RNA-associated protein 55 (RAP55) localizes to mRNA processing bodies and stress granules

Abstract

The mRNA processing body (P-body) is a cellular structure that has an important role in mRNA degradation. P-bodies have also been implicated in RNAi-mediated post-transcriptional gene silencing. The objective of this study was to identify and characterize novel components of the mammalian P-body. Approximately 5% of patients with the autoimmune disease primary biliary cirrhosis have antibodies directed against this structure. Serum from one of these patients was used to identify a cDNA encoding RAP55, a 463-amino acid protein. RAP55 colocalized with previously identified P-body components DCP1a and Ge-1. RAP55 contains an N-terminal Sm-like domain and two C-terminal RGG-rich domains separated by an FDF motif. The two RGG domains and the FDF domain were necessary and sufficient to target the protein to P-bodies. A fragment of RAP55 consisting of the FDF and the second RGG domains did not localize to P-bodies, but was able to displace other P-body components from this structure. After cells were subjected to arsenite-induced stress, RAP55 was detected in TIA-containing stress granules. The second RGG domain was necessary and sufficient for stress granule localization. siRNA-mediated knock-down of RAP55 resulted in loss of P-bodies, suggesting that RAP55 acts prior to the 5¢-decapping step in mRNA degradation. The results of this study show that RAP55 is a component of P-bodies in cells at rest and localizes in stress granules in arsenite-treated cells. RAP55 may serve to shuttle mRNAs between P-bodies and stress granules.

LSm14A is a processing body-associated sensor of viral nucleic acids that initiates cellular antiviral response in the early phase of viral infection

“…LSm14A was originally discovered to be a component of the Pbodies (18). Our study demonstrate that LSm14A is a sensor for both viral RNA and DNA and acts as a switch point on viral RNA-and DNA-induced IFN pathways, respectively.…”

“…In the present study, we identified a component of the processing bodies (P-bodies), LSm14A (also called RAP55), as an activator of IRF3 in expression screen experiments (18). Our results indicated that LSm14A bound to synthetic or viral RNA and DNA and was essential for initiating the induction of IFN-β in the early phase of virus infection.…”

“…To identify candidate molecules involved in virus-triggered innate immune response, we screened ∼10,000 independent human cDNA expression plasmids for their ability to regulate ISRE activity by reporter assays. These efforts led to the identification of LSm14A, a member of the LSm family of proteins that are involved in RNA metabolism (18). As shown in Fig.…”

“…LSm14A is a member of the LSm family of proteins, which have been shown to be involved in RNA processing events (18,21). The N terminus of LSm14A, amino acids 1-76, is a conserved LSm domain, whereas the C terminus contains two domains called "DFDF box" (amino acids 291-316) and "FDF_TFG box" (amino acids 361-397), respectively ( Fig.…”

“…It has been demonstrated that LSm14A is localized at the P-bodies (18). To further exclude the possibility that the defects in the innate response in LSm14A knockdown cells are due to disrupting the function of the P-bodies, we designed two RNAi vectors for DDX6 (also named p54), which is a critical component of the P-bodies, and knockdown of DDX6 leads to disassembly of the P-bodies (22,23).…”