Glutaminyl-tRNA Synthetase and Modeling of Its Interaction with tRNA. J Mol Biol. 2013 Jul 24; 425(14):2480-93.
    View in: PubMed
  3. Dean KM, Grayhack EJ. RNA-ID, a highly sensitive and robust method to identify cis-regulatory sequences using superfolder GFP and a fluorescence-based assay. RNA. 2012 Dec; 18(12):2335-44.
    View in: PubMed
  4. Grant TD, Snell EH, Luft JR, Quartley E, Corretore S, Wolfley JR, Snell ME, Hadd A, Perona JJ, Phizicky EM, Grayhack EJ. Structural conservation of an ancient tRNA sensor in eukaryotic glutaminyl-tRNA synthetase. Nucleic Acids Res. 2012 Apr; 40(8):3723-31.
    View in: PubMed
  5. Letzring DP, Dean KM, Grayhack EJ. Control of translation efficiency in yeast by codon-anticodon interactions. RNA. 2010 Dec; 16(12):2516-28.
    View in: PubMed
  6. Quartley E, Alexandrov A, Mikucki M, Buckner FS, Hol WG, DeTitta GT, Phizicky EM, Grayhack EJ. Heterologous expression of L. major proteins in S. cerevisiae: a test of solubility, purity, and gene recoding. J Struct Funct Genomics. 2009 Sep; 10(3):233-47.
    View in: PubMed
  7. Chernyakov I, Whipple JM, Kotelawala L, Grayhack EJ, Phizicky EM. Degradation of several hypomodified mature tRNA species in Saccharomyces cerevisiae is mediated by Met22 and the 5'-3' exonucleases Rat1 and Xrn1. Genes Dev. 2008 May 15; 22(10):1369-80.
    View in: PubMed
  8. Chernyakov I, Baker MA, Grayhack EJ, Phizicky EM. Chapter 11. Identification and analysis of tRNAs that are degraded in Saccharomyces cerevisiae due to lack of modifications. Methods Enzymol. 2008; 449:221-37.
    View in: PubMed
  9. Jackman JE, Grayhack EJ, Phizicky EM. The use of Saccharomyces cerevisiae proteomic libraries to identify RNA-modifying proteins. Methods Mol Biol. 2008; 488:383-93.
    View in: PubMed
  10. Fan E, Baker D, Fields S, Gelb MH, Buckner FS, Van Voorhis WC, Phizicky E, Dumont M, Mehlin C, Grayhack E, Sullivan M, Verlinde C, Detitta G, Meldrum DR, Merritt EA, Earnest T, Soltis M, Zucker F, Myler PJ, Schoenfeld L, Kim D, Worthey L, Lacount D, Vignali M, Li J, Mondal S, Massey A, Carroll B, Gulde S, Luft J, Desoto L, Holl M, Caruthers J, Bosch J, Robien M, Arakaki T, Holmes M, Le Trong I, Hol WG. Structural genomics of pathogenic protozoa: an overview. Methods Mol Biol. 2008; 426:497-513.
    View in: PubMed
  11. Kotelawala L, Grayhack EJ, Phizicky EM. Identification of yeast tRNA Um(44) 2'-O-methyltransferase (Trm44) and demonstration of a Trm44 role in sustaining levels of specific tRNA(Ser) species. RNA. 2008 Jan; 14(1):158-69.
    View in: PubMed
  12. Malkowski MG, Quartley E, Friedman AE, Babulski J, Kon Y, Wolfley J, Said M, Luft JR, Phizicky EM, DeTitta GT, Grayhack EJ. Blocking S-adenosylmethionine synthesis in yeast allows selenomethionine incorporation and multiwavelength anomalous dispersion phasing. Proc Natl Acad Sci U S A. 2007 Apr 17; 104(16):6678-83.
    View in: PubMed
  13. Wilkinson ML, Crary SM, Jackman JE, Grayhack EJ, Phizicky EM. The 2'-O-methyltransferase responsible for modification of yeast tRNA at position 4. RNA. 2007 Mar; 13(3):404-13.
    View in: PubMed
  14. Jackman JE, Kotelawala L, Grayhack EJ, Phizicky EM. Identification and characterization of modification enzymes by biochemical analysis of the proteome. Methods Enzymol. 2007; 425:139-52.
    View in: PubMed
  15. White MA, Clark KM, Grayhack EJ, Dumont ME. Characteristics affecting expression and solubilization of yeast membrane proteins. J Mol Biol. 2007 Jan 19; 365(3):621-36.
    View in: PubMed
  16. Phizicky EM, Grayhack EJ. Proteome-scale analysis of biochemical activity. Crit Rev Biochem Mol Biol. 2006 Sep-Oct; 41(5):315-27.
    View in: PubMed
  17. Alexandrov A, Chernyakov I, Gu W, Hiley SL, Hughes TR, Grayhack EJ, Phizicky EM. Rapid tRNA decay can result from lack of nonessential modifications. Mol Cell. 2006 Jan 6; 21(1):87-96.
    View in: PubMed
  18. Gelperin DM, White MA, Wilkinson ML, Kon Y, Kung LA, Wise KJ, Lopez-Hoyo N, Jiang L, Piccirillo S, Yu H, Gerstein M, Dumont ME, Phizicky EM, Snyder M, Grayhack EJ. Biochemical and genetic analysis of the yeast proteome with a movable ORF collection. Genes Dev. 2005 Dec 1; 19(23):2816-26.
    View in: PubMed
  19. Gu W, Hurto RL, Hopper AK, Grayhack EJ, Phizicky EM. Depletion of Saccharomyces cerevisiae tRNA(His) guanylyltransferase Thg1p leads to uncharged tRNAHis with additional m(5)C. Mol Cell Biol. 2005 Sep; 25(18):8191-201.
    View in: PubMed
  20. Ma X, Yang C, Alexandrov A, Grayhack EJ, Behm-Ansmant I, Yu YT. Pseudouridylation of yeast U2 snRNA is catalyzed by either an RNA-guided or RNA-independent mechanism. EMBO J. 2005 Jul 6; 24(13):2403-13.
    View in: PubMed
  21. Alexandrov A, Grayhack EJ, Phizicky EM. tRNA m7G methyltransferase Trm8p/Trm82p: evidence linking activity to a growth phenotype and implicating Trm82p in maintaining levels of active Trm8p. RNA. 2005 May; 11(5):821-30.
    View in: PubMed
  22. Alexandrov A, Vignali M, LaCount DJ, Quartley E, de Vries C, De Rosa D, Babulski J, Mitchell SF, Schoenfeld LW, Fields S, Hol WG, Dumont ME, Phizicky EM, Grayhack EJ. A facile method for high-throughput co-expression of protein pairs. Mol Cell Proteomics. 2004 Sep; 3(9):934-8.
    View in: PubMed
  23. Van Slyke C, Grayhack EJ. The essential transcription factor Reb1p interacts with the CLB2 UAS outside of the G2/M control region. Nucleic Acids Res. 2003 Aug 1; 31(15):4597-607.
    View in: PubMed
  24. Phizicky EM, Martzen MR, McCraith SM, Spinelli SL, Xing F, Shull NP, Van Slyke C, Montagne RK, Torres FM, Fields S, Grayhack EJ. Biochemical genomics approach to map activities to genes. Methods Enzymol. 2002; 350:546-59.
    View in: PubMed
  25. Grayhack EJ, Phizicky EM. Genomic analysis of biochemical function. Curr Opin Chem Biol. 2001 Feb; 5(1):34-9.
    View in: PubMed
  26. Martzen MR, McCraith SM, Spinelli SL, Torres FM, Fields S, Grayhack EJ, Phizicky EM. A biochemical genomics approach for identifying genes by the activity of their products. Science. 1999 Nov 5; 286(5442):1153-5.
    View in: PubMed
  27. Kuo MH, Nadeau ET, Grayhack EJ. Multiple phosphorylated forms of the Saccharomyces cerevisiae Mcm1 protein include an isoform induced in response to high salt concentrations. Mol Cell Biol. 1997 Feb; 17(2):819-32.
    View in: PubMed
  28. Kuo MH, Grayhack E. A library of yeast genomic MCM1 binding sites contains genes involved in cell cycle control, cell wall and membrane structure, and metabolism. Mol Cell Biol. 1994 Jan; 14(1):348-59.
    View in: PubMed
  29. Grayhack EJ. The yeast alpha 1 and MCM1 proteins bind a single strand of their duplex DNA recognition site. Mol Cell Biol. 1992 Aug; 12(8):3573-82.
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  30. Yang XJ, Hart CM, Grayhack EJ, Roberts JW. Transcription antitermination by phage lambda gene Q protein requires a DNA segment spanning the RNA start site. Genes Dev. 1987 May; 1(3):217-26.
    View in: PubMed
  31. Grayhack EJ, Yang XJ, Lau LF, Roberts JW. Phage lambda gene Q antiterminator recognizes RNA polymerase near the promoter and accelerates it through a pause site. Cell. 1985 Aug; 42(1):259-69.
    View in: PubMed
  32. Grayhack EJ, Roberts JW. Purification of the bacteriophage lambda late gene regulator encoded by gene Q. J Biol Chem. 1983 Aug 10; 258(15):9192-6.
    View in: PubMed
  33. Grayhack EJ, Roberts JW. The phage lambda Q gene product: activity of a transcription antiterminator in vitro. Cell. 1982 Sep; 30(2):637-48.
    View in: PubMed

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