Cantu Morin, L. M. | Search bioRxiv

bioRxiv2025

A widespread Actinobacterial G Protein System regulates production of specialized metabolites in Streptomyces coelicolor

Actinobacterial G protein systems (AGPSs), also known as conservons, are regulatory systems that are broadly distributed within Actinomycetota. AGPSs are composed of a minimum of four proteins, including a sensor histidine kinase, a small Ras-like GTPase, a roadblock/MglB protein (likely a GTPase activating protein), and protein with a domain of unknown function that likely functions as a guanine-nucleotide exchange factor (GEF). While progress has been made in understanding AGPS function at the mechanistic level, the phylogenetic distribution of individual AGPSs, and the genes and processes they regulate, remain largely unmapped. Previously, the Cvn8 AGPS of Streptomyces coelicolor was found to influence expression of genes in multiple specialized metabolic pathways during interspecies interactions with other actinomycetes. However, the impact of the Cvn8 AGPS on specialized metabolism has not been assessed at the chemical level. Here, we investigated the phylogenetic distribution of the Cvn8 AGPS clade,and assessed the impact of the Cvn8 AGPS on natural product biosynthesis using untargeted metabolomics. In a set of 485 actinobacterial genomes, we found that members of the clade that includes the Cvn8 AGPS from S. coelicolor are widely distributed in the lineages known to produce specialized metabolites. We also found that in S. coelicolor, the pattern of specialized metabolite production varied in mutants lacking specific components of the Cvn8 AGPS. Specifically, normal production of the pigmented antibiotic actinorhodin during interspecies interactions required cvnA8 and cvnF8, while a{Delta} cvnD8 overproduced undecylprodigiosin. Together, these results connect a widespread AGPS to control of specialized metabolism in a model actinomycete. ImportanceActinobacterial G protein systems (AGPSs) are found widely in bacteria in the phylum Actinomycetota, including genera like Streptomyces that produce many useful molecules and pathogens such as Mycobacterium tuberculosis. The genes and functions regulated by these regulatory systems are largely unknown. Here we investigated the role of the Cvn8 AGPS in controlling the production of specialized metabolites, like antibiotics, in the model actinomycete Streptomyces coelicolor. We found that specific components of the Cvn8 AGPS were required for normal production of the antibiotic actinorhodin during interactions between S. coelicolor and another actinomycete. We also show that Cvn8 belongs to a group of AGPSs that is found broadly in the genus Streptomyces and in more distantly related orders of Actinomycetota such as the Pseudonorcardiales and Micromonosporales. Together, these results raise the possibility that this group of AGPSs may influence specialized metabolism across a broad range of Actinomycetota lineages.

microbiology↗

bioRxiv2025

The accessory protein CvnF8 modulates histidine kinase activity in an Actinobacterial G protein system in Streptomyces coelicolor

Conservons are 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 MglA-type roadblock protein (CvnB homolog), a protein containing a domain of unknown function (CvnC homolog), and small Ras-like GTPase (CvnD homolog). Based on their conserved small GTPase components and their phylogenetic distribution, we propose that conservons should be known as Actinobacterial G protein systems (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 expression of multiple pathways for specialized metabolism and that the Cvn8 AGPS 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 serve as a modulator of CvnA8 activity. We found that 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 StatementMany 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, a small Ras-like GTPase, a likely GTPase activating protein, and a protein containing a domain of unknown function. Given this composition, we propose that these regulatory modules 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. Together this work indicates that these CvnF-family accessory proteins likely serve as sensory inputs for AGPSs found broadly in Actinomycetota.

microbiology↗