Sertoli cell­specific coxsackievirus and adenovirus receptor regulates cell adhesion and gene transcription via ß-catenin inactivation and Cdc42 activation.

Blood-testis barrier (BTB) and apical ectoplasmic specialization (ES) serve as structural supports for germ cell (GC) development. We demonstrated that the Sertoli cell (SC)-specific coxsackievirus and adenovirus receptor (CXADR) knockout (SC-CXADR-/-), but not the GC-specific knockout, impaired spermatogenesis. An increase in GC apoptosis and premature loss of elongated spermatids were observed in SC-CXADR-/- testes. The BTB function was compromised in SC-CXADR-/- testes with dysregulation of oocludin and zonula occludens-1 expression at the basal compartment of the seminiferous epithelium. An integrated omics analyses confirmed that altered gene ontology terms identified in SC-CXADR-/- testes are highly associated with spermatid development and differentiation, spermatogenesis, and sperm motility and are considered as unique testicular function terms. Leptin, Nasp, Tektin3, Larp 7, and acrosin, which are highly associated with male fertility, were found to be down-regulated in SC-CXADR-/- testes. Based on the data from the omics analyses, we employed the CXADR-deficient SC model to further investigate the molecular mechanisms involved. We unraveled that SC-CXADRs are required for ß-catenin inactivation and cell division cycle protein 42 (Cdc42) activation, resulting in maintaining the integrity and function of the BTB and apical ES as well as inhibiting gene transcription, such as the Myc gene, in the testes. We demonstrated for the first time that CXADR is an important mediator governing ß-catenin and Cdc42 signaling that is essential for spermatogenesis. The molecular mechanisms identified herein may provide new insights to unravel the novel functions and signaling cascades of CXADR in other key CXADR-expressing tissues.-Huang, K., Ru, B., Zhang, Y., Chan, W.-L., Chow, S.-C., Zhang, J., Lo, C., Lui, W.-Y. Sertoli cell-specific coxsackievirus and adenovirus receptor knockout regulates cell adhesion and gene transcription via ß-catenin inactivation and Cdc42 activation.

A potential antitumor ellagitannin, davidiin, inhibited hepatocellular tumor growth by targeting EZH2.

Hepatocellular carcinoma (HCC) is the fifth most common cancer worldwide and is the third most common cause of cancer-related deaths. Currently available treatment options for HCC patients are scarce resulting in an urgent need to develop a novel effective cure. Polygonum capitatum is a medicinal herb which has been used to treat inflammatory diseases in Miao nationality of China. We recently isolated a pure compound davidiin from P. capitatum extract. Four HCC cell lines were treated with davidiin. Cell viability was recorded by MTT assay. siRNAs targeting enhancer of zeste homolog 2 (EZH2) were applied to modulate the expression of EZH2. Established xenograft mice models of HCC were applied to evaluate the in vivo anticancer activity of davidiin. We investigated the anticancer activity and the underlying mechanism of davidiin. The compound inhibited HCC cell growth and also suppressed tumor growth in xenografted HCC mouse. Such inhibition was facilitated by specifically downregulation on EZH2. The compound possesses anticancer activity both in vitro and in vivo which warrants further clinical investigation as a potential anti-HCC agent.

Transcriptomic and iTRAQ proteomic approaches reveal novel short-term hyperosmotic stress responsive proteins in the gill of the Japanese eel (Anguilla japonica).

Osmoregulation is critical for the survival of fishes that migrate between freshwater (FW) and seawater (SW). The eel, as a catadromous fish, has been studied for decades to reveal the mechanisms of osmoregulation. These studies, however, have been limited by the lack of a genomic database to decipher the mechanism of osmoregulation at a molecular level. In this study, using high-throughput transcriptomic and proteomic technologies, we have provided the first genome-wide study to identify hyperosmotic responsive proteins in the gills of the Japanese eel. Deep sequencing using the 454 platform produced over 660,000 reads with a mean length of 385 bp. For the proteomic study, we collected gill samples from three different treatment groups of fish that had fully adapted to FW/SW or were transferred from FW to SW for 6h. The respective group of gill proteins were extracted and labeled using an isobaric tag for relative and absolute quantitation (iTRAQ) using LTQ-Orbitrap, a high resolution mass spectrometer. Among the 1519 proteins identified from the gill samples, 96 proteins were differentially expressed between FW and SW adapted fish. Nineteen hyperosmotic responsive proteins were detected (10 up-regulated and 9 down-regulated proteins) after 6h post FW to SW transfer. BIOLOGICAL SIGNIFICANCE: The study has provided the most comprehensive, targeted investigation of eel gill proteins to date, and shown the powerfulness of combining transcriptomic and proteomic approaches to provide molecular insights of osmoregulation mechanisms in a non-model organism, eel.

Enhancer of zeste homolog 2 silences microRNA-218 in human pancreatic ductal adenocarcinoma cells by inducing formation of heterochromatin.

BACKGROUND & AIMS: Enhancer of zeste homolog 2 (EZH2) is a histone methyltransferase that is overexpressed by pancreatic ductal adenocarcinoma (PDAC) cells and increases their aggressiveness. We identified microRNAs (miRs) that are regulated by EZH2 and studied their functions in PDAC cells. METHODS: We performed miR profile analysis of PDAC cells incubated with EZH2 inhibitor 3-deazaneplanocin A, and pancreatic ductal epithelial cells that overexpressed EZH2. Expression levels of miRs and the targets of miRs were analyzed by quantitative reverse transcription polymerase chain reaction and immunohistochemistry. We expressed different forms of EZH2 to analyze functional domains and used small interfering RNAs to reduce its level in PDAC cells. RESULTS: Expression of miR-218 was repressed by EZH2 in PDAC cells. Levels of miR-218 were significantly reduced in primary PDAC tumor samples compared with paired, adjacent nontumor tissue. Overexpression of miR-218 in SW1990 cells reduced their proliferation and tumor formation and metastasis in nude mice. Loss of miR-218 from SW1990 cells increased levels of UDP-glycosyltransferase 8 and miR-218 was found to bind to its 3'-UTR. Levels of UDP-glycosyltransferase protein and messenger RNA were associated with the metastatic potential of PDAC cell lines and progression of tumors in patients. EZH2 was found to silence miR-218 by binding to its promoter, promoting heterochromatin formation, and recruiting the DNAs methyltransferase 1, 3A, and 3B. CONCLUSIONS: EZH2 is up-regulated in PDAC samples from patients and silences miR-218. MicroRNA-218 prevents proliferation of PDAC cells in culture, and tumor growth and metastasis in nude mice. MicroRNA-218 reduces levels of UDP-glycosyltransferase, which is associated with the metastatic potential of PDAC tumors in mice and progression of human PDAC.