Tumor-suppressive miR-145 co-repressed by TCF4-ß-catenin and PRC2 complexes forms double-negative regulation loops with its negative regulators in colorectal cancer.

The frequently dysregulated Wnt/ß-catenin signaling in different malignancies, by activation of its own or orchestration with other co-factors, regulates various oncogenic or tumor-suppressive genes. Among these genes, miRNAs, which are negative posttranscriptional regulators, are also embedded in the Wnt signaling network. Different from the Wnt-induced oncogenic miRNAs, the specific mechanism underlying the Wnt-repressed tumor-suppressive miRNAs is much less understood. In our study, firstly by analyzing a ChIP-seq dataset against TCF4, the core transcription factor for initiation of Wnt signaling in colorectal cancer (CRC) cells, we screened out several tumor-suppressive miRNAs potentially regulated by Wnt signaling. Then through siRNA-mediated knock-down tests and protein and chromatin immunoprecipitations, we found the TCF4-ß-catenin complex can recruit the histone trimethylation complex PRC2 as a co-repressor while binding to the TCF4-binding element (TBE) in the promoter regions of miR-145, miR-132 and miR-212. Thus, upon Wnt signaling activation, the PRC2-mediated trimethylation of histone H3 at lysine 27 increases at these promoter regions, leading to decreased miRNA levels. Furthermore, we found that by targeting TCF4 and SUZ12, the key components of the negative regulation complexes, the tumor-suppressive miR-145 co-repressed by Wnt signaling and histone trimethylation, forms double-negative regulation loops with its negative regulators in CRC cells. And the inverse associations between miR-145 and its targets/negative regulators have also been demonstrated in nude mice and clinical samples. Collectively, we elucidated the detailed molecular mechanism of how dysregulated Wnt/ß-catenin signaling and tumor-suppressive miRNAs reciprocally regulate each other in CRC cells.

Targeting and suppression of HER3-positive breast cancer by T lymphocytes expressing a heregulin chimeric antigen receptor.

Chimeric antigen receptor-modulated T lymphocytes (CAR-T) have emerged as a powerful tool for arousing anticancer immunity. Endogenous ligands for tumor antigen may outperform single-chain variable fragments to serve as a component of CARs with high cancer recognition efficacy and minimized immunogenicity. As heterodimerization and signaling partners for human epidermal growth factor receptor 2 (HER2), HER3/HER4 has been implicated in tumorigenic signaling and therapeutic resistance of breast cancer. In this study, we engineered T cells with a CAR consisting of the extracellular domain of heregulin-1ß (HRG1ß) that is a natural ligand for HER3/HER4, and evaluated the specific cytotoxicity of these CAR-T cells in cultured HER3 positive breast cancer cells and xenograft tumors. Our results showed that HRG1ß-CAR was successfully constructed, and T cells were transduced at a rate of 50%. The CAR-T cells specifically recognized and killed HER3-overexpressing breast cancer cells SK-BR-3 and BT-474 in vitro, and displayed potent tumoricidal effect on SK-BR-3 xenograft tumor models. Our results suggest that HRG1ß-based CAR-T cells effectively suppress breast cancer driven by HER family receptors, and may provide a novel strategy to overcome cancer resistance to HER2-targeted therapy.

MYSM1 Is Essential for Maintaining Hematopoietic Stem Cell (HSC) Quiescence and Survival.

BACKGROUND Histone H2A deubiquitinase MYSM1 has recently been shown to be essential for hematopoiesis and hematopoietic stem cell (HSC) function in both mice and humans. However, conventional MYSM1 knockouts cause partial embryonic lethality and growth retardation, and it is difficult to convincingly remove the effects of environmental factors on HSC differentiation and function. MATERIAL AND METHODS MYSM1 conditional knockout (cKO) mice were efficiently induced by using the Vav1-cre transgenic system. The Vav-Cre MYSM1 cKO mice were then analyzed to verify the intrinsic role of MYSM1 in hematopoietic cells. RESULTS MYSM1 cKO mice were viable and were born at normal litter sizes. At steady state, we observed a defect in hematopoiesis, including reduced bone marrow cellularity and abnormal HSC function. MYSM1 deletion drives HSCs from quiescence into rapid cycling, and MYSM1-deficient HSCs display impaired engraftment. In particular, the immature cycling cKO HSCs have elevated reactive oxygen species (ROS) levels and are prone to apoptosis, resulting in the exhaustion of the stem cell pool during stress response to 5-FU. CONCLUSIONS Our study using MYSM1 cKO mice confirms the important role of MYSM1 in maintaining HSC quiescence and survival.

MiR-200c suppresses TGF-ß signaling and counteracts trastuzumab resistance and metastasis by targeting ZNF217 and ZEB1 in breast cancer.

Resistance to trastuzumab and concomitantly distal metastasis are leading causes of mortality in HER2-positive breast cancers, the molecular basis of which remains largely unknown. Here, we generated trastuzumab-resistant breast cancer cells with increased tumorigenicity and invasiveness compared with parental cells, and observed robust epithelial-mesenchymal transition (EMT) and consistently elevated TGF-ß signaling in these cells. MiR-200c, which was the most significantly downregulated miRNA in trastuzumab-resistant cells, restored trastuzumab sensitivity and suppressed invasion of breast cancer cells by concurrently targeting ZNF217, a transcriptional activator of TGF-ß, and ZEB1, a known mediator of TGF-ß signaling. Given the reported backward inhibition of miR-200c by ZEB1, ZNF217 also exerts a feedback suppression of miR-200c via TGF-ß/ZEB1 signaling. Restoration of miR-200c, silencing of ZEB1 or ZNF217 or blockade of TGF-ß signaling increased trastuzumab sensitivity and suppressed invasiveness of breast cancer cells. Therefore, our study unraveled nested regulatory circuits of miR-200c/ZEB1 and miR-200c/ZNF217/TGF-ß/ZEB1 in synergistically promoting trastuzumab resistance and metastasis of breast cancer cells. These findings provide novel insights into the common role of EMT and related molecular machinery in mediating the malignant phenotypes of breast cancers.

Re-recognition of pseudogenes: From molecular to clinical applications.

Pseudogenes were initially regarded as "nonfunctional" genomic elements that did not have protein-coding abilities due to several endogenous inactivating mutations. Although pseudogenes are widely expressed in prokaryotes and eukaryotes, for decades, they have been largely ignored and classified as gene "junk" or "relics". With the widespread availability of high-throughput sequencing analysis, especially omics technologies, knowledge concerning pseudogenes has substantially increased. Pseudogenes are evolutionarily conserved and derive primarily from a mutation or retrotransposon, conferring the pseudogene with a "gene repository" role to store and expand genetic information. In contrast to previous notions, pseudogenes have a variety of functions at the DNA, RNA and protein levels for broadly participating in gene regulation to influence the development and progression of certain diseases, especially cancer. Indeed, some pseudogenes have been proven to encode proteins, strongly contradicting their "trash" identification, and have been confirmed to have tissue-specific and disease subtype-specific expression, indicating their own value in disease diagnosis. Moreover, pseudogenes have been correlated with the life expectancy of patients and exhibit great potential for future use in disease treatment, suggesting that they are promising biomarkers and therapeutic targets for clinical applications. In this review, we summarize the natural properties, functions, disease involvement and clinical value of pseudogenes. Although our knowledge of pseudogenes remains nascent, this field deserves more attention and deeper exploration.

DNA methylation-regulated and tumor-suppressive roles of miR-487b in colorectal cancer via targeting MYC, SUZ12, and KRAS.

Human colorectal cancer (CRC), characterized by its high morbidity and lethality, seriously threatens human health and lives. MicroRNA-487b (miR-487b) is currently reported to be aberrantly expressed in several tumors, but the detailed functions and underlying mechanisms of miR-487b in CRC remain unclear. Here, we found that miR-487b is downregulated in CRC cell lines and is markedly decreased in tumor specimens derived from CRC patients. MiR-487b inhibits cell proliferation, migration and invasion and promotes the apoptosis of CRC cells in vitro. Statistical analysis of clinical samples indicates that miR-487b may serve as a biomarker for early CRC diagnosis. Inverse correlations between the expression levels of MYC, SUZ12, and KRAS and that of miR-487b exist in vitro and in CRC patient tissue specimens. Further experiments demonstrated the regulatory effects of miR-487b on MYC, SUZ12, and KRAS, and the disruption of these genes partially restores the miR-487b inhibitor-induced phenotype. Additionally, miR-487b promoter region is in a DNA hypermethylated condition and the DNA methyltransferase inhibitor 5-aza-2'-deoxycytidine (5-Aza) increases the levels of miR-487b but suppresses the expression of MYC, SUZ12, and KRAS in a time- and concentration-dependent manner in CRC cells. Collectively, miR-487b is regulated by DNA methylation and it functions as a tumor suppressor in CRC mainly through targeting MYC, SUZ12, and KRAS. Our study provides insight into the regulatory network in CRC cells, offering a new target for treating CRC patients.

CD4+T cell specific B7-H1 selectively inhibits proliferation of naïve T cells and Th17 differentiation in experimental autoimmune encephalomyelitis.

It is widely acknowledged that interleukin 17-producing T helper (Th17) cells are critically participant in the pathogenesis of multiple sclerosis. In the current study, we identified that the expression of CD4+T cells specific co-inhibitory molecule B7-homologue 1(B7-H1) in spleenocytes and mononuclear cells isolated from brains and spinal cord were positive correlated with Th1 and Th17 cells generation and disease severity in experimental autoimmune encephalomyelitis (EAE). Furthermore, B7-H1 transgenic mice developed milder EAE symptoms and fewer Th17 cells than B7-H1 wild type mice. We also found the proliferation of naïve CD4+CD62+T cells isolated from B7-H1 transgenic mice was inhibited. And naïve T cells isolated from B7-H1 transgenic mice produced fewer Th17 cells than WT mice in Th17-polarizing conditions, but the Th1, Th2, and inducible Treg differentiation were the similar in naïve T cells isolated from B7-H1 transgenic mice and WT mice. In conclusion, our study show CD4+T cells specific B7-H1 is a slective inhibitor in proliferation of naïve T cells, Th17 differentiation and pathogenesis of multiple sclerosis.

Epigenetically regulated miR-145 suppresses colon cancer invasion and metastasis by targeting LASP1.

MiR-145 is a tumor-suppressive microRNA that participates in the malignant progression of colorectal cancer (CRC). Although miR-145 has been reported to inhibit proliferation and to induce apoptosis of CRC cells, the reports about its role in invasion and metastasis are controversial. The regulation of miR-145 its own expression also requires further elucidation. In this study, we firstly found that miR-145 is markedly downregulated in the metastatic tumors of CRC patients. Then through gain- and loss-of function studies, we demonstrated that miR-145 suppresses the invasion and metastasis of CRC cells. We also provided experimental evidences which include direct binding assays and verifications on tissue specimens to confirm that LIM and SH3 protein 1 (LASP1) is a direct target of miR-145. Furthermore, we identified the core promoter regions of miR-145 and observed the cooperation between histone methylation and transcription factors through binding to these core promoter regions to regulate the expression of miR-145 in CRC cells. Our study provides an insight into the regulatory network in CRC cells, thus offering new targets for treating CRC patients.