Anti-YY1 (aa 221-320) polyclonal antibody

We tested the hypothesis that Let-7d-3p contributes to cardiac cell protection during hypoxic challenge. Myoblast H9c2 cells and primary neonatal rat ventricular cardiomyocytes (NRVM) were transfected with five selected miRNA mimics. Both cell lines were subjected to 0.2% oxygen hypoxia. The protective effects of these miRNAs were determined by assessment of cell metabolic activity by CCK8 assay and measurement of lactate dehydrogenase (LDH) release as a marker of cell injury. Apoptosis and autophagy flux were assessed by Annexin V/7-AAD double staining and the ratio of LC3 II/I with Baf-A1 treatment, an autophagy flux inhibitor, respectively. Luciferase-reporter assay, RT-qPCR and Western blots were performed to identify the changes of relevant gene targets. Among five miRNA mimic transfections, Let-7d-3p increased CCK8 activity, and decreased LDH release in both H9c2 and NRVM during hypoxia. Apoptosis was significantly reduced in H9c2 cells transfected with Let-7d-3p mimic. Autophagy and autophagy flux were not affected. In silico, mRNAs of HMGA2, YY1, KLF9, KLF12, and MEX3C are predicted targets for Let-7d-3p. Luciferase-reporter assay confirmed that Let-7d-3p bound directly to the 3'-UTR region of HMGA2, MEX3C, and YY1, the down-regulations of these mRNAs were verified in both H9c2 and NRVM. The protein expression of HMGA2, but not others, was downregulated in H9c2 and NRVM. It is known that HMGA2 is a strong apoptosis trigger through the blocking of DNA repair. Thus, we speculate that the anti-apoptotic effects of Let-7d-3p mimic during hypoxia challenge are due to direct targeting of HMGA2.

Laser-capture microdissection was used to isolate external germinal layer tissue from three developmental periods of mouse cerebellar development: embryonic days 13, 15, and 18. The cerebellar granule cell-enriched mRNA library was generated with next-generation sequencing using the Helicos technology. Our objective was to discover transcriptional regulators that could be important for the development of cerebellar granule cells-the most numerous neuron in the central nervous system. Through differential expression analysis, we have identified 82 differentially expressed transcription factors (TFs) from a total of 1311 differentially expressed genes. In addition, with TF-binding sequence analysis, we have identified 46 TF candidates that could be key regulators responsible for the variation in the granule cell transcriptome between developmental stages. Altogether, we identified 125 potential TFs (82 from differential expression analysis, 46 from motif analysis with 3 overlaps in the two sets). From this gene set, 37 TFs are considered novel due to the lack of previous knowledge about their roles in cerebellar development. The results from transcriptome-wide analyses were validated with existing online databases, qRT-PCR, and in situ hybridization. This study provides an initial insight into the TFs of cerebellar granule cells that might be important for development and provide valuable information for further functional studies on these transcriptional regulators.