RESUMEN
Chromatin modification contributes to pluripotency maintenance in embryonic stem cells (ESCs). However, the related mechanisms remain obscure. Here, we show that Npac, a "reader" of histone H3 lysine 36 trimethylation (H3K36me3), is required to maintain mouse ESC (mESC) pluripotency since knockdown of Npac causes mESC differentiation. Depletion of Npac in mouse embryonic fibroblasts (MEFs) inhibits reprogramming efficiency. Furthermore, our chromatin immunoprecipitation followed by sequencing (ChIP-seq) results of Npac reveal that Npac co-localizes with histone H3K36me3 in gene bodies of actively transcribed genes in mESCs. Interestingly, we find that Npac interacts with positive transcription elongation factor b (p-TEFb), Ser2-phosphorylated RNA Pol II (RNA Pol II Ser2P), and Ser5-phosphorylated RNA Pol II (RNA Pol II Ser5P). Furthermore, depletion of Npac disrupts transcriptional elongation of the pluripotency genes Nanog and Rif1. Taken together, we propose that Npac is essential for the transcriptional elongation of pluripotency genes by recruiting p-TEFb and interacting with RNA Pol II Ser2P and Ser5P.
Asunto(s)
Histonas , Células Madre Embrionarias de Ratones , Animales , Cromatina/genética , Fibroblastos/metabolismo , Histonas/metabolismo , Lisina , Ratones , Células Madre Embrionarias de Ratones/metabolismo , Factor B de Elongación Transcripcional Positiva/metabolismo , ARN Polimerasa II/metabolismo , Transcripción GenéticaRESUMEN
Topics on the molecular basis underlying cancer are quite popular among students. Also, excellent textbooks abound that provide interesting materials for discussion during lectures and tutorials about major events leading to cancer formation and progression. However, much less is available for students to conduct experiments for the analysis of cancer samples in undergraduate modules where there is a limited time-frame. Given the difficulty of working with cancer samples and the scarcity of good samples even in the clinical laboratories, it is impossible to run large-class practicals using patients' samples. Here, we describe the use of tissue slides in combination with polymerase-chain reaction (PCR) as a means of simulating an investigative approach to supplement students' learning of clinical research. By using tissue slides for histo-pathological examinations and specific budding yeast genomic DNA and primers adapted to demonstrate methylation-specific PCR, we designed an inquiry-based lab session to simulate the clinical investigation of a cohort of biopsies that students could analyze in a one-session practical.
