Graduate Program Information
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Professor of Biochemistry and Biophysics
Ph.D. Cornell University 1983

tRNA

We are interested in understanding how tRNA is processed, and why it has its modifications. tRNA is by far the most numerous class of RNA found in the cell, and a large number of proteins are required to trim the 5' leader and 3' trailer, to splice the intron if present, and to modify the appropriate nucleosides, as well as to direct tRNA trafficking and regulate its quality. In the yeast S. cerevisiae there are 25 biochemically distinct modifications, many of which are highly conserved in different organisms, biochemically fascinating, and of unknown function. We have identified 10 genes encoding proteins that modify tRNAs, including four dihydrouridine synthases responsible for modification of specific residues in different tRNAs, a tRNA m⁷G methyltransferase comprised of a complex of two proteins, an unusual m¹G₉ methyltransferase unlike other members of the widely conserved methyltransferase family, and an essential tRNAHis guanylyltransferase that adds a guanosine residue to the 5' end of tRNA^His, formally elongating the tRNA in a 3'-5' direction. We are currently working to define the basis of the substrate specificity of these proteins, to determine their mechanisms, and to define the role of these modifications in vivo. Recently, this research has led to the discovery of a new quality control pathway responsible for the degradation of tRNA lacking certain modifications, and to the discovery of a novel reverse polymerase activity of tRNA^His guanylyltransferase.

Biochemical Analysis of the Proteome

We previously developed a biochemical genomics approach for rapidly mapping biochemical activities to genes. To this end, we constructed a genomic library of yeast strains, each expressing a unique yeast protein fused to a purification tag. Then we purify pools of fusion proteins derived from these strains, assay the pools for biochemical activity, and then prepare and assay appropriate subpools of fusion proteins to assign the activity to a specific gene. We have used this approach for a large number of different types of biochemical activities. Recently we constructed a comprehensive new genomic library of plasmids and yeast strains (called the MORF library) for sophisticated biochemical and genetic analysis or the proteome (with M. Snyder at Yale and a group led by E. Grayhack at Rochester). We are using this library to define new activities and classes of activities, to evaluate factors that globally affect expression of proteins, and to help define cell circuitry by systematic genetic analysis of suppression caused by overexpression of genes.

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Recent Publications

Chernyakov I, Whipple JM, Kotelawala L, Grayhack EJ, Phizicky EM (2008) Degradation of several hypomodified mature tRNA species in Saccharomyces cerevisiae is mediated by Met22 and the 5'-3' exonucleases Rat1 and Xrn1. Genes Dev, 22:1369-80

Kotelawala L, Grayhack EJ, Phizicky EM (2008) 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, 14:158-69

Phizicky EM (2008) Waste not, want not: a case for tRNA repair. Nat Struct Mol Biol, 15:783-4

Jackman JE, Phizicky EM (2008) Identification of critical residues for G-1 addition and substrate recognition by tRNA(His) guanylyltransferase. Biochemistry, 47:4817-25

Jackman JE, Grayhack EJ, Phizicky EM (2008) The use of Saccharomyces cerevisiae proteomic libraries to identify RNA-modifying proteins. Methods Mol Biol, 488:383-93

Wilkinson ML, Crary SM, Jackman JE, Grayhack EJ, Phizicky EM (2007) The 2'-O-methyltransferase responsible for modification of yeast tRNA at position 4. Rna, 13:404-13

Malkowski MG, Quartley E, Friedman AE, Babulski J, Kon Y, Wolfley J, Said M, Luft JR, Phizicky EM, DeTitta GT, Grayhack EJ (2007) Blocking S-adenosylmethionine synthesis in yeast allows selenomethionine incorporation and multiwavelength anomalous dispersion phasing. Proc Natl Acad Sci U S A, 104:6678-83

Jackman JE, Kotelawala L, Grayhack EJ, Phizicky EM (2007) Identification and characterization of modification enzymes by biochemical analysis of the proteome. Methods Enzymol, 425:139-52

Phizicky EM, Grayhack EJ (2006) Proteome-scale analysis of biochemical activity. Crit Rev Biochem Mol Biol, 41:315-27

Jackman JE, Phizicky EM (2006) tRNAHis guanylyltransferase catalyzes a 3'-5' polymerization reaction that is distinct from G-1 addition. Proc Natl Acad Sci U S A, 103:8640-5

Jackman JE, Phizicky EM (2006) tRNAHis guanylyltransferase adds G-1 to the 5' end of tRNAHis by recognition of the anticodon, one of several features unexpectedly shared with tRNA synthetases. Rna, 12:1007-14

Alexandrov A, Chernyakov I, Gu W, Hiley SL, Hughes TR, Grayhack EJ, Phizicky EM (2006) Rapid tRNA decay can result from lack of nonessential modifications. Mol Cell, 21:87-96

Gu W, Hurto RL, Hopper AK, Grayhack EJ, Phizicky EM (2005) Depletion of Saccharomyces cerevisiae tRNA(His) guanylyltransferase Thg1p leads to uncharged tRNAHis with additional m(5)C. Mol Cell Biol, 25:8191-201

Cartlidge RA, Knebel A, Peggie M, Alexandrov A, Phizicky EM, Cohen P (2005) The tRNA methylase METTL1 is phosphorylated and inactivated by PKB and RSK in vitro and in cells. Embo J, 24:1696-705

Alexandrov A, Grayhack EJ, Phizicky EM (2005) tRNA m7G methyltransferase Trm8p/Trm82p: evidence linking activity to a growth phenotype and implicating Trm82p in maintaining levels of active Trm8p. Rna, 11:821-30

Steiger MA, Jackman JE, Phizicky EM (2005) Analysis of 2'-phosphotransferase (Tpt1p) from Saccharomyces cerevisiae: evidence for a conserved two-step reaction mechanism. Rna, 11:99-106

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 (2005) Biochemical and genetic analysis of the yeast proteome with a movable ORF collection. Genes Dev, 19:2816-26

Phizicky EM (2005) Have tRNA, will travel. Proc Natl Acad Sci U S A, 102:11127-8

Shull NP, Spinelli SL, Phizicky EM (2005) A highly specific phosphatase that acts on ADP-ribose 1''-phosphate, a metabolite of tRNA splicing in Saccharomyces cerevisiae. Nucleic Acids Res, 33:650-60

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 (2004) A facile method for high-throughput co-expression of protein pairs. Mol Cell Proteomics, 3:934-8

Xing F, Hiley SL, Hughes TR, Phizicky EM (2004) The specificities of four yeast dihydrouridine synthases for cytoplasmic tRNAs. J Biol Chem, 279:17850-60

Graduate students in my laboratory work toward a Ph.D. degree in the following program[s]:

Ph.D. in Biochemistry
Ph.D. in Biophysics
Ph.D. in Genetics

Ph.D. candidates in my laboratory may also be affiliated with these programs:

Contact Information

E-Mail: Eric_Phizicky@urmc.rochester.edu

Eric Phizicky
Department of Biochemistry and Biophysics
University of Rochester School of Medicine and Dentistry
601 Elmwood Ave, Box 712
Rochester, New York 14642

Office: Medical Center 3-7411
Telephone: (585) 275-7268; Fax: (585) 271-2683