TMEM16E (GDD1) Exhibits Protein Instability and Distinct Characteristics in Chloride Channel/Pore Forming Ability

Abstract

TMEM16E/GDD1 has been shown to be responsible for the bone-related late-onset disease gnathodiaphyseal dysplasia (GDD), with the dominant allele (TMEM16E(gdd) ) encoding a missense mutation at Cys356. Additionally, several recessive loss-of-function alleles of TMEM16E also cause late-onset limb girdle muscular dystrophy. In this study, we found that TMEM16E was rapidly degraded via the proteasome pathway, which was rescued by inhibition of the PI3K pathway and by the chemical chaperone, sodium butyrate. Moreover, TMEM16E(gdd) exhibited lower stability than TMEM16E, but showed similar propensity to be rescued. TMEM16E did not exhibit cell surface calcium-dependent chloride channel (CaCC) activity, which was originally identified in TMEM16A and TMEM16B, due to their intracellular vesicle distribution. A putative pore-forming domain of TMEM16E, which shared 39.8% similarity in 98 amino acids with TMEM16A, disrupted CaCC activity of TMEM16A via domain swapping. However, the Thr611Cys mutation in the swapped domain, which mimicked conserved cysteine residues between TMEM16A and TMEM16B, reconstituted CaCC activity. In addition, the GDD-causing cysteine mutation made in TMEM16A drastically altered CaCC activity. Based on these findings, TMEM16E possesses distinct function other than CaCC and another protein-stabilizing machinery toward the TMEM16E and TMEM16E(gdd) proteins should be considered for the on-set regulation of their phenotypes in tissues.

A novel ANO5 splicing variant in a LGMD2L patient leads to production of a truncated aggregation‐prone Ano5 peptide

“…Consistent with previous studies , our data also show that recombinant Ano5 resides primarily on the ER membrane and intracellular vesicles (Figure ). To study the expression and subcellular localization of endogenous Ano5 in human muscles, we performed immunofluorescence staining with the N421A/85 antibody and the antibody against dystrophin or calnexin.…”

“…Previous studies showed that Ano5 was rapidly degraded via the proteasome pathway and transient transfection with a plasmid encoding Ano5‐GFP resulted in very low GFP fluorescence , despite the high transfection efficiency. To validate the antibodies raised against human/mouse Ano5 antigens, we constructed a recombinant adenovirus (Ad) expressing human Ano5 with FLAG and YFP tags at the N‐ and C‐terminus, respectively.…”

A Role of TMEM16E Carrying a Scrambling Domain in Sperm Motility

“…To confirm this possibility, it would be necessary to determine whether the TMEM16A-E SCRD chimera can support microparticle production. Point mutations found in GDD patients appear to make the TMEM16E dominant-active (11,59). It will be interesting to examine the structure of intracellular membranes in cells expressing these mutants.…”

TMEM16, LRRC8A, bestrophin: chloride channels controlled by Ca2+ and cell volume

“…TMEM16E, which was not found to produce CaCC [32]. Instability of TMEM16E may be the cause for gnathodiaphyseal dysplasia [32,33].…”

“…Instability of TMEM16E may be the cause for gnathodiaphyseal dysplasia [32,33]. Although ANO5 is expressed primarily in muscle, bone, and testis, other studies also suggest expression in the gastrointestinal tract and in lung [19,34].…”