Structural basis of DNA binding by the WhiB-like transcription factor WhiB3 in Mycobacterium tuberculosis
Creators
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1.
University of Nebraska–Lincoln
Abstract
Mycobacterium tuberculosis (Mtb) WhiB3 is an iron–sulfur cluster-containing transcription factor belonging to a subclass of the WhiB-Like (Wbl) family that is widely distributed in the phylum Actinobacteria. WhiB3 plays a crucial role in the survival and pathogenesis of Mtb. It binds to the conserved region 4 of the principal sigma factor (σᴬ₄) in the RNA polymerase holoenzyme to regulate gene expression like other known Wbl proteins in Mtb. However, the structural basis of how WhiB3 coordinates with σᴬ4 to bind DNA and regulate transcription is unclear. Here we determined crystal structures of the WhiB3:σᴬ₄ complex without and with DNA at 1.5 Å and 2.45 Å, respectively, to elucidate how WhiB3 interacts with DNA to regulate gene expression. These structures reveal that the WhiB3:σᴬ₄ complex shares a molecular interface similar to other structurally characterized Wbl proteins and also possesses a subclass-specific Arg-rich DNA-binding motif. We demonstrate that this newly defined Arg-rich motif is required for WhiB3 binding to DNA in vitro and transcriptional regulation in Mycobacterium smegmatis. Together, our study provides empirical evidence of how WhiB3 regulates gene expression in Mtb by partnering with σᴬ₄ and engaging with DNA via the subclass-specific structural motif, distinct from the modes of DNA interaction by WhiB1 and WhiB7.
Additional Information
© 2023 The Authors. Published by Elsevier Inc on behalf of American Society for Biochemistry and Molecular Biology. This is an open access article under the CC BY license. We thank the staff at beamlines 9-2 and 12-2 of SSRL for the assistance during the X-ray diffraction data collection and Dr M. Kato-Maeda for the Mtb H37Rv genomic DNA. This work was supported in part by National Institutes of Health (R35 GM138157-01 to L-M. Z.) and by National Science Foundation CAREER Award (CLP 1846908 to L-M. Z.). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health and the National Science Foundation. Use of the Stanford Synchrotron Radiation Lightsource, SLAC National Accelerator Laboratory, is supported by the U.S. Department of Energy, Office of Science, Office of Basic Energy Sciences under Contract No. DE-AC02-76SF00515. The SSRL Structural Molecular Biology Program is supported by the DOE Office of Biological and Environmental Research, and by the National Institutes of Health, National Institute of General Medical Sciences (including P41GM103393). Data availability: Atomic coordinates and structure factors have been deposited in the RCSB Protein Data Bank (PDB) under the accession codes 8CWT and 8CWR for the WhiB3:σᴬ₄-β_(tip) complex in the P4₃2₁2 form and the R3 form, respectively, and 8CYF for the WhiB3:σᴬ₄-β_(tip):P_(whiB7) complex, respectively. Author contributions: L-M. Z. project administration; L-M. Z. and T. W. methodology; T. W., V. K., M. H., C. J., S. L., and A. S. investigation; X. S. supervision; T. W and L-M. Z. writing-original draft; .T. W., V. K., M. H., C. J., S. L., X. S., L-M. Z., and A. S. writing–review and editing.Attached Files
Published - PIIS0021925823018057.pdf
Supplemental Material - mmc1.docx
Files
PIIS0021925823018057.pdf
Additional details
Identifiers
- PMCID
- PMC10245118
- Eprint ID
- 122315
- Resolver ID
- CaltechAUTHORS:20230717-55375900.16
Funding
- NIH
- R35 GM138157-01
- NSF
- CHE-1846908
- Department of Energy (DOE)
- DE-AC02-76SF00515
- NIH
- P41GM103393
Dates
- Created
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2023-07-25Created from EPrint's datestamp field
- Updated
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2023-07-25Created from EPrint's last_modified field