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Human RNA polymerase III transcriptomes and relationships to Pol II promoter chromatin and enhancer-binding factors

Nature Structural & Molecular Biology volume 17, pages 620–628 (2010)Cite this article

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Abstract

RNA polymerase (Pol) III transcribes many noncoding RNAs (for example, transfer RNAs) important for translational capacity and other functions. We localized Pol III, alternative TFIIIB complexes (BRF1 or BRF2) and TFIIIC in HeLa cells to determine the Pol III transcriptome, define gene classes and reveal 'TFIIIC-only' sites. Pol III localization in other transformed and primary cell lines reveals previously uncharacterized and cell type–specific Pol III loci as well as one microRNA. Notably, only a fraction of the in silico–predicted Pol III loci are occupied. Many occupied Pol III genes reside within an annotated Pol II promoter. Outside of Pol II promoters, occupied Pol III genes overlap with enhancer-like chromatin and enhancer-binding proteins such as ETS1 and STAT1. Moreover, Pol III occupancy scales with the levels of nearby Pol II, active chromatin and CpG content. These results suggest that active chromatin gates Pol III accessibility to the genome.

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Figure 1: Occupancy analysis of Pol III and associated machinery in HeLa cells.
Figure 2: Differential Pol III occupancy in various cell types.
Figure 3: Genomic features of Pol III–occupied and unoccupied tDNAs in HeLa cells.
Figure 4: Chromatin features at Pol III–bound tDNAs in HeLa cells.
Figure 5: Chromatin marks and factors associated with Pol III–bound regions in Jurkat cells.
Figure 6: Model depicting how chromatin features affect Pol III recruitment at three different regions: promoters, enhancer-like promoters and heterochromatin.

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Acknowledgements

We thank D. Ayer and S. Lessnick at the Huntsman Cancer Institute for cells, B. Dalley for his expertise in Illumina sequencing, D. Nix for suggestions for ChIP-seq analysis and R. Roeder (Rockefeller University) for the gift of anti-RPB90 antibody. Financial support was from the Howard Hughes Medical Institute (HHMI; supplies and genomics resources), the US National Institutes of Health (grants GM38663 to B.J.G., CA42014 to the Huntsman Cancer Institute for support of core facilities and CA63640 to C.H.H.), the Huntsman Cancer Institute and Huntsman Cancer Foundation and the Agilent Technologies Foundation (supplies). B.R.C. is an investigator with the HHMI.

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Authors and Affiliations

  1. Department of Oncological Sciences, Huntsman Cancer Institute, University of Utah School of Medicine, Salt Lake City, Utah, USA

    Andrew J Oler, Ravi K Alla, Peter C Hollenhorst, Katherine J Chandler, Barbara J Graves & Bradley R Cairns

  2. Howard Hughes Medical Institute, University of Utah School of Medicine, Salt Lake City, Utah, USA

    Andrew J Oler, Ravi K Alla & Bradley R Cairns

  3. Agilent Technologies, Life Science Solutions Unit, Santa Clara, California, USA

    Douglas N Roberts & Alexander Wong

  4. Department of Medicine and Experimental Pathology, University of Utah School of Medicine, Salt Lake City, Utah, USA

    Patrick A Cassiday, Cassie A Nelson & Curt H Hagedorn

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Contributions

B.R.C. and A.J.O., overall scope and design; A.J.O., overall experimental execution; D.N.R. and A.W., ChIP-array experiments; C.A.N. and C.H.H., cap-purified RNA; P.A.C., qPCR; P.C.H., K.J.C. and B.J.G., ETS1 and CBP data sets and analysis; A.J.O., R.K.A. and B.R.C., data analysis and interpretation; A.J.O. and R.K.A., figures, tables and data organization. B.R.C. and A.J.O. wrote the manuscript, with comments from all authors.

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Correspondence to Bradley R Cairns.

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Supplementary Figures 1–8 and Supplementary Table 1 (PDF 8603 kb)

Supplementary Data 1

Lists of genes and enriched regions. (XLS 2068 kb)

Supplementary Data 2 (XLS 372 kb)

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Oler, A., Alla, R., Roberts, D. et al. Human RNA polymerase III transcriptomes and relationships to Pol II promoter chromatin and enhancer-binding factors. Nat Struct Mol Biol 17, 620–628 (2010). https://doi.org/10.1038/nsmb.1801

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