Abstract

Many non-coding RNAs (ncRNAs) serve as regulatory molecules in various physiological pathways, including gene expression in mammalian cells. Distinct from protein-coding RNA expression, ncRNA expression is regulated solely by transcription and RNA processing/stability. It is thus important to understand transcriptional regulation in ncRNA genes but is yet to be known completely. Previously, we identified that a subset of mammalian ncRNA genes is transcriptionally regulated by RNA polymerase II (Pol II) promoter-proximal pausing and in a tissue-specific manner. In this study, human ncRNA genes that are expressed in the early G1 phase, termed immediate early ncRNA genes, were monitored to assess the function of positive transcription elongation factor b (P-TEFb), a master Pol II pausing regulator for protein-coding genes, in ncRNA transcription. Our findings indicate that the expression of many ncRNA genes is induced in the G0–G1 transition and regulated by P-TEFb. Interestingly, a biphasic characteristic of P-TEFb-dependent transcription of serum responsive ncRNA genes was observed: Pol II carboxyl-terminal domain phosphorylated at serine 2 (S2) was largely increased in the transcription start site (TSS, -300 to +300) whereas overall, it was decreased in the gene body (GB, > +350) upon chemical inhibition of P-TEFb. In addition, the three representative, immediate early ncRNAs, whose expression is dependent on P-TEFb, metastasis-associated lung adenocarcinoma transcript 1 (MALAT1), nuclear enriched abundant transcript 1 (NEAT1), and X-inactive specific transcript (XIST), were further analyzed for determining P-TEFb association. Taken together, our data suggest that transcriptional activation of many human ncRNAs utilizes the pausing and releasing of Pol II, and that the regulatory mechanism of transcriptional elongation in these genes requires the function of P-TEFb. Furthermore, we propose that ncRNA and mRNA transcription are regulated by similar mechanisms while P-TEFb inhibition unexpectedly increases S2 Pol II phosphorylation in the TSSs in many ncRNA genes.One Sentence Summary: P-TEFb regulates Pol II phosphorylation for transcriptional activation in many stimulus-inducible ncRNA genes.

Highlights

  • Gene expression regulation is the most fundamental and crucial event to ensure development, growth, and homeostasis in living organisms

  • A heatmap with the 15,220 non-coding RNAs (ncRNAs) genes generated by subtracting S0 from S15 shows the patterns of phospho-serine 2 (S2) polymerase II (Pol II) occupancy changes in the

  • These data showed that S2 Pol II phosphorylation is up- or down-regulated in a large number of ncRNA genes (53.9%), and a majority of these ncRNA genes are enriched with phospho-S2 Pol II in the early G1 phase

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Summary

Introduction

Gene expression regulation is the most fundamental and crucial event to ensure development, growth, and homeostasis in living organisms. It is estimated that about 70% of the human genome is transcribed and yet only < 2% of the transcripts are mRNAs that are translated into proteins (Fatica and Bozzoni, 2014; Palazzo and Gregory, 2014). This indicates the vastness of non-protein coding RNA genes in mammalian cells. In addition to their large number, more and more non-coding RNAs (ncRNAs) have been found to exert important, diverse cellular functions. Understanding the functions and mechanisms of ncRNAs has become essential in biology and medicine

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