Identification of a gain-of-function mutation in a Golgi P-type ATPase that enhances Mn 2+ efflux and protects against toxicity

• DOI: 10.1073/pnas.1013642108
• Publication Date: 2010-12-28T01:45:14Z
• Authors: Somshuvra Mukhopa...; Adam D. Linstedt
• Keywords: Adenosine Triphosphatases; Models, Molecular; Manganese; 0303 health sciences; Alanine; Molecular Conformation; Vesicular Transport Proteins; Golgi Apparatus; Calcium-Transporting ATPases; Proton-Translocating ATPases; 03 medical and health sciences; Cytosol; FOS: Biological sciences; Mutation; Humans; Point Mutation; Phosphorylation; 69999 Biological Sciences not elsewhere classified; HeLa Cells

P-type ATPases transport a wide array of ions, regulate diverse cellular processes, and are implicated in a number of human diseases. However, mechanisms that increase ion transport by these ubiquitous proteins are not known. SPCA1 is a P-type pump that transports Mn 2+ from the cytosol into the Golgi. We developed an intra-Golgi Mn 2+ sensor and used it to screen for mutations introduced in SPCA1, on the basis of its predicted structure, which could increase its Mn 2+ pumping activity. Remarkably, a point mutation (Q747A) predicted to increase the size of its ion permeation cavity enhanced the sensor response and a compensatory mutation restoring the cavity to its original size abolished this effect. In vivo and in vitro Mn 2+ transport assays confirmed the hyperactivity of SPCA1-Q747A. Furthermore, increasing Golgi Mn 2+ transport by expression of SPCA1-Q747A increased cell viability upon Mn 2+ exposure, supporting the therapeutic potential of increased Mn 2+ uptake by the Golgi in the management of Mn 2+ -induced neurotoxicity.