Nonsense-mediated mRNA decay (NMD) in mammalian cells can be viewed as (i) a quality control mechanism that degrades abnormal mRNAs so as to eliminate the production of potentially deleterious truncated proteins and (ii) a regulatory mechanism that targets normal mRNAs so as to allow for proper levels of gene expression. We have shown that NMD generally degrades mRNAs that terminatetranslation more than 50-55 nucleotides upstream of a splicing-generated exon-exon junction. During the past funding period, we have demonstrated that the role of the exon-exon junction reflects the splicing-dependent deposition of an exon-junction complex (EJC) of proteins, which recruits Upf factors that elicit NMD when translation terminates sufficiently upstream. Furthermore, we have shown that Upfl is phosphorylated by the PIK-related protein kinase Smg1 and dephosphorylated in reactions that depend on at least two of three Smg5/7 factors, which were renamed Smg5, Smg6 and Smg7 once sequences for the C. elegans orthologs became available. This application aims to extend these and related studies.
Aim 1 is to continue to examine Upf1 function, including its role in a new mRNA decay pathway that we have uncovered: Staufenl- mediated mRNA decay (SMD).
Aim 2 is to define and characterize what we have called mRNA """"""""failsafe"""""""" sequences, which can mediate NMD in place of the 3'-most exon-exon junction (i.e., when the 3'-most intron has been deleted). Notably, data indicate that a failsafe sequence elicits NMD only if the mRNA derives from pre-mRNA that harbors at least one intron.
Aim 3 is to study the structure of mRNP that is targeted for NMD in S. cerevisiae. To date, we have demonstrated that NMD targets both Cbclp-bound and elF4E- bound mRNAs. The proposed experiments logically extend our long-time studies, and they should lend important insight into aspects of SMD and NMD that have yet to be understood.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM074593-25
Application #
7523238
Study Section
Molecular Genetics B Study Section (MGB)
Program Officer
Bender, Michael T
Project Start
1984-04-01
Project End
2009-11-30
Budget Start
2008-12-01
Budget End
2009-11-30
Support Year
25
Fiscal Year
2009
Total Cost
$333,247
Indirect Cost
Name
University of Rochester
Department
Biochemistry
Type
Schools of Dentistry
DUNS #
041294109
City
Rochester
State
NY
Country
United States
Zip Code
14627
Gong, Chenguang; Maquat, Lynne E (2015) Affinity purification of long noncoding RNA-protein complexes from formaldehyde cross-linked mammalian cells. Methods Mol Biol 1206:81-6
Elbarbary, Reyad A; Maquat, Lynne E (2014) Dodging two bullets with one dsRNA-binding protein. Cell Cycle 13:345-6
Gleghorn, Michael L; Maquat, Lynne E (2014) 'Black sheep' that don't leave the double-stranded RNA-binding domain fold. Trends Biochem Sci 39:328-40
Popp, Maximilian Wei-Lin; Maquat, Lynne E (2014) Defective secretory-protein mRNAs take the RAPP. Trends Biochem Sci 39:154-6
Gong, Chenguang; Tang, Yalan; Maquat, Lynne E (2013) mRNA-mRNA duplexes that autoelicit Staufen1-mediated mRNA decay. Nat Struct Mol Biol 20:1214-20
Park, Eonyoung; Maquat, Lynne E (2013) Staufen-mediated mRNA decay. Wiley Interdiscip Rev RNA 4:423-35
Elbarbary, Reyad A; Li, Wencheng; Tian, Bin et al. (2013) STAU1 binding 3' UTR IRAlus complements nuclear retention to protect cells from PKR-mediated translational shutdown. Genes Dev 27:1495-510
Wang, Jiashi; Gong, Chenguang; Maquat, Lynne E (2013) Control of myogenesis by rodent SINE-containing lncRNAs. Genes Dev 27:793-804
Gleghorn, Michael L; Gong, Chenguang; Kielkopf, Clara L et al. (2013) Staufen1 dimerizes through a conserved motif and a degenerate dsRNA-binding domain to promote mRNA decay. Nat Struct Mol Biol 20:515-24
Park, Eonyoung; Gleghorn, Michael L; Maquat, Lynne E (2013) Staufen2 functions in Staufen1-mediated mRNA decay by binding to itself and its paralog and promoting UPF1 helicase but not ATPase activity. Proc Natl Acad Sci U S A 110:405-12

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