Regulation of Yeast Cell Growth and Division

Research

My laboratory studies molecular mechanisms that regulate growth and division in the budding yeast Saccharomyces cerevisiae. A surprisingly large number of fundamental cellular processes are conserved between yeast and higher eukaryotes. The combination of the powerful classical and molecular genetic methodologies available, the well annotated genome and the ability to carry out sophisticated biochemistry and cell biology have turned budding yeast into a valuable model system in which to study conserved regulatory mechanisms.

A major current focus is on Elongator, a conserved, six-subunit protein complex. Although Saccharomyces cerevisiae Elongator is non-essential for growth in the laboratory, it is essential for mammalian development and mutations in its Elp1 subunit are associated with Familial Dysautonomia, a neurodevelopmental disease. Over the past decade a variety of roles have been proposed for Elongator, but the principal and possibly only role of Elongator in yeast is to promote two related chemical modifications to the uridine residue that is present at the anticodon ‘wobble’ position (U34) in a subset of tRNAs. These modifications to U34 (termed mcm⁵U and ncm⁵U) are required for wobble uridine-containing tRNAs to function efficiently in protein synthesis, and Elongator’s role in wobble uridine modification is conserved in plants and worms (C. elegans).

Yeast Elp1 shows Hrr25-dependent phosphorylation and we have recently shown that this is required for Elongator functionality. Hrr25, a yeast casein kinase I orthologue, directly phosphorylates Elp1 and Elongator-dependent tRNA wobble uridine modification depends on phosphorylation of the Hrr25 sites along with other, adjacent phosphorylation sites that are not directly phosphorylated by Hrr25. We are currently dissecting the phosphoregulatory domain in Elp1 and trying to understanding in molecular terms how Elp1 phosphorylation promotes Elongator function.

A second area of interest is in mechanisms that ensure chromosomes segregate properly when cells divide. Correct chromosome segregation is vital for maintenance of genome integrity, and gain or loss of chromosomes due to errors in chromosome segregation is a hallmark of cancer cells. The conserved protein kinase Ipl1/Aurora B is important as part of a correction mechanism that ensures sister chromatids become attached to microtubules from opposite spindle poles during mitotic metaphase – a state termed chromosome bi-orientation – so that they are pulled in opposite directions during anaphase when chromosomes segregate. Ipl1 kinase activity is regulated by its association with other components of the ‘chromosome passenger complex’ – Sli15 (INCENP), Bir1 (Survivin) and Nbl1 (Borealin) – and we are using yeast to understand how these regulatory subunits control Ipl1 kinase activity during chromosome bi-orientation. We would like to understand how tension regulates Ipl1-dependent phosphorylation of its targets at the kinetochore and our current focus is on the role of Bir1, part of the Ipl1-associated chromosome passenger complex that like Ipl1 is required for chromosome biorientation.

Teaching

Publications

Mehlgarten, C., Jablonowski, D., Breunig, K. D., Stark, M.J.R. and Schaffrath, R. (2009) Elongator function depends on antagonistic regulation by casein kinase Hrr25 and protein phosphatase Sit4. Mol Microbiol. 79, 869-881. PMID: 19656297  View Paper

Keating, P., Rachidi, N., Tanaka, T.U., and Stark, M.J.R., (2009) lpl1-dependent phosphorylation of Dam1 is reduced by tension applied on kinetochores. J Cell Sci 122:4375-4382 PMID: 19923271  View Paper

Makrantoni, V., and Stark, M.J.R., (2009) Efficient chromosome bi-orientation and the tension checkpoint in Saccharomyces cerevisiae both require Bir1p. Mol Cell Biol 29:4552-4562 PMID: 19528231  View Paper

Lain, S., Hollick, J.J., Campbell, J., Staples, O.D., Higgins, M., Aoubala, M., McCarthy, A., Appleyard, V., Murray, K.E., Baker, L., Thompson, A., Mathers, J., Holland, S.J., Stark, M.J.R., Pass, G., Woods, J., Lane, D.P., Westwood, N.J. (2008) Discovery, in vivo activity, and mechanism of action of a small-molecule p53 activator. Cancer Cell 13:454-463 PMID: 18455128 View Paper

King, E.M., Rachidi, N., Morrice, N., Hardwick, K.G., Stark, M.J.R., (2007) IpI1p-dependent phosphorylation of Mad3p is required for the spindle checkpoint response to lack of tension at kinetochores. Genes Devel. 21:1163-1168. PMID: 17504936 View Paper

Fox, G.C., Shafiq, M., Briggs, D.C., Knowles, P.P., Collister, M., Didmon, M.J., Makrantoni, V., Dickinson, R.J., Hanrahan, S., Totty, N., Stark, M.J.R., Keyse, S.M., McDonald, N.Q. (2007) Redox-mediated substrate recognition by Sdp1 defines a new group of tyrosine phosphates. Nature 447: 487-492. PMID: 17495930 View Paper

Rudolf, J., Makrantoni, V., Ingledew, W.J., Stark, M.J.R., White, M.F. (2006) The DNA repair helicases XPD and FancJ have essential iron-sulfur domains. Mol. Cell 23:801-808. PMID: 16973432 View Paper

Tanaka, T.U., Rachidi, N., Janke, C., Pereira, G., Galova, M., Schiebel, E., Stark, M.J.R., Nasmyth, K. (2002) Evidence that the IpI1-Sli15 (Aurora kinase-INCENP) complex promotes chromosome bi-orientation by altering kinetochore-spindle pole connections. Cell 108: 317-329. PMID: 11853667 View Paper

Impact