Domains within RbpA serve specific functional roles that regulate the expression of distinct mycobacterial gene subsets

Jerome Prusa, Drake Jensen, Gustavo Santiago-Collazo, Steven S. Pope, Ashley L. Garner, Justin J. Miller, Ana Ruiz Manzano, Eric A. Galburt, Christina L. Stallings

Research output: Contribution to journalArticlepeer-review

7 Scopus citations

Abstract

The RNA polymerase (RNAP) binding protein A (RbpA) contributes to the formation of stable RNAP-promoter open complexes (RPo) and is essential for viability in mycobacteria. Four domains have been identified in the RbpA protein, i.e., an N-terminal tail (NTT) that interacts with RNAP β' and σ subunits, a core domain (CD) that contacts the RNAP β' subunit, a basic linker (BL) that binds DNA, and a σ-interaction domain (SID) that binds group I and group II σ factors. Limited in vivo studies have been performed in mycobacteria, however, and how individual structural domains of RbpA contribute to RbpA function and mycobacterial gene expression remains mostly unknown. We investigated the roles of the RbpA structural domains in mycobacteria using a panel of rbpA mutants that target individual RbpA domains. The function of each RbpA domain was required for Mycobacterium tuberculosis viability and optimal growth in Mycobacterium smegmatis. We determined that the RbpA SID is both necessary and sufficient for RbpA interaction with the RNAP, indicating that the primary functions of the NTT and CD are not solely association with the RNAP. We show that the RbpA BL and SID are required for RPo stabilization in vitro, while the NTT and CD antagonize this activity. Finally, RNA-sequencing analyses suggest that the NTT and CD broadly activate gene expression, whereas the BL and SID activate or repress gene expression in a genedependent manner for a subset of mycobacterial genes. Our findings highlight specific outcomes for the activities of the individual functional domains in RbpA.

Original languageEnglish
Article numbere00690-17
JournalJournal of bacteriology
Volume200
Issue number13
DOIs
StatePublished - Jul 1 2018

Keywords

  • Eubacteria
  • Mycobacterium
  • RNA polymerases
  • RbpA
  • Transcription
  • Transcriptional regulation

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