LYNNE MAQUAT

..Failure is most likely because each mRNA lacks a cis-acting destabilizing element, because insertion of a spliceable intron a sufficient distance downstream of a nonsense codon within either gene is sufficient to elicit NMD...

..Translation factors, RNA-binding proteins, and targets of signaling pathways that are particular to newly synthesized mRNAs regulate critical functions of the pioneer round...

..We describe how a factor that is common to both pathways results in their competition. We also explain how competition between the two pathways contributes to the differentiation of C2C12 myoblasts to multinucleated myotubes...

..Here, we overview cellular processes by which CBC-bound mRNPs are remodeled to eIF4E-bound mRNPs. We also describe the molecular movements of certain factors during NMD in view of the influential role of CBP80...

..The acquisition and loss of mRNA-associated proteins accompanies the transition from the pioneer round to subsequent rounds of translation, and from translational competence to substrate for nonsense-mediated mRNA decay...

..Such evidence derives from the initial finding that GPx1 mRNA is a natural substrate of nonsense-mediated decay. Here, recent work on GPx1 RNA metabolism is reviewed and future directions of study are defined...

..They also indicate that CBP80-bound mRNA undergoes a "pioneer" round of translation, before CBP80-CBP20 are replaced by eIF4E, and Upf2 and Upf3 proteins dissociate from upstream of exon-exon junctions...

..Each of these proteins is also detected on CBP80-bound mRNA in the cytoplasmic fraction, indicating a presence on mRNA after export. The dynamics of mRNP composition before and after mRNA export are discussed...

..Our results are discussed in the context of pre-mRNA processing and stability and mRNA trafficking and the pioneer round of translation...

..Here, we will review how to determine which PTCs elicit NMD, what is currently known about the mechanism of NMD, and additional information that is pertinent to establishing therapies for PTC-associated diseases...

..Along with other types of quality control that occur during the complex processes of mRNA biogenesis, these mRNA surveillance mechanisms help to ensure the integrity of protein-encoding gene expression...

..Our studies, including analyses of factor dependence, reveal important shared features of the two mammalian-cell NMD pathways as well as fundamental differences between NMD in mammals and Saccharomyces cerevisiae...

..Our results uncover an additional role for SF2/ASF and indicate that the efficiency of the pioneer round of translation influences the efficiency of subsequent rounds of translation...

..We show that EMCV IRES-initiated translation undergoes a CBP80/20-associated pioneer round of translation that results in CBP80/20-dependent and Upf factor-dependent NMD when translation terminates prematurely...

..Our results uncover a critical step during NMD that converts a pioneer translation initiation complex to a translationally compromised mRNP...

..The results further implicate general spliceosomal components in AS regulation...

..Our results suggest that most PTC-introducing AS events are not under positive selection pressure and therefore may not contribute important functional roles...

..SMD depends on the recruitment of Upf1 by the RNA-binding protein Stau1 but does not depend on the other Upf proteins. We find that CBP80 interacts with Upf1 and promotes the interaction of Upf1 with Upf2 but not with Stau1...

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..We also describe a modification using beta-mercaptoethanol that facilitates analyzing proteins that comigrate with antibody light or heavy chains...

..Polysome profiles indicate that CBP80-bound mRNAs are translated less efficiently than their eIF4E-bound counterparts...

..We discuss this pathway as a means for cells to downregulate the expression of Stau1 binding transcripts...

..We propose a model in which eIF4G serves to connect CBP80/20 with other initiation factors during the pioneer round of translation...

..These results suggest that both Cbc1p- and eIF4E-mediated pioneer rounds of translation occur in yeast...

..Although NMD is conserved in all eukaryotes that have been examined, it can manifest mechanistic differences in different organisms. A recent study using Drosophila melanogaster describes a new mechanistic twist to NMD...

..We propose that Stau1 influences the expression of a wide variety of physiologic transcripts and metabolic pathways...

..This assay is simpler and more versatile than the first assay because it can be used to assay any cellular gene in situ provided the cells can be stably transfected with the hUpf1p expression vector...

..From these and other data, we conclude that NMD in mammalian cells degrades mRNAs from both 5' and 3' ends by recruiting decapping and 5'-->3' exonuclease activities as well as deadenylating and 3'-->5' exonuclease activities...

..These data suggest that an endonuclease with preference for UG dinucleotides is involved in the degradation of nonsense-containing and, to a lesser extent, nonsense-free human beta-globin mRNAs in mouse erythroid cells...

..Finally, NMD is suppressed in hSMG-1- but not ATM-deficient cells. These results indicate that hSMG-1 plays important roles in the maintenance of both genome and transcriptome integrity in human cells...

..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. ..

..This program has greatly promoted scientific interactions among laboratories, increased flexibility in the choice of research mentors, and improved graduate recruiting. ..

..We expect that renewed support of this grant will allow us to continue to make major advances in understanding the mechanism of NMD in mammalian cells. ..

..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. ..

..As an example, NMD provides a means by which many RNA binding proteins negatively regulate the genes from which they derived so that an appropriate level of gene expression can be achieved. ..

..We expect that renewed support of this grant will allow us to continue to make major advances in understanding the mechanism of NMD in mammalian cells. ..

..We expect that renewed support of this grant will allow us to continue to make major advances in understanding the mechanism of NMD in mammalian cells. ..

..elegans. Characterize the human homologue of Ubf1 and its role in NMD, in particular studying the role of its ATPase and putative helicase activities, and what cellular proteins it interacts with. ..

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..2. Determine why premature termination of PHGPx mRNA translation at the position of the Sec codon fails to mediate a significant reduction in PHGPx mRNA abundance. ..

..As an example, NMD provides a means by which many RNA binding proteins negatively regulate the genes from which they derived so that an appropriate level of gene expression can be achieved. ..