Lott, J. S. | bioRxiv Science

bioRxiv2026

A remarkable case of conserved domain swapping in the COMMD family of proteins

The eukaryotic COMMD family of proteins are core subunits of the Commander protein complex, with a central role in endosomal membrane trafficking and signalling. Previous crystal and cryoEM structures show that COMMD and COMMD-like proteins form homo-oligomeric and hetero-decameric assemblies with a central ring structure formed by their small C-terminal COMM domains. Their -helical N-terminal (HN) domains decorate each side of these rings, and in the eukaryotic Commander complex engage the coiled-coil proteins CCDC22 and CCDC93. Here, we have determined new crystal structures of the isolated HN domains of human Commd4, Commd9 and Commd10, and find that all three proteins form domain swapped structures with a remarkably consistent topology. This occurs via a conformational change in a hinge-loop between helices 2 and 3, leading to exchange of helices 3 to 6 between protomers. The hinge-loops of Commd9 and Commd10 possess several serine and threonine residues that can be phosphorylated, and we find that specific phospho-mimicking mutations in Commd10 can promote or inhibit domain swapping. Whether the unique COMMD HN domains play any roles beyond assembly with CCDC proteins is unclear, but this work suggests a common conformational switch exists with a potential to regulate their function.

biochemistry↗

bioRxiv2025

Ancestral neuronal receptors are bacterial accessory toxins

Symbiosis and horizontal gene transfer events were crucial in the emergence of eukaryotic and multicellular life. A striking example is the acquisition of Teneurins, putative surface-exposed toxins in bacteria that function as cell adhesion receptors in metazoan neuronal development. Here, we demonstrate the evolutionary relationships between metazoan and bacterial Teneurins. We show that bacterial Teneurins are accessory proteins, that harbour diverse encapsulated toxins. They predominate in species with complex social behaviours, suggesting roles in both cell-to-cell competition and cooperation. Metazoan Teneurins are therefore repurposed bacterial toxins central to cell-to-cell communication in nervous systems. Their acquisition was a foundational event in the evolution of metazoans.

biochemistry↗

bioRxiv2021

RNase HI depletion strongly potentiates cell killing by rifampicin in mycobacteria

Multidrug resistant (MDR) tuberculosis (TB) is defined by the resistance of Mycobacterium tuberculosis, the causative organism, to the first-line antibiotics rifampicin and isoniazid. Mitigating or reversing resistance to these drugs offers a means of preserving and extending their use in TB treatment. R-loops are RNA/DNA hybrids that are formed in the genome during transcription, and can be lethal to the cell if not resolved. RNase HI is an enzyme that removes R-loops, and this activity is essential in M. tuberculosis: knockouts of rnhC, the gene encoding RNase HI, are non-viable. This essentiality supports it as a candidate target for the development of new antibiotics. In the model organism Mycolicibacterium smegmatis, RNase HI activity is provided by two RNase HI enzymes, RnhA and RnhC. We show that the partial depletion of RNase HI activity in M. smegmatis, by knocking out either of the genes encoding RnhA or RnhC, led to the accumulation of R-loops. The sensitivity of the knockout strains to the antibiotics moxifloxacin, streptomycin and rifampicin was increased, with sensitivity to the transcriptional inhibitor rifampicin strikingly increased by nearly 100-fold. We also show that R-loop accumulation accompanies partial transcriptional inhibition, suggesting a mechanistic basis for the synergy between RNase HI depletion and transcriptional inhibition. A model of how transcriptional inhibition can potentiate R-loop accumulation is presented. Finally, we identified four small molecules that inhibit recombinant RnhC activity and that also potentiated rifampicin activity in whole-cell assays against M. tuberculosis, supporting an on-target mode of action, and providing the first step in developing a new class of anti-mycobacterial drug. ImportanceThis study validates mycobacterial RNase HI as a druggable, vulnerable candidate for a new therapeutic treatment of M. tuberculosis with a novel mode of action. RNase HI depletion shows synergistic bacterial killing with some current first- and second-line antibiotics, suggesting that RNase HI inhibitors would combine well with these regimens, and could potentially accelerate the clearance of drug-sensitive strains. RNase HI inhibitors also have the potential to reduce the effective dose of rifampicin, with the comcommitant reduction in side effects. The potentiation of rifampicin efficacy conferred by RNase HI deficiency suggests that RNase HI inhibitors may be able to mitigate against development of rifampicin resistance. The synergy may also be able to reverse rifampicin resistance, rescuing this antibiotic for therapy. The surprising finding that low levels of transcriptional inhibition potentiate R-loop formation provides a key new insight into R-loop metabolism.

microbiology↗