Cofilin recruits F-actin to SPCA1 and promotes Ca2+-mediated secretory cargo sorting.
Bottom Line: How these proteins interact and activate the pump to facilitate cargo sorting, however, is not known.A 132-amino acid portion of the SPCA1 phosphorylation domain (P-domain) interacted with actin in a CFL-1-dependent manner.Altogether, our findings reveal the mechanism of CFL-1-dependent recruitment of actin to SPCA1 and the significance of this interaction for Ca(2+) influx and secretory cargo sorting.
Affiliation: Max Planck Institute of Biochemistry, 82152 Martinsried, Germany.Show MeSH
License 1 - License 2
Mentions: To confirm the ss-HRP secretion phenotype with additional sorting markers, control HeLa cells and HeLa cells expressing siRNA-resistant SPCA1-wt and -mut4 were transfected with control or SPCA1 siRNA, respectively. Cell pellets and supernatants were analyzed for the presence of Cab45 and Cathepsin D by Western blotting. The data clearly showed that SPCA1-wt rescued Cab45 and Cathepsin D sorting, whereas the -mut4 did not (Fig. 10, A–D). Finally, we measured the TGN Ca2+ uptake in HeLa cells stably expressing SPCA1-wt and -mut4 treated with SPCA1 siRNA and transfected with Go-D1-cpv. SPCA1-wt rescued the siRNA-induced defect of Ca2+ uptake, whereas SPCA1-mut4 did not (Fig. 10 E).
Affiliation: Max Planck Institute of Biochemistry, 82152 Martinsried, Germany.