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Crnkovic, A.

Publications and source records attributed to Crnkovic, A..

2 recordsLinked to original sources

High-resolution cryo-EM structures of a protein pore reveal diverse roles of membrane lipids

The structure and function of membrane proteins depend on their interactions with the lipid molecules that constitute lipid membranes. Actinoporins are a family of -pore-forming proteins that bind specifically to sphingomyelin-containing lipid membranes, where they oligomerize and form transmembrane pores. The numerous contacts they form with the lipid membrane make them an exemplary object for studying the different roles that lipids play in the structure and function of membrane proteins. Through a comprehensive cryo-electron microscopic analysis of a pore formed by an actinoporin Fav from the coral Orbicella faveolata, we show that the octameric pore interacts with 112 lipids in the upper leaflet of the membrane. The structures of Fav pores formed on different lipid membranes reveal the different roles of lipids and demonstrate that the actinoporin surface is perfectly suited for binding multiple receptor sphingomyelin molecules. When cholesterol is present in the membrane, it forms nanodomains associated with the pore, leading to a tighter arrangement of lipids, which in turn increases the stability of the pores. Atomistic simulations support the structural data, show that the protein-bound lipids are not mobile, and reveal additional effects of the pore on the lipid membrane. Overall, these data reveal a complex network of protein-lipid and lipid-lipid interactions, and an underrated role of lipids in the structure and function of transmembrane protein complexes.

biochemistry↗

An Aedes aegypti seryl-tRNA synthetase paralog controls bacteroidetes growth in the midgut

Insect gut microbiota plays important roles in host physiology, such as nutrition, digestion, development, fertility, and immunity. We have found that in the intestine of Aedes aegypti, SLIMP (seryl-tRNA synthetase like insect mitochondrial protein) knockdown followed by a blood meal promotes dysbiosis, characterized by the overgrowth of a specific bacterial phylum, Bacteroidetes. In turn, the latter decreased both infection rates and Zika virus prevalence in the mosquitoes. Previous work in Drosophila melanogaster showed that SLIMP is involved in protein synthesis and mitochondrial respiration in a network directly coupled to mtDNA levels. There are no other reports on this enzyme and its function in other insect species. Our work expands the knowledge of the role of these SerRS paralogs. We show that A. aegypti SLIMP (AaeSLIMP) clusters with SLIMPs of the Nematocera sub-order, which have lost both the tRNA binding domain and active site residues, rendering them unable to activate amino acids and aminoacylate tRNAs. Knockdown of AaeSLIMP did not significantly influence the mosquitoes survival, oviposition, or eclosion. It also neither affected midgut cell respiration nor mitochondrial ROS production. However, it caused dysbiosis, which led to the activation of Dual oxidase and resulted in increased midgut ROS levels. Our data indicate that the intestinal microbiota can be controlled in a blood-feeding vector by a novel, unprecedent mechanism, impacting also mosquito vectorial competence towards zika virus and possibly other pathogens as well. Author SummaryAminoacyl-tRNA synthetases (aaRS) are a family of ubiquitous enzymes responsible for the attachment of specific amino acids to their cognate tRNAs. During evolution some aaRS acquired new domains and/or suffered gene duplications, resulting in the improvement and expansion of their functions some of them being specific to a group of organisms. A paralog of seryl-tRNA synthetase restricted to the class Insecta (SLIMP) is found in Arthropoda. Our goal was to explore the role of SLIMP in the female mosquito Aedes aegypti using RNA interference. We showed that A. aegypti SLIMP (AaeSLIMP) gene expression is up-regulated upon blood feeding through a heme-dependent signaling. Although AaeSLIMP knockdown neither impacted the mosquito survival nor oviposition, it provoked ROS levels augmentation in the midgut via Dual Oxidase activity in order to control the increase in the intestinal native microbiota, specifically bacteria of the Bacteroidetes phylum. Although dysbiosis can result from mitochondrial impairment, this is the first time that the absence of a mitochondrial enzyme is linked to intestinal microbiota without any visible effects in mitochondrial respiration and mitochondrial ROS production. Furthermore, Zika Virus infection of AaeSLIMP silenced mosquitoes is decreased when comparing to control, meaning that Bacteroidetes overgrowth may be protecting the female mosquito. Our data indicate that the intestinal microbiota can be controlled in a blood-feeding vector by a novel, unprecedent mechanism, impacting also mosquito vectorial competence towards zika virus and possibly other pathogens as well.

biochemistry↗