PAF49: An RNA Polymerase I subunit essential for rDNA transcription and stabilization of PAF53

Rachel McNamar, Emma Freeman, Kairo N. Baylor, Aula M. Fakhouri, Sui Huang, Bruce A. Knutson, Lawrence I. Rothblum*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

2 Scopus citations

Abstract

The application of genetic and biochemical techniques in yeast has informed our knowledge of transcription in mammalian cells. Such systems have allowed investigators to determine whether a gene was essential and to determine its function in rDNA transcription. However, there are significant differences in the nature of the transcription factors essential for transcription by Pol I in yeast and mammalian cells, and yeast RNA polymerase I contains 14 subunits while mammalian polymerase contains 13 subunits. We previously reported the adaptation of the auxin-dependent degron that enabled a combination of a “genetics-like” approach and biochemistry to study mammalian rDNA transcription. Using this system, we studied the mammalian orthologue of yeast RPA34.5, PAF49, and found that it is essential for rDNA transcription and cell division. The auxin-induced degradation of PAF49 induced nucleolar stress and the accumulation of P53. Interestingly, the auxin-induced degradation of AID-tagged PAF49 led to the degradation of its binding partner, PAF53, but not vice versa. A similar pattern of co-dependent expression was also found when we studied the non-essential, yeast orthologues. An analysis of the domains of PAF49 that are essential for rDNA transcription demonstrated a requirement for both the dimerization domain and an “arm” of PAF49 that interacts with PolR1B. Further, we demonstrate this interaction can be disrupted to inhibit Pol I transcription in normal and cancer cells which leads to the arrest of normal cells and cancer cell death. In summary, we have shown that both PAF53 and PAF49 are necessary for rDNA transcription and cell growth.

Original languageEnglish (US)
Article number104951
JournalJournal of Biological Chemistry
Volume299
Issue number8
DOIs
StatePublished - Aug 2023

Funding

L. R. was supported by 1P20 GM103639 and OCAST HR 23-155. R. M. was supported by F31CA250352. B. A. K. was supported by NIH NIGMS – 5R01GM141033 . S. H. was supported by U10CA260699. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. R. M. and L. I. R. conceptualization; R. M. L. I. R. B. A. K. A. M. F. K. N. B. S. H. and E. F. investigation; R. M. B. A. K. A. M. F. K. N. B. S. H. and E. F. methodology; B. A. K. A. M. F. and K. N. B. formal analysis; R. M. and L. I. R. writing–original draft; R. M. S. H. B. A. K. and L. I. R. writing–review and editing; R. M. S. H. B. A. K. and L. I. R. funding acquisition. L. R. was supported by 1P20 GM103639 and OCAST HR 23-155. R. M. was supported by F31CA250352. B. A. K. was supported by NIH NIGMS – 5R01GM141033. S. H. was supported by U10CA260699. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

Keywords

  • PAF49
  • PAF53
  • RNA polymerase I (RNAP1)
  • RNA polymerase I subunit E (POLR1E)
  • RNA polymerase I subunit G (POLR1G)
  • SCF (TIR1)
  • auxin
  • conditional protein degradation
  • nucleolus
  • ribosomal RNA
  • transcription

ASJC Scopus subject areas

  • Molecular Biology
  • Biochemistry
  • Cell Biology

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