Unraveling regulation and new components of human P-bodies through a protein interaction framework and experimental validation

Department of Biochemistry and Molecular Biology, The University of Texas Medical School, Houston, Texas 77021, USA.
RNA (Impact Factor: 4.94). 09/2011; 17(9):1619-34. DOI: 10.1261/rna.2789611
Source: PubMed


The cellular factors involved in mRNA degradation and translation repression can aggregate into cytoplasmic domains known as GW bodies or mRNA processing bodies (P-bodies). However, current understanding of P-bodies, especially the regulatory aspect, remains relatively fragmentary. To provide a framework for studying the mechanisms and regulation of P-body formation, maintenance, and disassembly, we compiled a list of P-body proteins found in various species and further grouped both reported and predicted human P-body proteins according to their functions. By analyzing protein-protein interactions of human P-body components, we found that many P-body proteins form complex interaction networks with each other and with other cellular proteins that are not recognized as P-body components. The observation suggests that these other cellular proteins may play important roles in regulating P-body dynamics and functions. We further used siRNA-mediated gene knockdown and immunofluorescence microscopy to demonstrate the validity of our in silico analyses. Our combined approach identifies new P-body components and suggests that protein ubiquitination and protein phosphorylation involving 14-3-3 proteins may play critical roles for post-translational modifications of P-body components in regulating P-body dynamics. Our analyses provide not only a global view of human P-body components and their physical interactions but also a wealth of hypotheses to help guide future research on the regulation and function of human P-bodies.

Download full-text


Available from: Dinghai Zheng, Sep 03, 2015
  • Source
    • "VCS contains two protein–protein interaction WD40 motifs, although they are not necessary to the interaction with DCP1 and DCP2. Deletion of WD40 repeats in Ge-1, the human homolog of VCS, did not compromised the localization into PB, then, WD40 repeats probably function in the recruitment of another mediators of decapping (Yu et al., 2005). The Arabidopsis DCP5 is required for decapping and repression of translation in vivo (Xu and Chua, 2009). "
    [Show abstract] [Hide abstract]
    ABSTRACT: mRNA accumulation is tightly regulated by diverse molecular pathways. The identification and characterization of enzymes and regulatory proteins involved in controlling the fate of mRNA offers the possibility to broaden our understanding of posttranscriptional gene regulation. Processing bodies (P bodies, PB) are cytoplasmic protein complexes involved in degradation and translational arrest of mRNA. Composition and dynamics of these subcellular structures have been studied in animal systems, yeasts and in the model plant Arabidopsis. Their assembly implies the aggregation of specific factors related to decapping, deadenylation, and exoribonucleases that operate synchronously to regulate certain mRNA targets during development and adaptation to stress. Although the general function of PB along with the flow of genetic information is understood, several questions still remain open. This review summarizes data on the composition, potential molecular roles, and biological significance of PB and potentially related proteins in Arabidopsis.
    Frontiers in Plant Science 05/2014; 5:201. DOI:10.3389/fpls.2014.00201 · 3.95 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Messenger RNAs transcribed by RNA polymerase II are modified at their 5'-end by the cotranscriptional addition of a 7-methylguanosine (m7G) cap. The cap is an important modulator of gene expression and the mechanism and components involved in its removal have been extensively studied. At least two decapping enzymes, Dcp2 and Nudt16, and an array of decapping regulatory proteins remove the m7G cap from an mRNA exposing the 5'-end to exonucleolytic decay. In contrast, relatively less is known about the decay of mRNAs that may be aberrantly capped. The recent demonstration that the Saccharomyces cerevisiae Rai1 protein selectively hydrolyzes aberrantly capped mRNAs provides new insights into the modulation of mRNA that lack a canonical m7G cap 5'-end. Whether an mRNA is uncapped or capped but missing the N7 methyl moiety, Rai1 hydrolyzes its 5'-end to generate an mRNA with a 5' monophosphate. Interestingly, Rai1 heterodimerizes with the Rat1 5'-3' exoribonuclease, which subsequently degrades the 5'-end monophosphorylated mRNA. Importantly, Rat1 stimulates the 5'-end hydrolysis activities of Rai1 to generate a 5'-end unprotected mRNA substrate for Rat1 and, in turn, Rai1 stimulates the activity of Rat1. The Rai1-Rat1 heterodimer functions as a molecular motor to detect and degrade mRNAs with aberrant caps and defines a novel quality control mechanism that ensures mRNA 5'-end integrity. The increase in aberrantly capped mRNA population following nutritional stress in S. cerevisiae demonstrates the presence of aberrantly capped mRNAs in cells and further reinforces the functional significance of the Rai1 in ensuring mRNA 5'-end integrity.
    01/2012: pages 165-180;
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The mammalian TOB1 and TOB2 proteins have emerged as key players in repressing cell proliferation. Accumulating evidence indicates that TOBs regulate mRNA deadenylation. A recruitment model was proposed in which TOBs promote deadenylation by recruiting CAF1-CCR4 deadenylase complex to the 3' end of mRNAs by simultaneously binding CAF1 and PABP. However, the exact molecular mechanism underlying TOB-promoted deadenylation remains unclear. It is also unclear whether TOBs' antiproliferative and deadenylation-promoting activities are connected. Here, we combine biochemical analyses with a functional assay directly monitoring deadenylation and mRNA decay to characterize the effects of tethering TOBs or their mutant derivatives to mRNAs. The results provide direct evidence supporting the recruitment model and reveal a link between TOBs' antiproliferative and deadenylation-promoting activities. We also find that TOBs' actions in deadenylation are independent of the phosphorylation state of three serines known to regulate antiproliferative actions, suggesting that TOBs arrest cell growth through at least two different mechanisms. TOB1 and TOB2 were interchangeable in the properties tested here, indicating considerable functional redundancy between the two proteins. We propose that their multiple modes of modulating mRNA turnover and arresting cell growth permit the TOB proteins to coordinate their diverse roles in controlling cell growth and differentiation.
    Molecular and Cellular Biology 03/2012; 32(6):1089-98. DOI:10.1128/MCB.06370-11 · 4.78 Impact Factor
Show more