The heterodimeric structure of heterogeneous nuclear ribonucleoprotein C1/C2 dictates 1,25-dihydroxyvitamin D-directed transcriptional events in osteoblasts

Heterogeneous nuclear ribonucleoprotein （hnRNP） C plays a key role in RNA processing but also exerts a dominant negative effect on responses to 1,25-dihydroxyvitamin D （1,25（OH）2D） by functioning as a vitamin D response element-binding protein （VDRE-BP）. hnRNPC acts a tetramer of hnRNPC1 （huC1） and hnRNPC2 （huC2）, and organization of these subunits is critical to in vivo nucleic acid-binding. Overexpression of either huC1 or huC2 in human osteoblasts is sufficient to confer VDRE-BP suppression of 1,25（OH）2D-mediated transcription. However, huC1 or huC2 alone did not suppress 1,25（OH）2D-induced transcription in mouse osteoblastic cells. By contrast, overexpression of huC1 and huC2 in combination or transfection with a bone-specific polycistronic vector using a ＂self-cleaving＂ 2A peptide to co-express huC1/C2 suppressed 1,25D-mediated induction of osteoblast target gene expression. Structural diversity of hnRNPC between human/NWPs and mouse/rat/rabbit/dog was investigated by analysis of sequence variations within the hnRNP CLZ domain. The predicted loss of distal helical function in hnRNPC from lower species provides an explanation for the altered interaction between huC1/C2 and their mouse counterparts. These data provide new evidence of a role for hnRNPC1/C2 in 1,25（OH）2D-driven gene expression, and further suggest that species-specific tetramerization is a crucial determinant of its actions as a regulator of VDR-directed transactivation.

A candidate protective factor in amyotrophic lateral sclerosis:heterogenous nuclear ribonucleoprotein G

Heterogenous nuclear ribonucleoprotein G is down-regulated in the spinal cord of the Tg(SOD1*G93A)1Gur(TG)amyotrophic lateral sclerosis mouse model.However,most studies have only examined heterogenous nuclear ribonucleoprotein G expression in the amyotrophic lateral sclerosis model and heterogenous nuclear ribonucleoprotein G effects in amyotrophic lateral sclerosis pathogenesis such as in apoptosis are unknown.In this study,we studied the potential mechanism of heterogenous nuclear ribonucleoprotein G in neuronal death in the spinal cord of TG and wild-type mice and examined the mechanism by which heterogenous nuclear ribonucleoprotein G induces apoptosis.Heterogenous nuclear ribonucleoprotein G in spinal cord was analyzed using immunohistochemistry and western blotting,and cell proliferation and proteins(TAR DNA binding protein 43,superoxide dismutase 1,and Bax)were detected by the Cell Counting Kit-8 and western blot analysis in heterogenous nuclear ribonucleoprotein G siRNA-transfected PC12 cells.We analyzed heterogenous nuclear ribonucleoprotein G distribution in spinal cord in the amyotrophic lateral sclerosis model at various time points and the expressions of apoptosis and proliferation-related proteins.Heterogenous nuclear ribonucleoprotein G was mainly localized in neurons.Amyotrophic lateral sclerosis mice were examined at three stages:preonset(60-70 days),onset(90-100 days)and progression(120-130 days).The number of heterogenous nuclear ribonucleoprotein G-positive cells was significantly higher in the anterior horn of the lumbar spinal cord segment of TG mice at the preonset stage than that of control group but lower than that of the control group at the onset stage.The number of heterogenous nuclear ribonucleoprotein G-positive cells in both central canal and surrounding gray matter of the whole spinal cord of TG mice at the onset stage was significantly lower than that in the control group,whereas that of the lumbar spinal cord segment of TG mice was significantly higher than that in the control group at preonset stage and significantly lower than that in the control group at the progression stage.The numbers of heterogenous nuclear ribonucleoprotein G-positive cells in the posterior horn of cervical and thoracic segments of TG mice at preonset and progression stages were significantly lower than those in the control group.The expression of heterogenous nuclear ribonucleoprotein G in the cervical spinal cord segment of TG mice was significantly higher than that in the control group at the preonset stage but significantly lower at the progression stage.The expression of heterogenous nuclear ribonucleoprotein G in the thoracic spinal cord segment of TG mice was significantly increased at the preonset stage,significantly decreased at the onset stage,and significantly increased at the progression stage compared with the control group.heterogenous nuclear ribonucleoprotein G expression in the lumbar spinal cord segment of TG mice was significantly lower than that of the control group at the progression stage.After heterogenous nuclear ribonucleoprotein G gene silencing,PC12 cell survival was lower than that of control cells.Both TAR DNA binding protein 43 and Bax expressions were significantly increased in heterogenous nuclear ribonucleoprotein G-silenced cells compared with control cells.Our study suggests that abnormal distribution and expression of heterogenous nuclear ribonucleoprotein G might play a protective effect in amyotrophic lateral sclerosis development via preventing neuronal death by reducing abnormal TAR DNA binding protein 43 generation in the spinal cord.

Functional foods effective for hepatitis C: Identification of oligomeric proanthocyanidin and its action mechanism

Hepatitis C virus(HCV)is a major cause of viral hepatitis and currently infects approximately 170 million people worldwide.An infection by HCV causes high rates of chronic hepatitis(】75%)and progresses to liver cirrhosis and hepatocellular carcinoma ultimately.HCV can be eliminated by a combination of pegylatedα-interferon and the broad-spectrum antiviral drug ribavirin;however,this treatment is still associated with poor efficacy and tolerability and is often accompanied by serious side-effects.While some novel direct-actingantivirals against HCV have been developed recently,high medical costs limit the access to the therapy in cost-sensitive countries.To search for new natural anti-HCV agents,we screened local agricultural products for their suppressive activities against HCV replication using the HCV replicon cell system in vitro.We found a potent inhibitor of HCV RNA expression in the extracts of blueberry leaves and then identified oligomeric proanthocyanidin as the active ingredient.Further investigations into the action mechanism of oligomeric proanthocyanidin suggested that it is an inhibitor of heterogeneous nuclear ribonucleoproteins(hn RNPs)such as hn RNP A2/B1.In this review,we presented an overview of functional foods and ingredients efficient for HCV infection,the chemical structural characteristics of oligomeric proanthocyanidin,and its action mechanism.

Significance of the antigenic epitopes in heterogenous nuclear ribonucleoproteins-Ⅰfor the diagnosis and prognosis of systematic sclerosis

To assess the presence of autoantibodies against epitopes of heterogenous nuclear ribonucleoprotein- Ⅰ （hnRNP- Ⅰ ） in systematic sclerosis （SSc） and to analyze their clinical significance, polypeptides of hnRNP- Ⅰ were designed by biological technical software and analyzed with both the Wonderful Biology Information System and DNA Star-Protean Analysis Software at the same time. In these ways, two polypeptides of hnRNP- Ⅰ were obtained based on their amino acid sequences, folding features, hydrophilic, curl style, dough kneading sensation and the possibility on the surface of proteins. They are named as hnRNP- Ⅰ -1 （ NVKYNNDKSRDYTRPDLPSGDSQPSLDQT, 264-292 aa） and hnRNP- Ⅰ -2 （QLP4REGQEDQGLTKDYGNSOL, 441-461 aa）, simply designated as Ⅰ -1 and Ⅰ -2. The autoantibodies against hnRNPs were detected by means of ELISA using the synthetic epitopes polypeptides as antigen. It was found that the positive rate of detection for anti- Ⅰ -1 and anti- Ⅰ -2 autoantibodies were rather higher in SSc patients than that in other CTDs and the sensitivities and specificities of the testing with ELISA for anti- Ⅰ -1 and anti- Ⅰ -2 antibodies in SSc patients were 47.62%/93.43% and 38.1%/ 91.08%, without any significant difference between these two groups of testings. Also, there was no significant difference in the clinical features and laboratory findings, such as age, involvements in digestive and respiratory tracts and erythrocyte sedimentation rate etc., between the anti- Ⅰ -Ⅰ-positive and -negative groups in SSc patients. However, the hnRNP- Ⅰ -autoantibody-positive group of patients had obviously shorter duration of disease course compared with that of the autoantibody-negative group. Anti- Ⅰ -1 and anti- Ⅰ -2 autoantibodies also had no association with antinuclear antibody, anti-Sc170 and anti-centromere antibody （ACA） in SSc patients. So, it is apparent that the autoantibodies related with SSc may act through different pathways in the pathogenesis of SSc, and the hnRNP- Ⅰ autoantibody is a new type antibody occuring during the early stage of disease and appearing to have diagnostic and prognostic significances.

Overexpression of c-Myc-dependent heterogeneous nuclear ribonucleoprotein A1 promotes proliferation and inhibits apoptosis in NOTCH1-mutated chronic lymphocytic leukemia cells

Background: NOTCH1 mutation is an essential molecular biologic aberration in chronic lymphocytic leukemia (CLL). CLL patients withNOTCH1 mutation have shown an unfavorable survival and a poor response to chemoimmunotherapy. This study aims to present the mechanisms of adverse prognosis caused byNOTCH1 mutation from the perspective of the splicing factor heterogeneous nuclear ribonucleoprotein A1 (hnRNPA1).Methods: The microarray data in Gene Expression Omnibus datasets were analyzed by bioinformatics and the function of hnRNPA1 was checked by testing the proliferation and apoptosis of CLL-like cell lines. Afterward, quantitative reverse transcription-polymerase chain reaction and Western blotting were applied to explore the relationship among NOTCH1, c-Myc, and hnRNPA1.Results: RNA splicing was found to play a vital part inNOTCH1-mutated CLL cells;hence, hnRNPA1 was selected as the focus of this study. Higher expression of hnRNPA1 validated in primaryNOTCH1-mutated CLL samples could promote proliferation and inhibit apoptosis in CLL. The expression of hnRNPA1 increased when NOTCH1 signaling was activated by transfection with NOTCH1 intracellular domain (NICD)-overexpressed adenovirus vector and declined after NOTCH1 signaling was inhibited by NOTCH1-shRNA. Higher expression of c-Myc was observed in NICD-overexpressed cells and hnRNPA1 expression was downregulated after applying c-Myc inhibitor 10058-F4. Moreover, in NICD-overexpressed cells, hnRNPA1 expression decreased through c-Myc inhibition.Conclusion: Overexpression of c-Myc-dependent hnRNPA1 could promote proliferation and inhibit apoptosis inNOTCH1- mutated CLL cells, which might partly account for the poor prognosis of patients withNOTCH1 mutation.

Characterization of the protein expression and localization of hnRNP family members during murine spermatogenesis

Mammalian testis exhibits remarkably high transcriptome complexity,and spermatogenesis undergoes two periods of transcriptional cessation.These make the RNA-binding proteins(RBPs)the utmost importance during male germ cell development.Heterogeneous nuclear ribonucleoproteins(hnRNPs)are a large family of RBPs implicated in many steps of RNA processing;however,their roles in spermatogenesis are largely unknown.Here,we investigated the expression pattern of 12 hnRNP family members in mouse testes and found that most detected members are highly expressed in the testis.Furthermore,we found that most of the detected hnRNP proteins(hnRNPD,hnRNPK,hnRNPQ,hnRNPU,and hnRNPUL1)display the highest signals in the nuclei of pachytene spermatocytes,round spermatids,and Sertoli cells,whereas hnRNPE1 exclusively concentrates in the manchette of elongating spermatids.The expression of these hnRNP proteins showed both similarities and specificity,suggesting their diverse roles in spermatogenesis.