Structural basis of non-canonical transcriptional regulation by the σ(A)-bound iron-sulfur protein WhiB1 in M. tuberculosis

WhiB1 is a monomeric iron–sulfur cluster-containing transcription factor in the WhiB-like family that is widely distributed in actinobacteria including the notoriously persistent pathogen Mycobacterium tuberculosis (M. tuberculosis). WhiB1 plays multiple roles in regulating cell growth and responding to nitric oxide stress in M. tuberculosis, but its underlying mechanism is unclear. Here we report a 1.85 Å-resolution crystal structure of the [4Fe–4S] cluster-bound (holo-) WhiB1 in complex with the C-terminal domain of the σ(70)-family primary sigma factor σ(A) of M. tuberculosis containing the conserved region 4 (σ(A)(4)). Region 4 of the σ(70)-family primary sigma factors is commonly used by transcription factors for gene activation, and holo-WhiB1 has been proposed to activate gene expression via binding to σ(A)(4). The complex structure, however, unexpectedly reveals that the interaction between WhiB1 and σ(A)(4) is dominated by hydrophobic residues in the [4Fe–4S] cluster binding pocket, distinct from previously characterized canonical σ(70)(4)-bound transcription activators. Furthermore, we show that holo-WhiB1 represses transcription by interaction with σ(A)(4)in vitro and that WhiB1 must interact with σ(A)(4) to perform its essential role in supporting cell growth in vivo. Together, these results demonstrate that holo-WhiB1 regulates gene expression by a non-canonical mechanism relative to well-characterized σ(A)(4)-dependent transcription activators.