Aug 2026· Current Biology· 0 citations· 55 references
Medicine
TL;DR
It is proposed that MAIN domains allow microbes to release proteins from their cell surface in response to shear force, enabling broader nutrient scavenging, intoxication of neighboring cells, and dispersal through surface detachment.
Abstract
Adhesion G protein-coupled receptors (aGPCRs) transduce mechanical stimuli across the cytoplasmic membrane in eukaryotes. These receptors contain extracellular GPCR autoproteolysis-inducing (GAIN) domains that undergo autoproteolysis but maintain stable associations of their cleavage products. A diverse set of adhesion domains appended to the GAIN domain binds surface ligands on neighboring cells or the extracellular matrix. Shear force is thought to disrupt the interaction between the cleavage products, exposing a tethered agonist that triggers GPCR signaling. Here, we report that proteins with structural homology to GAIN domains are broadly conserved among bacteria and archaea. The microbial domains lack strong sequence conservation to their eukaryotic counterparts but are predicted to adopt a similar fold. We demonstrate that these microbial autoproteolysis-inducing (MAIN) domains undergo autoproteolysis both in vitro and in vivo, using conserved catalytic residues. Furthermore, proteolysis occurs in a conserved β turn that allows stable non-covalent interaction between the cleavage products. MAIN domains are tethered to the cell envelope of bacteria and archaea and are fused to diverse sets of adhesion and enzymatic domains. Many of the same adhesion domains are appended to both MAIN and GAIN domains, suggesting these protein families share a common origin and function. We propose that MAIN domains allow microbes to release proteins from their cell surface in response to shear force, enabling broader nutrient scavenging, intoxication of neighboring cells, and dispersal through surface detachment.
Bacteroidales secreted antimicrobial proteins (BSAPs) are diffusible MACPF-domain toxins that mediate intra-species antagonism in the gut microbiota. Here we define the mechanism of action of BSAP-1 from Bacteroides fragilis, showing how target specificity encoded within the N- and C-terminal domains is coordinated with pore-forming activity of the MACPF. We show that specificity of the toxin for its receptor is mediated by an extended interface comprised of the BSAP-1 C-terminal domain and residues on the receptor that differ from the orthologous protein of BSAP-1 producing strains. On the surface of susceptible cells, BSAP-1 undergoes proteolytic cleavage of an N-terminal regulatory domain, triggering its assembly into oligomeric pores. Cryo-electron microscopy of membrane-inserted BSAP-1 reveals a 13-subunit transmembrane β-barrel pore formed through canonical MACPF rearrangements. Comparative modelling supports a conserved oligomerization mechanism across the BSAP family despite diversification of receptor-binding domains that target either proteins or glycan receptors. Together, these findings establish BSAP-1 as a receptor-targeted, protease-activated antibacterial MACPF toxin and provide a framework for understanding how gut Bacteroidales spatially restrict toxin activation to shape strain-level competition.
S. N. Mostyn, K. Flores, G. Hedger et al.· bioRxiv· 0 citations
Several members of the adhesion subfamily of G protein-coupled receptors (aGPCRs) are capable of self-activation by an internal agonist sequence (aka the Stachel) that's exposed upon removal or conformational changes of the N-terminal fragment of the receptor. Synthetic peptides derived from the Stachel sequence can be used as exogenous agonists. In the inactive form of the full-length receptor, the Stachel is sequestered as the β13-strand within the GPCR Autoproteolysis-INducing (GAIN) domain, but it engages the seven transmembrane region as a helix when it is either an intramolecular sequence or a synthetic peptide. Little is known about the molecular details underlying this transition, but we hypothesize that a disordered conformation is central to this intermediate state in receptor activation. Despite the primarily helical Stachel AlphaFold3 and PEP-FOLD4 models predicted with high confidence for the entire aGPCR subfamily, computational predictions and biophysical experiments reveal a predominantly disordered conformation in solution. Investigating the ADGRG6 (also known as GPR126) Stachel peptide, circular dichroism (CD) and nuclear magnetic resonance (NMR) experiments reveal a predominantly random coil conformation in aqueous buffer, polar detergent micelles, and zwitterionic lipids. Titration of trifluoroethanol uncovered a two-state equilibrium between an unfolded and helix-containing conformation with NMR localizing a single-turn helix to residues L846-L849. Taken together, these data indicate the ADGRG6 Stachel peptide is primarily disordered with a subset adopting partial helical structures, likely requiring the steric hindrance of the receptor binding pocket to fully induce helix formation in an induced fit mechanism.
Tucker J. Shriver, Sandra Berndt, Scott A. Robson et al.· Biophysical Journal· 0 citations
It is suggested that bacterial START domains participate in a remarkably broad range of biological processes-including small-molecule binding, metabolic regulation, enzymatic catalysis, and stress adaptation-rather than traditional lipid transport.
Ece Aslan, Ksenia I. Lubova, Alexander Speer et al.· FEMS Microbiology Reviews· 0 citations
Targeted protein degradation mediated by antibodies has emerged as a promising strategy for degrading extracellular or membrane-bound proteins. Proteolysis-Targeting Antibodies (PROTABs) are bispecific antibodies specifically designed to induce the degradation of membrane proteins by tethering them to a cell surface E3 ligase, which promotes ubiquitination and subsequent degradation. Recent studies have demonstrated the potential of PROTABs to degrade oncogenic receptors, but their underlying mechanisms remain to be fully elucidated. Here, we investigated the mechanism of action of a HER2-targeting PROTAB comprising an anti-Zinc and RING finger protein 3 (ZNRF3) arm and an anti-receptor tyrosine-protein kinase erbB-2 (HER2) arm. We show that PROTAB induces rapid ternary complex formation, followed by receptor internalization and degradation, resulting in ~ 85% target depletion within 24 h. Mechanistically, ubiquitination enhances but is not strictly required for internalization, and degradation proceeds predominantly through the lysosomal pathway. Notably, ZNRF3 is not codegraded but instead accumulates at the cell surface, while the PROTAB antibody itself is largely recycled. Importantly, target degradation does not consistently translate into growth inhibition, highlighting the role of cellular context and target dependency. Together, these findings provide a mechanistic framework for PROTAB function and inform the rational design of next-generation antibody-based degraders.
Jieyan He, Tao Sun, Mengwen Zhang et al.· The FEBS Journal· 0 citations
Plasma membrane lipid asymmetry is tightly regulated and fundamental to mammalian cell physiology. TMEM30A is the β-subunit of P4-ATPases, flippase enzymes that maintain strict phosphatidylserine (PS) asymmetry by pumping it from the outer to the cytosolic leaflet. Loss of TMEM30A function causes constitutive PS externalization and has been implicated in diseases such as diffuse large B-cell lymphoma and tumor immune evasion. Here, we systematically define the biophysical and molecular consequences of TMEM30A deletion in immune cells. Using a live-cell lipid reporter, membrane order probe, and surface proteome mapping, we show that TMEM30A-knockout cells display robust PS externalization accompanied by faster lateral diffusion of membrane constituents and decreased plasma membrane order. Surface proteome reorganization includes increased abundance of tetraspanins and CD47. Furthermore, TMEM30A loss triggers glycocalyx remodeling via ADAM10-dependent shedding, which removes major transmembrane mucins, including CD43 and CD162 (also known as SPN and SELPLG, respectively). Together, these data reveal a coordinated reorganization of lipids, glycans and proteins upon TMEM30A loss, suggesting mechanistic links between flippase dysfunction and increased plasma membrane dynamics and potential sensitization to immune therapy. Furthermore, our study provides an integrated surfaceome framework that might shed light on the relationship between TMEM30A expression and clinical outcomes in cancer.
C. Gurdap, F. Ragaller, Marion Muller et al.· Journal of Cell Science· 0 citations
Microbial organisms assemble a diverse array of surface structures to facilitate critical functions including motility, adhesion, and biofilm formation. As extracellular organelles, pili and related surface structures must be able to function in harsh environments and withstand various stressors. Microbes have evolved different strategies to assemble structures able to function under these challenging conditions. This review focuses on bacterial and archaeal systems that utilize donor-strand exchange (DSE) interactions between subunit proteins. DSE is a noncovalent assembly mechanism where one subunit contributes a β-strand to complete the structure of its neighboring subunit. This subunit-subunit interaction is one of the strongest noncovalent interactions known, with the resulting fiber being capable of withstanding extreme environmental stresses and shear forces. We summarize the structural biology and biogenesis of these surface structures, highlighting how DSE-mediated polymerization contributes to the assembly of extracellular structures in both the bacterial and archaeal domains.
Karla Cardenas Arevalo, D. Thanassi· Annual Review of Microbiolog...· 0 citations
