PDCD10 (programmed cell death 10, TFAR15), a novel protein associated with cell apoptosis has been recently implicated in mutations associated with Cerebral Cavernous Malformations (CCM). Yeast two-hybrid screening revealed that PDCD10 interacts with MST4, a member of Ste20-related kinases. This interaction was confirmed by coimmunoprecipitation and colocalization assays in mammalian cells. Furthermore, the co-overexpression of PDCD10 and MST4 promoted cell proliferation and transformation via modulation of the extracellular signal-regulated kinase (ERK) pathway. Potent short interfering RNAs (siRNAs) against PDCD10 (siPDCD10) and MST4 (siMST4) were designed to specifically inhibit the expression of PDCD10 and MST4 mRNA, respectively. The induction of siPDCD10 or siMST4 resulted in decreased expression of endogenous PDCD10 or MST4, which was accompanied by reduced ERK activity and attenuated cell growth and anchorage-independent growth. On the other hand, siMST4 had similar effects in PDCD10-overexpressed cells. And more importantly, we confirmed that either overexpressing or endogenous PDCD10 can increase the MST4 kinase activity in vitro. Our results demonstrated that PDCD10 modulation of ERK signaling was mediated by MST4, and PDCD10 could be a regulatory adaptor necessary for MST4 function, suggesting a link between cerebral cavernous malformation pathogenesis and the ERK-MAPK cascade via PDCD10/MST4.

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Purpose: CMTM5 (CKLF-like MARVEL transmembrane domain containing member 5) is located at 14q11.2, a locus associated with multiple cancers. It has six RNA splicing variants with CMTM5-v1as the major one.We explored its expression pattern in normal tissues and tumor cell lines, as well as its functions in carcinoma cells. Experimental Design: We evaluated CMTM5 expression by semiquantitative reverse transcription-PCR (RT-PCR) in normal tissues and carcinoma cell lines of cervical, breast, nasopharyngeal, lung, hepatocellular, esophageal, gastric, colon, and prostate. We further examined CMTM5 promoter methylation in these cell lines. We also analyzed CMTM5 expression after 5-aza-2 ¶-deoxycytidine treatment and genetic demethylation and the functional consequences of restoring CMTM5 in HeLa and PC-3 cells. Results: CMTM5-v1is broadly expressed in human normal adult and fetal tissues, but undetectable or down-regulated in most carcinoma cell lines. Its promoter methylation was detected in virtually all the silenced or down-regulated cell lines. The silencing of CMTM5 could be reversed by pharmacologic demethylation or genetic double-knockout of DNMT1and DNMT3B, indicating methylation-mediated mechanism. Restoration of CMTM5-v1suppressed carcinoma cell proliferation, migration, and invasion. Conclusions: These results indicate that CMTM5 exhibits tumor suppressor activities, but with frequent epigenetic inactivation in carcinoma cell lines.CKLFSF is a novel family of proteins linking chemokines and transmembrane 4 super family (TM4SF). In human, nine genes, CKLF and CKLFSF1 to CKLFSF8, have been discovered. Most CKLFSF genes have different RNA splicing variants, and the protein product of at least one variant of each member has a MARVEL (MAL and related proteins for vesicle trafficking and membrane link) domain. For this reason, in December 2005, CKLFSF1 to CKLFSF8 were renamed as CMTM1 to CMTM8 (CKLF-like MARVEL transmembrane domain containing member 1 to 8). CKLF is the first identified member of this family; with four RNA splicing variants designated CKLF1 to CKLF4. Among them, CKLF1 is more related to chemokine, because it has chemotactic activity on leukocytes with CCR4 as one of its functional receptors (1, 2). CMTM1 to CMTM8 were identified based on the cDNA and amino acid sequences of CKLF2 that is the full-length cDNA product of CKLF. Of them, CMTM1 and CMTM2 are more related to chemokine, whereas CMTM8 is more related to TM4SF11, with 39% sequence homology at the amino acid level. The characteristics of other CMTM members are intermediate between CMTM1 and CMTM8 (3).Generally, TM4SF includes several types of proteins possessing the four transmembrane-helix structure, such as typical TM4SF (tetraspanin) and MARVEL domain -containing proteins. Tetraspanin-like proteins KAI1/CD82 and CD63 contain a larger second outer loop (EC2) and four to six conserved extracellular cysteine residues. Whereas the MARVEL domaincontaining proteins, including MAL, physins, gyrins, and occludin families, lack the descr...

MicroRNAs (miRs) play an essential role in the regulation of bone formation and homeostasis. miR-185 has been reported to negatively regulate osteogenesis in vitro. However, whether it has an impact on in vivo bone homeostasis remains unknown. Here, we demonstrated that primary osteoblasts and mesenchymal stem cells derived from miR-185-knockout (KO) mice exhibited enhanced osteogenesis. Further, we constructed an ovariectomized mouse model to investigate the role of miR-185 during osteoporosis. Micro-computed tomography revealed an increased bone volume in KO compared to wild-type mice 6 weeks after surgery, indicating redundant bone formation after miR-185 depletion. Dual-luciferase reporter assays identified biglycan (Bgn), which promotes bone formation through the BMP/Smad pathway, as the direct target of miR-185. Taken together, these findings indicate that blocking miR-185 expression increases bone formation during osteoporosis, which may partly occur through the regulation of Bgn expression and BMP/Smad signaling.

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Using cDNA microarrays we determined the gene expression patterns in the human acute promyelocytic leukemia (APL) cell line NB4 during all-trans retinoic acid (ATRA)-induced differentiation. We analyzed the expression of 12,288 genes in the NB4 cells after 12 hours, 24 hours, 48 hours, 72 hours, and 96 hours of ATRA exposure. During this time course, we found 168 up-regulated and more than 179 down-regulated genes, most of which have not been reported before. Many of the altered genes encode products that participate in signaling pathways, cell differentiation, programmed cell death, transcription regulation, and production of cytokines and chemokines. Of interest, the CD52 and protein kinase A regulatory subunit ␣ (PKA-Rl␣) genes, whose products are being used as therapeutic targets for certain human neoplasias in currently ongoing clinical trials, were among the genes observed to be markedly up-regulated after ATRA treatment. The present study provides valuable data to further understand the mechanism of ATRAinduced APL cell differentiation and suggests potential therapeutic alternatives for this leukemia. Acute promyelocytic leukemia (APL) is a subtype of acute myeloid leukemia characterized by the accumulation of cells arrested at the promyelocytic stage of myeloid differentiation. This leukemia exhibits a specific chromosomal translocation t(15;17) involving the promyelocytic leukemia (PML) gene locus on chromosome 15, and the retinoid acid receptor ␣ (RAR␣) locus on chromosome 17. This translocation generates a chimeric fusion gene PML-RAR␣, which encodes a protein that functions as an aberrant nuclear receptor considered to be the cause of APL.