Expression of Ribosomal RNA and Protein Genes in Human Embryonic Stem Cells Is Associated With the Activating H3K4me3 Histone Mark.

Embryonic stem cells (ESCs) exhibit unrestricted and indefinite, but stringently controlled, proliferation, and can differentiate into any lineage in the body. In the current study, we test the hypothesis that expression of ribosomal RNA (rRNA) and ribosomal protein genes (RPGs) contribute to the ability of hESCs to proliferate indefinitely. Consistent with the accelerated growth rate of hESCs, we find that hESC lines H1 and H9 both exhibit significantly higher levels of rRNA when compared to a panel of normal and cancer human cell lines. Although many RPGs are expressed at levels that comparable to other human cell lines, a few RPGs also exhibit higher expression levels. In situ nuclear run-on assays reveal that both nucleoli in hESCs actively transcribe nascent rRNA. Employing genome-wide chromatin immunoprecipitation-deep sequencing and bioinformatics approaches, we discovered that, RPGs are dominantly marked by the activating H3K4me3 histone mark in the G1, M, and G2 phases of the cell cycle. Interestingly, the rDNA repeats are marked by the activating H3K4me3 only in the M phase, and repressive H3K27me3 histone mark in all three cell cycle phases. Bioinformatics analyses also reveal that Myc, a known regulator of cell growth and proliferation, occupies both the rRNA genes and RPGs. Functionally, down-regulation of Myc expression by siRNA results in a concomitant decrease in rRNA levels. Together, our results show that expression of rRNA, which is regulated by the Myc pluripotency transcription factor, and of RPGs in hESCs is associated with the activating H3K4me3 modification. J. Cell. Physiol. 231: 2007-2013, 2016.