Natural and induced mitochondrial phosphate carrier loss: differential dependence of mitochondrial metabolism and dynamics, and cell survival, on the extent of depletion

Seifert, Erin L. and Gál, Anikó and Acoba, Michelle G. and Li, Qipei and Anderson-Pullinger, Lauren and Golenár, Tünde and Hajnóczky, György (2016) Natural and induced mitochondrial phosphate carrier loss: differential dependence of mitochondrial metabolism and dynamics, and cell survival, on the extent of depletion. JOURNAL OF BIOLOGICAL CHEMISTRY, 2016. ISSN 0021-9258

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Abstract

The relevance of PiC, encoded by SLC25A3, in bioenergetics is well accepted. However, little is known about the mechanisms mediating the cellular impairments induced by pathological SLC25A3 variants. To this end, we investigated the pathogenicity of a novel compound heterozygous mutation in SLC25A3. First, each variant was modeled in yeast, revealing that substituting GSSAS for QIP within the 5th matrix loop is incompatible with survival on non-fermentable substrate whereas the L200W variant is functionally neutral. Next, using skin fibroblasts from an individual expressing these variants, and HeLa cells with varying degrees of PiC depletion, PiC loss of ~60% was still compatible with uncompromised maximal oxidative phosphorylation (oxphos) whereas lower maximal oxphos was evident at ~85% PiC depletion. Furthermore, intact mutant fibroblasts displayed suppressed mitochondrial bioenergetics consistent with a lower substrate availability rather than phosphate limitation. This was accompanied by slowed proliferation in glucose-replete media, however proliferation ceased when only mitochondrial substrate was provided. Both mutant fibroblasts and HeLa cells with 60% PiC loss showed a less interconnected mitochondrial network and a mitochondrial fusion defect that is not explained by altered abundance of OPA1, MFN1/2 or relative amount of different OPA1 forms. Altogether these results indicate that PiC depletion may need to be profound (>85%) to substantially affect maximal oxphos and that pathogenesis associated with PiC depletion or loss-of-function may be independent of phosphate limitation when ATP requirements are not high.

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