HPat provides a link between deadenylation and decapping in metazoa

Haas, Gabrielle; Braun, Joerg E.; Igreja, Cátia; Tritschler, Felix; Nishihara, Tadashi; Izaurralde, Elisa

doi:10.1083/jcb.200910141

Cited by 73 publications

(157 citation statements)

References 36 publications

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“…Finally, mutation of Y47 and W204 to alanine increase the half-life of reporter mRNA in vivo under conditions when Edc1 and 2 are not expressed, suggesting that additional proteins in yeast bind Dcp1 through the PRS binding site (Tharun and Parker 1999). It is noteworthy that additional decapping coactivators, such as Pat1 and PNRC2, have proline-rich sequences that are required for function (Cho et al 2009;Braun et al 2010;Haas et al 2010). The question arises whether these proline-rich sequences confer to the consensus motif found here.…”

Section: Discussionmentioning

confidence: 90%

Dcp1 links coactivators of mRNA decapping to Dcp2 by proline recognition

Borja

Piotukh²,

Freund³

et al. 2010

RNA

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Cap hydrolysis is a critical step in several eukaryotic mRNA decay pathways and is carried out by the evolutionarily conserved decapping complex containing Dcp2 at the catalytic core. In yeast, Dcp1 is an essential activator of decapping and coactivators such as Edc1 and Edc2 are thought to enhance activity, though their mechanism remains elusive. Using kinetic analysis we show that a crucial function of Dcp1 is to couple the binding of coactivators of decapping to activation of Dcp2. Edc1 and Edc2 bind Dcp1 via its EVH1 proline recognition site and stimulate decapping by 1000-fold, affecting both the K M for mRNA and rate of the catalytic step. The C-terminus of Edc1 is necessary and sufficient to enhance the catalytic step, while the remainder of the protein likely increases mRNA binding to the decapping complex. Lesions in the Dcp1 EVH1 domain or the Edc1 prolinerich sequence are sufficient to block stimulation. These results identify a new role of Dcp1, which is to link the binding of coactivators to substrate recognition and activation of Dcp2.

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Section: Discussionmentioning

confidence: 90%

Dcp1 links coactivators of mRNA decapping to Dcp2 by proline recognition

Borja

Piotukh²,

Freund³

et al. 2010

RNA

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“…However, evidence defining the molecular factors that physically link these two processes has been lacking. Dhh1p and metazoan Pat1 proteins have been likely candidates due to their abilities to enhance decapping and associate with CCR4-NOT (Coller et al 2001;Fischer and Weis 2002;Haas et al 2010;Nissan et al 2010;Ozgur et al 2010). Pat1 directly binds to DDX6 (Sharif et al 2013), suggesting that it may associate indirectly with CCR4-NOT via DDX6.…”

Section: Discussionmentioning

confidence: 99%

“…Finally, decapped miRNA targets are degraded by the 5 ′ -3 ′ exonuclease, Xrn1 (Rehwinkel et al 2005;Chen et al 2009). Coupling between deadenylation and decapping in eukaryotes occurs through an ill-understood molecular mechanism, although DDX6 and Pat1b are thought to functionally link both processes (Coller et al 2001;Haas et al 2010;Ozgur et al 2010). The contribution of DDX6 and Pat1b to the transition from mRNA deadenylation to decapping, as well as the protein-protein contacts that link the deadenylation and decapping complexes, remain unclear.…”

Section: Introductionmentioning

confidence: 99%

Human DDX6 effects miRNA-mediated gene silencing via direct binding to CNOT1

Rouya

Siddiqui

Morita

et al. 2014

RNA

121

145

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MicroRNAs (miRNAs) play critical roles in a variety of biological processes through widespread effects on protein synthesis. Upon association with the miRNA-induced silencing complex (miRISC), miRNAs repress target mRNA translation and accelerate mRNA decay. Degradation of the mRNA is initiated by shortening of the poly(A) tail by the CCR4-NOT deadenylase complex followed by the removal of the 5 ′ cap structure and exonucleolytic decay of the mRNA. Here, we report a direct interaction between the large scaffolding subunit of CCR4-NOT, CNOT1, with the translational repressor and decapping activator protein, DDX6. DDX6 binds to a conserved CNOT1 subdomain in a manner resembling the interaction of the translation initiation factor eIF4A with eIF4G. Importantly, mutations that disrupt the DDX6-CNOT1 interaction impair miRISC-mediated gene silencing in human cells. Thus, CNOT1 facilitates recruitment of DDX6 to miRNA-targeted mRNAs, placing DDX6 as a downstream effector in the miRNA silencing pathway.

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“…Overexpression of Pat1 induces p-body formation in both yeast and human cells (Coller and Parker 2005;Ozgur et al 2010). Pat1 facilitates p-body assembly, probably by acting as a scaffolding protein since it interacts with numerous decay factors, including the decapping enzyme, decapping activators, the 5 ′ -to-3 ′ exonuclease Xrn1, and components of the Ccr4 deadenylase complex as revealed by studies in human cells, Drosophila, and yeast Haas et al 2010;Nissan et al 2010;Ozgur et al 2010). Human and yeast Pat1 also have the potential to self-associate (Nissan et al 2010;Ozgur et al 2010), a property that may facilitate p-body assembly.…”

Section: Introductionmentioning

confidence: 99%

Pat1 contributes to the RNA binding activity of the Lsm1-7–Pat1 complex

Chowdhury

Kalurupalle

Tharun

2014

RNA

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A major mRNA decay pathway in eukaryotes is initiated by deadenylation followed by decapping of the oligoadenylated mRNAs and subsequent 5 ′ -to-3 ′ exonucleolytic degradation of the capless mRNA. In this pathway, decapping is a rate-limiting step that requires the hetero-octameric Lsm1-7-Pat1 complex to occur at normal rates in vivo. This complex is made up of the seven Sm-like proteins, Lsm1 through Lsm7, and the Pat1 protein. It binds RNA and has a unique binding preference for oligoadenylated RNAs over polyadenylated RNAs. Such binding ability is crucial for its mRNA decay function in vivo. In order to determine the contribution of Pat1 to the function of the Lsm1-7-Pat1 complex, we compared the RNA binding properties of the Lsm1-7 complex purified from pat1Δ cells and purified Pat1 fragments with that of the wild-type Lsm1-7-Pat1 complex. Our studies revealed that both the Lsm1-7 complex and purified Pat1 fragments have very low RNA binding activity and are impaired in the ability to recognize the oligo(A) tail on the RNA. However, reconstitution of the Lsm1-7-Pat1 complex from these components restored these abilities. We also observed that Pat1 directly contacts RNA in the context of the Lsm1-7-Pat1 complex. These studies suggest that the unique RNA binding properties and the mRNA decay function of the Lsm1-7-Pat1 complex involve cooperation of residues from both Pat1 and the Lsm1-7 ring. Finally our studies also revealed that the middle domain of Pat1 is essential for the interaction of Pat1 with the Lsm1-7 complex in vivo.

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HPat provides a link between deadenylation and decapping in metazoa

Cited by 73 publications

References 36 publications

Dcp1 links coactivators of mRNA decapping to Dcp2 by proline recognition

Dcp1 links coactivators of mRNA decapping to Dcp2 by proline recognition

Human DDX6 effects miRNA-mediated gene silencing via direct binding to CNOT1

Pat1 contributes to the RNA binding activity of the Lsm1-7–Pat1 complex

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