In recent years a tremendous amount of excitement has surrounded the finding that regulated proteolysis is intricately involved in controlling the cell cycle in yeast and mammals. We have found that regulated ribonuclease digestion also plays a crucial part in controlling the cell cycle in yeast. The major objective for this proposal is to determine the sites, mechanism and regulation of mRNA destabilization by the ribonucleoprotein endoribonuclease RNase MRP in late telophase of the cell division cycle. We have discovered that mutations of different components of the RNase MRP enzyme complex lead to a cell division cycle (CDC) arrest. Divided nuclei and an elongated spindle characterize this late cell cycle arrest, in which cells fail to disassemble their spindle or to undergo cytokinesis. This M to G1 arrest is classically defined by the CDC5, DBF2 group of mutations. These M to G1 mutants lead to the inability to turn off the Clb1&2/Cdc28 cyclin/kinase. Inactivation of the kinase is required for disassembly of the spindle, cytokinesis and the ability to enter G1. Results indicate that the cell division cycle arrest in RNase MRP mutants is the result of a failure to rapidly degrade the CLB2 mRNA. Increased CLB2 mRNA leads to increased CIb2 protein, associated kinase activity and cell cycle arrest. The specific goals during the project period will include characterizing the pathway of CLB2 mRNA degradation and determining direct targets of RNase MRP in addition to CLB2 mRNA. Genome technology will be used to identify all of the potential RNase MRP substrates in a cell. A series of biochemical and genetic analyses will identify instability elements and the direct sites of RNase MRP action. Lastly, we will investigate the regulation of RNase MRP during the cell cycle and determine the means of that regulation. The significance of the role of RNase MRP in the M to G1 transition cannot be overstated. Regulated degradation of specific mRNAs may play as large a part in cell cycle control as regulated proteolysis. Many of these late acting genes have human homologues (PTEN/MMAC1, PIk, Polo, p55CDC) that are intimately involved in regulating the cell cycle and cell proliferation. Humans also have an RNase MRP complex with RNA and protein subunits similar to the yeast enzyme. Mutations in human RNase MRP cause a pleotropic disease, Cartilage Hair Hypoplasia. The manifestations of this disease are caused by a general cell proliferation defect, similar to what we see in yeast. This will allow us to use a simple genetic model system to lend insights into human disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM063798-04
Application #
7007265
Study Section
Physiological Chemistry Study Section (PC)
Program Officer
Zatz, Marion M
Project Start
2003-02-01
Project End
2008-01-31
Budget Start
2006-02-01
Budget End
2008-01-31
Support Year
4
Fiscal Year
2006
Total Cost
$267,170
Indirect Cost
Name
Upstate Medical University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
058889106
City
Syracuse
State
NY
Country
United States
Zip Code
13210
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Jaag, Hannah M; Lu, Qiasheng; Schmitt, Mark E et al. (2011) Role of RNase MRP in viral RNA degradation and RNA recombination. J Virol 85:243-53
Lu, Qiaosheng; Wierzbicki, Sara; Krasilnikov, Andrey S et al. (2010) Comparison of mitochondrial and nucleolar RNase MRP reveals identical RNA components with distinct enzymatic activities and protein components. RNA 16:529-37
Esakova, Olga; Perederina, Anna; Quan, Chao et al. (2008) Footprinting analysis demonstrates extensive similarity between eukaryotic RNase P and RNase MRP holoenzymes. RNA 14:1558-67
Perederina, Anna; Esakova, Olga; Koc, Hasan et al. (2007) Specific binding of a Pop6/Pop7 heterodimer to the P3 stem of the yeast RNase MRP and RNase P RNAs. RNA 13:1648-55
Gill, Tina; Aulds, Jason; Schmitt, Mark E (2006) A specialized processing body that is temporally and asymmetrically regulated during the cell cycle in Saccharomyces cerevisiae. J Cell Biol 173:35-45
Thiel, Christian T; Horn, Denise; Zabel, Bernhard et al. (2005) Severely incapacitating mutations in patients with extreme short stature identify RNA-processing endoribonuclease RMRP as an essential cell growth regulator. Am J Hum Genet 77:795-806
Hermanns, Pia; Bertuch, Alison A; Bertin, Terry K et al. (2005) Consequences of mutations in the non-coding RMRP RNA in cartilage-hair hypoplasia. Hum Mol Genet 14:3723-40
Gill, Tina; Cai, Ti; Aulds, Jason et al. (2004) RNase MRP cleaves the CLB2 mRNA to promote cell cycle progression: novel method of mRNA degradation. Mol Cell Biol 24:945-53