Helicobacter pylori is recognised as a main risk factor for gastric cancer. However, approximately half of the patients with gastritis are negative for H. pylori infection, and the abundance of H. pylori decreases in patients with cancer. In the current study, we profiled gastric epithelium-associated bacterial species in patients with gastritis, intestinal metaplasia, and gastric cancer to identify additional potential pathogenic bacteria. The overall composition of the microbiota was similar between the patients with gastritis and those with intestinal metaplasia. H. pylori was present in half of the non-cancer group, and the dominant bacterial species in the H. pylori-negative patients were Burkholderia, Enterobacter, and Leclercia. The abundance of those bacteria was similar between the cancer and non-cancer groups, whereas the frequency and abundance of H. pylori were significantly lower in the cancer group. Instead, Clostridium, Fusobacterium, and Lactobacillus species were frequently abundant in patients with gastric cancer, demonstrating a gastric cancer-specific bacterial signature. A receiver operating characteristic curve analysis showed that Clostridium colicanis and Fusobacterium nucleatum exhibited a diagnostic ability for gastric cancer. Our findings indicate that the gastric microenvironment is frequently colonised by Clostridium and Fusobacterium in patients with gastric cancer.

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We have utilized Caenorhabditis elegans as a model to investigate the toxicity and underlying mechanism of untranslated CAG repeats in comparison to CUG repeats. Our results indicate that CAG repeats can be toxic at the RNA level in a length-dependent manner, similar to that of CUG repeats. Both CAG and CUG repeats of toxic length form nuclear foci and co-localize with C. elegans muscleblind (CeMBL), implying that CeMBL may play a role in repeat RNA toxicity. Consistently, the phenotypes of worms expressing toxic CAG and CUG repeats, including shortened life span and reduced motility rate, were partially reversed by CeMbl over-expression. These results provide the first experimental evidence to show that the RNA toxicity induced by expanded CAG and CUG repeats can be mediated, at least in part, through the functional alteration of muscleblind in worms.

The multicomponent exon junction complex (EJC) is deposited on the spliced mRNA during pre-mRNA splicing and is implicated in several post-splicing events, including mRNA export, nonsensemediated mRNA decay (NMD), and translation control. This report is the first to identify potential post-translational modifications of the EJC core component Y14. We demonstrate that Y14 is phosphorylated at its repeated arginine/serine (RS) dipeptides, likely by SR protein-specific kinases. Phosphorylation of Y14 abolished its interaction with EJC components as well as factors that function downstream of the EJC. A non-phosphorylatable Y14 mutant was equivalent to the wild-type protein with respect to its association with spliced mRNA and its ability in NMD activation, but the mutant sequestered EJC and NMD factors on ribosome-containing mRNA ribonucleoproteins (mRNPs). We therefore hypothesize that phosphorylation of Y14 occurs upon completion of mRNA surveillance, leading to dissociation of Y14 from ribosome-containing mRNPs. Moreover, we found that Y14 is possibly methylated at multiple arginine residues in the carboxyl-terminal domain and that methylation of Y14 was antagonized by phosphorylation of RS dipeptides. This study reveals antagonistic post-translational modifications of Y14 that may be involved in the remodeling of Y14-containing mRNPs.Eukaryotic mRNAs undergo several processing steps before export to the cytoplasm for translation. The splicing reaction removes introns from precursor mRNAs (pre-mRNAs) 4 and positions the exon junction complex (EJC) on spliced mRNA in a sequence-independent manner (1, 2). The EJC is a dynamic multicomponent complex consisting of a heterodimer of Y14 with Mago and a number of associated factors (1, 2). The EJC may be functionally connected to transcription and acts as an adaptor for recruiting factors involved in the RNA metabolism steps downstream of splicing (3).Previous work suggests that the EJC functions for the nuclear export of spliced mRNAs via the interaction of Y14/Mago, as well as other components, with the mRNA export receptor TAP (4 -6). However, depletion of EJC components only marginally affects bulk poly(A) ϩ RNA export in cultured Drosophila cells (7), suggesting that the EJC may be an accessory factor for mRNA export. On the other hand, Y14/ Mago and RNPS1, another EJC component, directly promote NMDmediated mRNA degradation (8, 9). Y14 interacts with the NMD initiator Upf3 in the nucleus, which subsequently recruits other Upf proteins to yield the active NMD complex (1, 2). Depletion of Y14 abolishes NMD, indicating its essential role in this pathway (10). Recent reports show that the EJC promotes efficient translation by enhancing polysome association with mRNAs (11, 12). In particular, Y14/Mago, as well as RNPS1, is implicated in this translation enhancement (11-13). Thus, the EJC participates in several post-splicing events, including mRNA export/surveillance and translation control (1, 2, 14). The Y14/ Mago heterodimer acts as a core that interacts with sever...

Pnn/DRS protein is associated with desmosomes and colocalizes with splicing factors in nuclear speckled domains. The potential interaction of Pnn with RNPS1, a pre-mRNA splicing factor and a component of the exon-exon junction complex, prompted us to examine whether Pnn is involved in nuclear mRNA processing. By immunoprecipitation, we found that Pnn associates preferentially with mRNAs produced by splicing in vitro. Oligonucleotide-directed RNase H digestion revealed that Pnn binds to the spliced mRNAs at a position immediately upstream of the splice junction and that 5 splice site utilization determines the location of Pnn in alternatively spliced mRNAs. Immunoprecipitation further showed that Pnn binds to mRNAs produced from a transiently expressed reporter in vivo. Although associated with mRNPs, Pnn is a nuclear-restricted protein as revealed by the heterokaryon assay. Overexpression of an amino-terminal fragment of Pnn that directly interacts with RNPS1 leads to blockage of pre-mRNA splicing. However, although suppression of Pnn expression shows no significant effect on splicing, it leads to some extent to nuclear accumulation of bulk poly(A) ؉ RNA. Therefore, Pnn may participate, via its interaction with RNPS1, in mRNA metabolism in the nucleus, including mRNA splicing and export.In eukaryotic cells, the step to remove the intron sequences from the pre-mRNA is carried out by the spliceosome, a macromolecular complex consisting of small nuclear ribonucleoproteins (snRNPs) and a number of protein factors (22). A family of Ser/Arg-dipeptide-rich proteins (SR proteins) play essential roles in constitutive splicing and/or can modulate alternative splice site selection (13). Atypical SR protein RNPS1 was previously characterized with a splicing activity that promotes utilization of distal alternative 3Ј splice sites (32). However, recombinant RNPS1 instead synergizes with prototypical SR proteins to activate both constitutive and alternative pre-mRNA splicing, suggesting the role of RNPS1 as a general splicing activator (32). On the other hand, RNPS1 associates with SAP18 and acinus proteins to form the apoptosis and splicing-associated protein (ASAP) complex, which inhibits in vitro splicing and promotes apoptosis (42). It appears that RNPS1 functions to activate or suppress splicing by forming complexes with different regulatory proteins.During the pre-mRNA splicing process, a multiprotein complex is deposited on the spliced mRNP (24,25). This complex occupies a region 20 to 24 nucleotides (nt) upstream of the splice junctions of mature mRNA and is thus termed the exonexon junction complex (EJC) (25). RNPS1 has been determined as a component of the EJC (25), which is consistent with the observation that RNPS1 specifically associates with spliced mRNAs in vitro (32). The EJC is thought to function as an adaptor platform that provides multiple postsplicing functions (26). This notion is apparently held true in the case of nonsense-mediated decay (NMD), which subjects aberrant mRNAs with premature termination co...