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.
| 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 |
| 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 |
| Elbarbary, Reyad A; Maquat, Lynne E (2014) Dodging two bullets with one dsRNA-binding protein. Cell Cycle 13:345-6 |
| 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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