XISTexpression in male leukocytes

Abstract

Abstract XISTexpression occurs in human somatic cells possessing more than one X chromosome to balance gene expression and in male germline cells to ensure X chromosome inactivation. A study designed to investigate the sex-biased expression of X-linked immune genes convincingly and repeatedly found low-level XISTexpression in male leukocytes. RNA was extracted from peripheral blood mononuclear cells (PBMC), granulocytes, and oral mucosa from healthy young adults of both sexes (n = 8) by the TRIzol method. After a DNase treatment, RNA was reverse transcribed with a mix of random hexamers and oligodT primers. Real-time PCR was performed in duplicate on 5 ng RNA-equivalent cDNA in duplicate using Taqman primer-probe sets. Negative controls included non-template controls and non-reverse transcribed samples; ref gene, ACTB.Delta Ct between parametric groups were compared by Welch’s t-test. Control genes behaved as expected: CDH1, was expressed higher in mucosa than in leukocytes (P < 0.04); the Y-linked gene TSMB4Ywas expressed in male cells only. Most X-linked immune genes were expressed higher in leukocytes than in mucosa (exception TLR7) with an equal expression between the sexes (0.3< P>0.93) or a sex-biased expression: FOXP3(male-biased, P= 0.04), SH2D1A (female-biased, P= 0.003). Only XISTexpression differed from the expected profile, as it was also found in all male leukocyte samples, both PBMC and granulocytes, but not in mucosa samples. The expression levels in male leukocytes were much lower than in female leukocytes (P< 0.0001). Further investigation on XISTexpression in male leukocytes will focus on chromosomal localization of lncRNA XIST within male nuclei and the life cycle stage of XIST-expressing male leukocytes. Supported by UDEM grant UIN20512