The accessory protein CvnF8 modulates histidine kinase activity in an Actinobacterial G protein system in <i>Streptomyces coelicolor</i>

ABSTRACT Conservons are operons that encode unusual regulatory systems found in bacteria of the phylum Actinomycetota. These regulatory systems are composed of four core proteins: a sensor histidine kinase-like protein (CvnA homolog), an MglB-type roadblock protein (CvnB homolog), a protein containing a domain of unknown function (CvnC homolog), and a small Ras-like GTPase (CvnD homolog). Based on their conserved small GTPase components and their phylogenetic distribution, we propose that the systems encoded by conservons should be known as a ctinobacterial G p rotein s ystems (AGPSs). The signal transduction path through AGPSs remains poorly understood, and some AGPSs have additional accessory proteins (CvnE and CvnF homologs) of unknown function. In this work, we show that AGPS accessory proteins are present when the cognate histidine kinase protein (CvnA homolog) lacks an extracytoplasmic sensory domain. It was previously shown that the Cvn8 AGPS of Streptomyces coelicolor controls the expression of multiple pathways for specialized metabolism. The Cvn8 AGPS also contains an accessory protein, CvnF8. Through protein modeling, we found that CvnF8 may share an interaction interface with the histidine kinase CvnA8, prompting the hypothesis that CvnF8 may act as a modulator of CvnA8 activity. Consistent with this hypothesis, we found that when co-expressed in a heterologous host, CvnA8 and CvnF8 were purified as a stable complex. In a purified system, CvnF8 strongly stimulated the ATPase activity and autophosphorylation of CvnA8. Taken together, these findings indicate that CvnF family accessory proteins likely serve as sensors and/or modulators of histidine kinases of AGPSs found broadly in Actinomycetota. IMPORTANCE Many lineages of bacteria in the phylum Actinomycetota contain conserved operons (conservons) that encode an unusual type of regulatory system whose function is poorly understood. These lineages include pathogens such as Mycobacterium tuberculosis and members of the genus Streptomyces that produce valuable natural products. These regulatory systems are composed of four proteins, including a sensor histidine kinase and a small Ras-like GTPase. We propose that these regulatory systems be known as actinobacterial G protein systems (AGPSs). We show that some AGPSs include accessory proteins that are only found with partner histidine kinases that lack sensory domains. We demonstrate that one such accessory protein can control the activity of its cognate histidine kinase. Our findings indicate that these CvnF-family accessory proteins likely serve as sensory inputs for AGPSs found broadly in Actinomycetota. This work sheds light on the initial steps of signal transduction within these unusual regulatory systems.

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