Biology subjects

Poulsen, M. H.

Publications and source records attributed to Poulsen, M. H..

4 recordsLinked to original sources

bioRxiv2025

Coupling between acid-sensing ion channel 1a and the monocarboxylate transporter family shapes cellular pH response

Proton-mediated activation of acid-sensing ion channel 1a (ASIC1a) leads to influx of sodium and, to a lesser extent, calcium, followed by rapid and complete desensitization. However, various ASIC1a-expressing cell types display atypical proton-induced currents characterized by altered proton sensitivity and slow and incomplete desensitization. The origin of this functional diversity and its physiological relevance remain unclear. Here, we show that the monocarboxylate transporter 1 (MCT1) interacts with ASIC1a, leading to subtype-specific modulation of proton sensitivity and desensitization. We delineate the molecular mechanism of the ASIC1a-MCT1 coupling and find that the presence of MCT1 is required for the distinct ASIC1a-mediated currents observed in glioblastoma cells, the most lethal brain cancer in adults. Strikingly, presence of ASIC1a together with MCT1 significantly prolongs the lifespan of mice intracranially injected with glioblastoma cells. We thus uncover the basis for altered ASIC1a function in glioblastoma and highlight the importance of deciphering the cell-type specific ASIC1a interactome.

biophysics↗

bioRxiv2024

Conformational plasticity of human acid-sensing ion channel 1a

Acid-sensing ion channels (ASICs) are typically activated by acidic environments and contribute to nociception and synaptic plasticity. ASIC1a is the most abundant subunit in the central nervous system and forms homomeric channels permeable to Na+ and Ca2+, making it a compelling therapeutic target for acidotic pathologies including stroke and traumatic brain injury. However, a complete conformational library of human ASIC1a in its various functional states has yet to be described. Using cryo-EM, we obtained hASIC1a structures across a pH range between 8.5 and 5.7, as well as in the presence of a toxin agonist and a gating-modulating mutation. We identify six major conformations that establish linear transmembrane helices to be associated with an open state, delineate mechanistic differences between proton and toxin activation, and demonstrate that desensitization leads to unexpected conformational diversity in the transmembrane domain. Together, they provide a three-dimensional rationalization of decades of structure-function studies on ASIC.

biochemistry↗

bioRxiv2021

Disrupting inter-subunit interactions favors channel opening in P2X2 receptors

Background and PurposeP2X receptors (P2XRs) are trimeric ligand-gated ion channels (LGICs) that open a cation-selective pore in response to ATP binding to their large extracellular domain (ECD). The seven known P2XR subtypes can assemble as homo- or heterotrimeric complexes and contribute to numerous physiological functions, including nociception, inflammation and hearing. The overall structure of P2XRs is well established, but little is known about the spectrum and prevalence of human genetic variations and the functional implications in specific domains. Experimental ApproachHere we examine the impact of P2X2 receptor (P2X2R) inter-subunit interface missense variants identified in the human population or through structural predictions. We test both single and double mutants through electrophysiological and biochemical approaches. Key resultsWe demonstrate that predicted ECD inter-subunit interfaces display a higher-than-expected density of missense variations and that the majority of mutations that disrupt putative inter-subunit interactions result in channels with higher apparent ATP affinity. Lastly, we show that double mutants at the subunit interface show significant energetic coupling, especially if located in close proximity. Conclusions and ImplicationsWe provide the first structural mapping of the mutational burden across the human population in a LGIC and show that the density of missense mutations is constrained between protein domains, indicating evolutionary selection at the domain level. Our data may indicate that, unlike other LGICs, P2X2Rs have evolved an intrinsically high threshold for activation, possibly to allow for additional modulation or as a cellular protection mechanism against overstimulation. Bullet point summary What is already known: O_LIP2X2 receptors are ATP-activated ion channels implicated in hearing and nociceptice pathways C_LI What this study adds: O_LIA structural mapping of missense variants observed in the human population C_LIO_LIWe identify the intersubunit-interface as a variant hotspot and decipher functional impact of mutations C_LI Clinical significance: O_LIThe development of both inhibitors and activators of P2X2 receptor function may be required C_LI

biophysics↗

bioRxiv2020

Chemical modification of proteins by insertion of synthetic peptides using tandem protein trans-splicing

Manipulation of proteins by chemical modification is a powerful way to decipher their function or harness that function for therapeutic purposes. Despite recent progress in ribosome-dependent and semi-synthetic chemical modifications, these techniques sometimes have limitations in the number and type of modifications that can be simultaneously introduced or their application in live eukaryotic cells. Here we present a new approach to incorporate single or multiple post-translational modifications or non-canonical amino acids into soluble and membrane proteins expressed in eukaryotic cells. We insert synthetic peptides into proteins of interest via tandem protein trans-splicing using two orthogonal split intein pairs and validate our approach by investigating different aspects of GFP, NaV1.5 and P2X2 receptor function. Because the approach can introduce virtually any chemical modification into both intracellular and extracellular regions of target proteins, we anticipate that it will overcome some of the drawbacks of other semi-synthetic or ribosome-dependent methods to engineer proteins.

synthetic biology↗