Vesicular distribution of Secretory Pathway Ca²⁺-ATPase isoform 1 and a role in manganese detoxification in liver-derived polarized cells

Manganese is a trace element that is an essential co-factor in many enzymes critical to diverse biological pathways. However, excess Mn²⁺ leads to neurotoxicity, with psychiatric and motor dysfunction resembling parkinsonism. The liver is the main organ for Mn²⁺ detoxification by excretion into bile. Although many pathways of cellular Mn²⁺ uptake have been established, efflux mechanisms remain essentially undefined. In this study, we evaluated a potential role in Mn²⁺ detoxification by the Secretory Pathway Ca²⁺, Mn²⁺-ATPase in rat liver and a liver-derived cell model WIF-B that polarizes to distinct bile canalicular and sinusoidal domains in culture. Of two known isoforms, only secretory pathway Ca²⁺-ATPase isoform 1 (SPCA1) was expressed in liver and WIF-B cells. As previously observed in non-polarized cells, SPCA1 showed overlapping distribution with TGN38, consistent with Golgi/TGN localization. However, a prominent novel localization of SPCA1 to an endosomal population close to, but not on the basolateral membrane was also observed. This was confirmed by fractionation of rat liver homogenates which revealed dual distribution of SPCA1 to the Golgi/TGN and a fraction that included the early endosomal marker, EEA1. We suggest that this novel pool of endosomes may serve to sequester Mn²⁺ as it enters from the sinusoidal/basolateral domains. Isoform-specific partial knockdown of SPCA1 delayed cell growth and formation of canalicular domain by about 30% and diminished viability upon exposure to Mn²⁺. Conversely, overexpression of SPCA1 in HEK 293T cells conferred tolerance to Mn²⁺ toxicity. Taken together, our findings suggest a role for SPCA1 in Mn²⁺ detoxification in liver.