A role for Q/N-rich aggregation-prone regions in P-body localization

P-bodies are cytoplasmic foci that are sites of mRNA degradation and translational repression. It is not known what causes the accumulation of RNA degradation factors in P-bodies, although RNA is required. The yeast Lsm1-7p complex is recruited to P-bodies under certain stress conditions. It is required for efficient decapping and degradation of mRNAs, but not for the assembly of P-bodies. Here we show that the Lsm4p subunit and its asparagine-rich carboxy-terminus are prone to aggregation and that this tendency to aggregate promotes efficient accumulation of Lsm1-7p in P-bodies. The presence of Q/N-rich regions in other P-body components suggests a more general role for aggregation-prone residues in P-body localization and assembly. This is supported by reduced P-body accumulation of Ccr4p, Pop2p and Dhh1p after deletion of these domains, and by the observed aggregation of the Q/N-rich region from Ccr4p

Defective removal of ribonucleotides from DNA promotes systemic autoimmunity

Genome integrity is continuously challenged by the DNA damage that arises during normal cell metabolism. Biallelic mutations in the genes encoding the genome surveillance enzyme ribonuclease H2 (RNase H2) cause Aicardi-Goutières syndrome (AGS), a pediatric disorder that shares features with the autoimmune disease systemic lupus erythematosus (SLE). Here we determined that heterozygous parents of AGS patients exhibit an intermediate autoimmune phenotype and demonstrated a genetic association between rare RNASEH2 sequence variants and SLE. Evaluation of patient cells revealed that SLE- and AGS-associated mutations impair RNase H2 function and result in accumulation of ribonucleotides in genomic DNA. The ensuing chronic low level of DNA damage triggered a DNA damage response characterized by constitutive p53 phosphorylation and senescence. Patient fibroblasts exhibited constitutive upregulation of IFN-stimulated genes and an enhanced type I IFN response to the immunostimulatory nucleic acid polyinosinic:polycytidylic acid and UV light irradiation, linking RNase H2 deficiency to potentiation of innate immune signaling. Moreover, UV-induced cyclobutane pyrimidine dimer formation was markedly enhanced in ribonucleotide-containing DNA, providing a mechanism for photosensitivity in RNase H2-associated SLE. Collectively, our findings implicate RNase H2 in the pathogenesis of SLE and suggest a role of DNA damage-associated pathways in the initiation of autoimmunity

Role of p54 RNA Helicase Activity and Its C-terminal Domain in Translational Repression, P-body Localization and Assembly

The RNA helicase p54 (DDX6, Dhh1, Me31B, Cgh-1, RCK) is a prototypic component of P-(rocessing) bodies in cells ranging from yeast to human. Previously, we have shown that it is also a component of the large cytoplasmic polyadenylation element-binding protein translation repressor complex in Xenopus oocytes and that when tethered to the 3′ untranslated region, Xp54 represses reporter mRNA translation. Here, we examine the role of the p54 helicase activity in translational repression and in P-body formation. Mutagenesis of conserved p54 helicase motifs activates translation in the tethered function assay, reduces accumulation of p54 in P-bodies in HeLa cells, and inhibits its capacity to assemble P-bodies in p54-depleted cells. Similar results were obtained in four helicase motifs implicated in ATP binding and in coupling ATPase and RNA binding activities. This is accompanied by changes in the interaction of the mutant p54 with the oocyte repressor complex components. Surprisingly, the C-terminal D2 domain alone is sufficient for translational repression and complete accumulation in P-bodies, although it is deficient for P-body assembly. We propose a novel RNA helicase model, in which the D2 domain acts as a protein binding platform and the ATPase/helicase activity allows protein complex remodeling that dictates the balance between repressors and an activator of translation