Polycomb-Mediated Loss of miR-31 Activates NIK-Dependent NF-κB Pathway in Adult T Cell Leukemia and Other Cancers

SummaryConstitutive NF-κB activation has causative roles in adult T cell leukemia (ATL) caused by HTLV-1 and other cancers. Here, we report a pathway involving Polycomb-mediated miRNA silencing and NF-κB activation. We determine the miRNA signatures and reveal miR-31 loss in primary ATL cells. MiR-31 negatively regulates the noncanonical NF-κB pathway by targeting NF-κB inducing kinase (NIK). Loss of miR-31 therefore triggers oncogenic signaling. In ATL cells, miR-31 level is epigenetically regulated, and aberrant upregulation of Polycomb proteins contribute to miR-31 downregulation in an epigenetic fashion, leading to activation of NF-κB and apoptosis resistance. Furthermore, this emerging circuit operates in other cancers and receptor-initiated NF-κB cascade. Our findings provide a perspective involving the epigenetic program, inflammatory responses, and oncogenic signaling

Plasmid analysis of clinically isolated Enterobacter cloacae in Showa University Hospital

We have reported the possibility of an outbreak of a plasmid-borne carbapenemase-producing Enterobacteriaceae at Showa University Hospital using conjugal transfer experiments; however, we could not perform plasmid profiling and fingerprinting to identify the plasmid responsible for the outbreak in clinical isolates. Therefore, to distinguish whether the appearance of metallo-β-lactamase IMP-11 （blaIMP-11）-producing Enterobacter cloacae （E. cloacae） was due to the same plasmid, we established a plasmid testing system involving plasmid isolation, typing, profiling, and fingerprinting, as well as DNA sequencing analysis of genes surrounding the carbapenemase-encoding gene. Plasmid fingerprinting is an essential tool for identifying plasmids when next-generation sequencing methods cannot be employed. Of note, an important step in fingerprinting is plasmid isolation, which is difficult when large plasmids are involved. In addition, plasmid profiling using S1 nuclease pulse-field gel electrophoresis （PFGE） Southern blotting is an important tool for profiling the size and number of plasmids in bacteria. In this study, we successfully isolated an approximately 90-kb IncL/M plasmid by employing our plasmid analysis system. Importantly, as different blaIMP-11-producing E. cloacae isolates carried the same type of plasmid, with similar size and fingerprinting pattern, we suggest that the isolated IncL/M plasmid is the one present in this strain

T-cell activation via CD26 and caveolin-1 in rheumatoid synovium

CD26 is a T-cell costimulatory molecule with dipeptidyl peptidase IV (DPPIV) activity in its extracellular region. We previously reported that recombinant soluble CD26 enhances peripheral blood T-cell proliferation induced by the recall antigen tetanus toxoid (TT). Recently, we demonstrated that CD26 binds caveolin-1 on antigen-presenting cell (APC), and that residues 201–211 of CD26 along with the serine catalytic site at residue 630, which constitute a pocket structure of CD26/DPPIV, contribute to binding to caveolin-1 scaffolding domain. In addition, following CD26–caveolin-1 interaction on TT-loaded monocytes, caveolin-1 is phosphorylated, with linkage to NF-κB activation, followed by upregulation of CD86. Finally, reduced caveolin-1 expression on APC inhibits CD26-mediated CD86 upregulation and abrogates CD26 effect on TT-induced T-cell proliferation, and immunohistochemical studies revealed an infiltration of CD26+ T cells in the sublining region of rheumatoid synovium and high expression of caveolin-1 in the increased vasculature and synoviocytes of the rheumatoid synovium. Taken together, these results strongly suggest that CD26–cavolin-1 interaction plays a role in the upregulation of CD86 on TT-loaded APC and subsequent engagement with CD28 on T cells, leading to antigen-specific T-cell activation such as the T-cell-mediated antigen-specific response in rheumatoid arthritis

CD26 Mediates Dissociation of Tollip and IRAK-1 from Caveolin-1 and Induces Upregulation of CD86 on Antigen-Presenting Cells

CD26 is a T-cell costimulatory molecule with dipeptidyl peptidase IV enzyme activity in its extracellular region. We have previously reported that the addition of recombinant soluble CD26 resulted in enhanced proliferation of human T lymphocytes induced by the recall antigen tetanus toxoid (TT) via upregulation of CD86 on monocytes and that caveolin-1 was a binding protein of CD26, and the CD26-caveolin-1 interaction resulted in caveolin-1 phosphorylation (p-cav-1) as well as TT-mediated T-cell proliferation. However, the mechanism involved in this immune enhancement has not yet been elucidated. In the present work, we perform experiments to identify the molecular mechanisms by which p-cav-1 leads directly to the upregulation of CD86. Through proteomic analysis, we identify Tollip (Toll-interacting protein) and IRAK-1 (interleukin-1 receptor-associated serine/threonine kinase 1) as caveolin-1-interacting proteins in monocytes. We also demonstrate that following stimulation by exogenous CD26, Tollip and IRAK-1 dissociate from caveolin-1, and IRAK-1 is then phosphorylated in the cytosol, leading to the upregulation of CD86 via activation of NF-κB. Binding of CD26 to caveolin-1 therefore regulates signaling pathways in antigen-presenting cells to induce antigen-specific T-cell proliferation

HIV-1-encoded antisense RNA suppresses viral replication for a prolonged period

<p>Abstract</p> <p>Background</p> <p>Recent evidence proposes a novel concept that mammalian natural antisense RNAs play important roles in cellular homeostasis by regulating the expression of several genes. Identification and characterization of retroviral antisense RNA would provide new insights into mechanisms of replication and pathogenesis. HIV-1 encoded-antisense RNAs have been reported, although their structures and functions remain to be studied. We have tried to identify and characterize antisense RNAs of HIV-1 and their function in viral infection.</p> <p>Results</p> <p>Characterization of transcripts of HEK293T cells that were transiently transfected with an expression plasmid with HIV-1_NL4–3 DNA in the antisense orientation showed that various antisense transcripts can be expressed. By screening and characterizing antisense RNAs in HIV-1_NL4–3-infected cells, we defined the primary structure of a major form of HIV-1 antisense RNAs, which corresponds to a variant of previously reported <it>ASP</it> mRNA. This 2.6 kb RNA was transcribed from the U3 region of the 3′ LTR and terminated at the <it>env</it> region in acutely or chronically infected cell lines and acutely infected human peripheral blood mononuclear cells. Reporter assays clearly demonstrated that the HIV-1 LTR harbours promoter activity in the reverse orientation. Mutation analyses suggested the involvement of NF-κΒ binding sites in the regulation of antisense transcription. The antisense RNA was localized in the nuclei of the infected cells. The expression of this antisense RNA suppressed HIV-1 replication for more than one month. Furthermore, the specific knockdown of this antisense RNA enhanced HIV-1 gene expression and replication.</p> <p>Conclusions</p> <p>The results of the present study identified an accurate structure of the major form of antisense RNAs expressed from the HIV-1_NL4–3 provirus and demonstrated its nuclear localization. Functional studies collectively demonstrated a new role of the antisense RNA in viral replication. Thus, we suggest a novel viral mechanism that self-limits HIV-1 replication and provides new insight into the viral life cycle.</p