Biology subjects

Hiotis, G.

Publications and source records attributed to Hiotis, G..

3 recordsLinked to original sources

bioRxiv2022

Dynamic HIV-1 spike motion creates vulnerability for its membrane-bound tripod to antibody attack

Vaccines targeting HIV-1s gp160 spike protein are stymied by high viral mutation rates and structural chicanery. gp160s membrane-proximal external region (MPER) is the target of naturally arising broadly neutralizing antibodies (bnAbs), yet MPER-based vaccines fail to generate bnAbs. Here, nanodisc-embedded spike protein was investigated by cryo-electron microscopy and molecular-dynamics simulations, revealing spontaneous ectodomain tilting that creates vulnerability for HIV-1. While each MPER protomer radiates centrally towards the three-fold axis contributing to a membrane-associated tripod structure that is occluded in the upright spike, tilting provides access to the opposing MPER. Structures of spike proteins with bound 4E10 bnAb Fabs reveal that the antibody binds exposed MPER, thereby altering MPER dynamics, modifying average ectodomain tilt, and imposing strain on the viral membrane and the spikes transmembrane segments, resulting in the abrogation of membrane fusion and informing future vaccine development.

biophysics↗

bioRxiv2022

A ubiquitin-independent proteasome pathway controls the CARD8 inflammasome

CARD8 is a pattern-recognition receptor that forms a caspase-1-activating inflammasome. CARD8 undergoes autoproteolysis, generating an N-terminal (NT) fragment with a disordered region and a ZU5 domain and a C-terminal (CT) fragment with UPA and CARD domains. DPP8 and DPP9 (DPP8/9) inhibitors, including Val-boroPro (VbP), accelerate the degradation of the NT fragment via a poorly characterized proteasome-mediated pathway, thereby releasing the inflammatory CT fragment from autoinhibition. Here, we show that the core 20S proteasome, which degrades disordered and misfolded proteins independent of ubiquitin, controls CARD8 activation. In unstressed cells, the 20S proteasome degrades just the NT disordered region, leaving behind the folded ZU5, UPA, and CARD domains to act as an inhibitor of inflammasome assembly. In VbP-stressed cells, the 20S proteasome degrades the entire NT fragment, perhaps due to ZU5 domain unfolding, freeing the CT fragment from autoinhibition. Overall, this work shows that CARD8 NTs susceptibility to 20S proteasome-mediated degradation controls inflammasome activation.

immunology↗

bioRxiv2021

Impaired Cx43 gap junction endocytosis causes cardiovascular defects in zebrafish

Gap junction proteins, termed connexins (Cx), mediate direct cell-to-cell communication by forming channels that physically couple cells, thereby linking their cytoplasm, permitting exchange of molecules, ions, and electrical impulses. The most ubiquitously expressed gap junction protein, connexin43 (Cx43) has been implicated in cardiovascular diseases including arrhythmias, cardiomyopathies, hypertension and diabetes. The Cx43 C-terminal (CT) domain serves as the regulatory hub of the protein affecting all aspects of gap junction function. Here, deletion within the Cx43 CT (amino acids 256-289), a region known to encode key residues regulating gap junction turnover is employed to examine the effects of dysregulated Cx43 gap junction endocytosis using cultured cells (Cx43{Delta}256-289) and zebrafish model (cx43lh10). We report that this CT deletion causes defective gap junction endocytosis as well as increased gap junction intercellular communication (GJIC). Increased Cx43 protein content in cx43lh10 zebrafish, specifically in the cardiac tissue, larger gap junction plaques and longer Cx43 protein half-lives coincide with severely impaired cardiovascular development. These findings suggest that normal, unimpaired Cx43 gap junction endocytosis and turnover is an essential aspect of gap junction function as demonstrated here for cardiovascular development that when impaired can give rise to arrhythmias, heart malformations and aberrant vasculature structure and function.

cell biology↗