The long term goal of this proposal is to understand, in detail, the process of cessation of mRNA translation. We have uncovered compelling evidence that shutting down mRNA translation is not simply a passive and default event in the lifetime of an mRNA, but, rather, that mRNA is actively removed from the translational machinery in response to specific cues. Importantly, we have identified the ATP-dependent RNA helicase, Dhh1 p, as a factor required for the dissociation of mRNA from translation. Furthermore, data suggests that Dhhlp functions to exit an mRNA from translation following loss of the poly(A) tail from the 3'end of the mRNA. We have also demonstrated that Dhhlp is used by the cell to achieve global regulation of mRNA expression under conditions of nutrient-starvation. Dhhlp acts, therefore, both as a cis-acitng . regulator of mRNA translation under normal cell growth conditions and a global regulator of mRNA expression upon cellular stress. We propose to extend our initial identification of this unexplored step of gene regulation under three specific aims. First, we will examine the role of Dhhlp in the repression of global mRNA translation that is induced by nutrient starvation. Our preliminary observations indicate that under either glucose and amino acid starvation, mRNA 3'poly(A) tail status or function is rapidly and radically altered. We will investigate how this dramatic alteration in poly(A) tail function is achieved and if it is required for the movement of mRNA into translational quiescence by Dhhlp upon stress. Second, we will dissect the nature of the observed interaction between Dhhlp and mRNA undergoing translation. In particular, we will determine the mode by which Dhhlp interacts with translating mRNAs, and if Dhhlp specifically interacts with either ribosomes or with the mRNA itself. Lastly, we will investigate the molecular mechanisms by which Dhhlp promotes the exit of mRNA out of translation under normal and nutrient stress conditions. Preliminary evidence suggests that the substrate for Dhhlp activity is the 40S ribosomal subunit itself. The nature of this interaction will be more rigorously tested, and genetic and biochemical analyses will be preformed to identify additional substrates of Dhhlp. In aggregate, we will explore further the mechanisms by which mRNA is removed from translation dependent upon Dhhlp. Importantly, Dhhlp activity appears to be functionally conserved. Homologues of Dhhlp are critical for translational silencing of maternal mRNAs in oocytes, and mutations lead to precocious activation of translationally silenced maternal mRNA and the manifestation of embryonic defects. Consistent with these observations, mouse and human Dhhlp has been implicated in gametogenesis and carcinogenesis by impacting mRNA translation. The proposed studies, therefore, can be anticipated to provide relevant insight into public health issues such as the manifestation of birth defects, sterility, and cellular transformation.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
3R01GM080465-03S1
Application #
7887958
Study Section
Molecular Genetics A Study Section (MGA)
Program Officer
Bender, Michael T
Project Start
2009-08-10
Project End
2012-07-31
Budget Start
2009-08-10
Budget End
2012-07-31
Support Year
3
Fiscal Year
2009
Total Cost
$181,321
Indirect Cost
Name
Case Western Reserve University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
077758407
City
Cleveland
State
OH
Country
United States
Zip Code
44106
Radhakrishnan, Aditya; Chen, Ying-Hsin; Martin, Sophie et al. (2016) The DEAD-Box Protein Dhh1p Couples mRNA Decay and Translation by Monitoring Codon Optimality. Cell 167:122-132.e9
Coller, Jeff (2016) mRNA decapping in 3D. Nat Struct Mol Biol 23:954-956
Chen, Ying-Hsin; Coller, Jeff (2016) A Universal Code for mRNA Stability? Trends Genet 32:687-688
Alhusaini, Najwa; Coller, Jeff (2016) The deadenylase components Not2p, Not3p, and Not5p promote mRNA decapping. RNA 22:709-21
Richter, Joel D; Coller, Jeff (2015) Pausing on Polyribosomes: Make Way for Elongation in Translational Control. Cell 163:292-300
Presnyak, Vladimir; Alhusaini, Najwa; Chen, Ying-Hsin et al. (2015) Codon optimality is a major determinant of mRNA stability. Cell 160:1111-24
Martin, Sophie; Coller, Jeff (2014) PAN-orama: three convergent views of a eukaryotic deadenylase. Nat Struct Mol Biol 21:577-8
Smith, Jenna E; Alvarez-Dominguez, Juan R; Kline, Nicholas et al. (2014) Translation of small open reading frames within unannotated RNA transcripts in Saccharomyces cerevisiae. Cell Rep 7:1858-66
Geisler, Sarah; Coller, Jeff (2013) RNA in unexpected places: long non-coding RNA functions in diverse cellular contexts. Nat Rev Mol Cell Biol 14:699-712
Presnyak, Vlad; Coller, Jeff (2013) The DHH1/RCKp54 family of helicases: an ancient family of proteins that promote translational silencing. Biochim Biophys Acta 1829:817-23

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