Effects of the HDACi SAHA on IgH transcription in B-cell splenocytes and effect of deletion of two cis-transcriptional enhancer on IgH transcription in femoral B-cells.

B-cell development is spatially and temporally regulated with the Ig heavy chain (IgH) locus as a conductor. Starting with the first steps of B-cell development, IgH DNA remodeling constantly occurs under the control of several cis-regulatory elements scattered all along the IgH locus. Their individual genomic deletion had partial or no effect on B-cell development. In this study we investigate the effect of the dual deletion of 5’Eµ and 3’RR enhancer on IgH transcription. Numerous B-cell lymphomas feature translocations linking oncogenes with the IgH locus and epigenetic drugs such as histone deacetylase inhibitors (HDACi) have been approved to treat some of them. In this study we investigated IgH locus transcription in B-cell splenocytes stimùulated with LPS and the HDACi SAHA. Femoral B-cells from 5’Eµ/3’RR-deficient mice and RAG2-deficient mice were recovered. RNA was extracted using miRNeasy kit from QIAGEN, according to the manufacturer’s instructions. Two pooled RNAs (each with mice samples) were obtained for the two genotypes. CD43- splenocytes were obtained from wt mice after 48h of in vitro stimulation (1x10^6 cells/ml in RPMI 1640 with 10% FCS) with 5µg/ml LPS ± 200 ng/ml SAHA. RNA was extracted using miRNeasy kit from QIAGEN, according to the manufacturer’s instructions. Two pooled RNAs (each with three samples) were obtained for each stimulatory condition. There are two replicates per condition.