Dataset Information

MicroRNA miR-328 regulates zonation morphogenesis by targeting CD44 expression.

ABSTRACT: Morphogenesis is crucial to initiate physiological development and tumor invasion. Here we show that a microRNA controls zonation morphogenesis by targeting hyaluronan receptor CD44. We have developed a novel system to study microRNA functions by generating constructs expressing pre-miRNAs and mature miRNAs. Using this system, we have demonstrated that expression of miR-328 reduced cell adhesion, aggregation, and migration, and regulated formation of capillary structure. Protein analysis indicated that miR-328 repressed CD44 expression. Activities of luciferase constructs harboring the target site in CD44, but not the one containing mutation, were repressed by miR-328. Zonation morphogenesis appeared in cells transfected by miR-328: miR-328-transfected cells were present on the surface of zonating structures while the control cells stayed in the middle. MiR-328-mediated CD44 actions was validated by anti-CD44 antibody, hyaluronidase, CD44 siRNA, and CD44 expression constructs. In vivo experiments showed that CD44-silencing cells appeared as layers on the surfaces of nodules or zonating structures. Immuno-histochemistry also exhibited CD44-negative cells on the surface layers of normal rat livers and the internal zones of Portal veins. Our results demonstrate that miR-328 targets CD44, which is essential in regulating zonation morphogenesis: silencing of CD44 expression is essential in sealing the zonation structures to facilitate their extension and to inhibit complex expansion.

SUBMITTER: Wang CH

PROVIDER: S-EPMC2409976 | biostudies-literature | 2008

REPOSITORIES: biostudies-literature

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MicroRNA miR-328 regulates zonation morphogenesis by targeting CD44 expression.

Wang Chia-Hui CH Lee Daniel Y DY Deng Zhaoqun Z Jeyapalan Zina Z Lee Shao-Chen SC Kahai Shireen S Lu Wei-Yang WY Zhang Yaou Y Yang Burton B BB

PloS one 20080618 6

Morphogenesis is crucial to initiate physiological development and tumor invasion. Here we show that a microRNA controls zonation morphogenesis by targeting hyaluronan receptor CD44. We have developed a novel system to study microRNA functions by generating constructs expressing pre-miRNAs and mature miRNAs. Using this system, we have demonstrated that expression of miR-328 reduced cell adhesion, aggregation, and migration, and regulated formation of capillary structure. Protein analysis indicat ...[more]

PMID: 18560585

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Project description:MicroRNA (miRNA) are non-coding small RNA that regulate gene expression. MiR-328 is reported to influence breast cancer resistance protein (BCRP) expression in cancer cells. As a large inter-individual difference in BCRP levels is observed in various human tissues, the contribution of miR-328 to these differences is of interest. We hypothesized that DNA methylation in the miR-328 promoter region is responsible for the difference in miR-328 levels, leading to inter-individual variability in BCRP levels in human placenta. The association between placental miR-328 and BCRP levels was analyzed, and then DNA methylation in the miR-328 5'-flanking region and regulatory mechanisms causing inter-individual differences in miR-328 and BCRP levels were examined. MiR-328 expression was significantly correlated with BCRP mRNA (Rs = -0.560, P < 0.01) and protein (Rs = -0.730, P < 0.01) levels. It was also up-regulated by the demethylating agent 5-aza-2'-deoxycytidine in BCRP-expressing cells. Luciferase assays with differentially methylated reporter constructs indicated that methylation in the miR-328 5'-flanking region including a predicted CpG island remarkably decreased transcriptional activity compared to that in unmethylated constructs. We selected CCAAT/enhancer binding protein ? (C/EBP?), located within the predicted CpG island, by in silico analysis. To elucidate the role of C/EBP? in miR-328 expression, a chromatin immunoprecipitation assay, promoter deletion analysis, and electrophoretic mobility shift assay (EMSA) were performed. C/EBP?-binding site-truncated constructs showed significantly decreased promoter activity, and EMSA indicated that the C/EBP?-binding sites were located in the CpG island. Finally, the methylation patterns of several CpG dinucleotides proximal to two C/EBP?-binding sites in the miR-328 5'-flanking region were correlated negatively with miR-328 levels, and positively with BCRP levels in human placental samples. These results suggest that methylation patterns in the miR-328 5'-flanking region are involved in the inter-individual difference in BCRP levels in human placenta.

Dataset Information

MicroRNA miR-328 regulates zonation morphogenesis by targeting CD44 expression.

Publications

MicroRNA miR-328 regulates zonation morphogenesis by targeting CD44 expression.

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