Cryo-EM structure of the mechanically activated ion channel OSCA1.2

Abstract

Main 41We purified Arabidopsis thaliana OSCA1.2 expressed in HEK293F cells and determined 42 cryo-EM reconstructions in lipidic nanodiscs and LMNG detergent micelle with cholesteryl 43 hemisuccinate (CHS) ( Fig. 1a-d, Extended Data Figs. 1,2). The resulting density maps 44 allowed building of 82% of the full-length OSCA1.2 sequence (Extended Data Fig. 3, 45 Extended Data Table 1). Structures of OSCA1.2 in nanodisc and LMNG were nearly 46 identical (RMSD=0.81 Å, Extended Data Fig. 4a-c). We refer to the nanodisc structure 47

Heterozygous variants in the mechanosensitive ion channelTMEM63Aresult in transient hypomyelination during infancy

“…The approximate location of the extracellular (Ex) and intracellular (In) surfaces of plasma membrane indicated by dashed black lines. The five pore lining transmembrane helices are shown in green [20]. C) Section of TMEM63A structural model looking through pore region towards the extracellular domain.…”

“…Gly576 residue shown as red spheres, pink mesh indicated the predicted position the serine side-chain would occupy in the p.Gly576Ser mutant protein. Conserved residue Glu571 (shown as brown sticks) that has been shown to reduce stretch activated ion conductance when mutated to alanine in OSCA1.2 can be seen adjacent to Gly576 [20]. D) Clustal alignment of TMEM63A protein from 10 vertebrate species and the Arabidopsis homologue OSCA1.2.…”

“…Using a cryo-EM structure of the TMEM63A homologue OSCA1.2 from Arabidopsis, 166 we generated a model of TMEM63A protein structure using SWISS-MODEL (Figure 167 3b) [20,21]. The Gly168 residue sits in an intracellular loop between helix 1 and helix 2 168 while both Ile462 and Gly567 are located within pore lining transmembrane helices of 169 TMEM63A.…”

“…The Gly168 residue sits in an intracellular loop between helix 1 and helix 2 168 while both Ile462 and Gly567 are located within pore lining transmembrane helices of 169 TMEM63A. Interestingly Gly567 resides on the internal face of the pore one helix turn 170 from the conserved residue Glu571 that has been shown to reduce stretch activated ion 171 conductance when mutated to alanine in OSCA1.2 [20]. 172 We introduced the variants seen in these individuals by site directed mutagenesis 173 into TMEM63A clones purchased from Origene (NM 014698, Cat No.…”

Overexpression of osmosensitive Ca2+-activated channel TMEM63B promotes migration in HEK293T cells

“…[9] Investigation of the conserved domain architecture among these transporters revealed the existence of orthologs present in various taxonomic groups, such as fungi, green algae, plants, birds and mammals, which suggests the functional conservation of this family throughout higher eukaryotes. With the availability of sequences of various ANO superfamily members and the recently reported 3D structures of some OSCA proteins in Arabidopsis thaliana [4][5][6] it is possible to get an insight into the structural aspects of TMEM63B domains involved in the ion translocation across the membrane.…”