The function of many of the pre-mRNA-binding proteins in mRNA biogenesis is unclear. We have analyzed the biochemical function of the hnRNP K protein by using a mouse cDNA clone. A previous study indicated that the expression of hnRNP K activates c-myc promoter in transient transfection assays. We show that the expression of hnRNP K results in a trans-activation of a variety of RNA polymerase II promoters. The trans-activation function depends on the sequences of hnRNP K that are also necessary for RNA binding. However, the RNA binding motifs are not sufficient for trans-activation. We could identify a mutant that bound RNA in vitro but was impaired in its ability to trans-activate the reporter genes. The trans-activation was not a result of the stabilization of the reporter mRNA, because hnRNP K increased the steady-state level of the reporter mRNA without altering its decay rate. By doing nuclear run-on assays, we provide evidence that the hnRNP K protein trans-activates the reporter genes by increasing the level of transcription.

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Although diverse immunomodulatory reactions of probiotic bacteria have been reported, this effect via Bacillus subtilis natto remains unclear, despite its long consumption history in Japan and usage in Natto production. Hence, we performed a comparative analysis of the immunomodulatory activities of 23 types of B. subtilis natto isolated from Natto products to elucidate the key active components. Among the isolated 23 strains, the supernatant from B. subtilis strain 1 fermented medium showed the highest induction of anti-inflammatory IL-10 and pro-inflammatory IL-12 in THP-1 dendritic cells (THP-1 DC) after co-incubation. We isolated the active component from strain 1 cultured medium and employed DEAE-Sepharose chromatography with 0.5 M NaCl elution for fractionation. IL-10-inducing activity was specific to an approximately 60 kDa protein, GroEL, which was identified as a chaperone protein and was significantly reduced with anti-GroEL antibody. Differential expression analysis of strains 1 and 15, which had the lowest cytokine-producing activity, showed a higher expression of various genes involved in chaperones and sporulation in strain 1. Furthermore, GroEL production was induced in spore-forming medium. The present study is the first to show that the chaperone protein GroEL, secreted by B. subtilis natto during sporulation, plays a crucial role in IL-10 and IL-12 production in THP-1 DC.

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