Novel Mutations in SLC25A3 Encoding the Mitochondrial Phosphate Carrier

Abstract

It has long been known that the mitochondrial phosphate carrier (PiC), encoded by the nuclear gene SLC25A3, serves a dual role as the primary means of phosphate (Pi) uptake into mitochondria for oxidative phosphorylation and to serve as the main buffering species for calcium entering mitochondria. Yet only lately have mutations in human PiC been identified. We recently described an infant who is a compound heterozygote for two novel PiC variants, c.599T>G (p.Leu200Trp) and c. 886_898delGGTAGCAGTGCTTins CAGATAC (p.Gly296_ Ser300delins GlnIlePro) (Bhoj et al., 2015; PMID 25681081). This compound mutation affects both the muscle-specific isoform, PiC-A, and the ubiquitously expressed isoform, PiC-B. Of note, these two mutations are the first to affect the PiC-B isoform. Phenotypically, the infant initially had severe cardiomyopathy requiring heart transplantation, but had minimal skeletal myopathy and is now relatively healthy. Here, we used patient fibroblasts and yeast expressing humanized versions of Pic1p (the yeast PiC homolog) to analyze these novel PiC variants. Analyses in yeast suggested that the Leu200Trp mutation, affecting a highly conserved leucine, yields functional PiC protein, whereas the del/ins mutation does not. In patient fibroblasts, PiC protein expression was barely detectable, despite the potentially neutral Leu200Trp variant, and phosphorylating oxygen consumption was decreased by ∼50%. The lower oxidative phosphorylation led us to question if mitochondrial fusion activity was altered, since inner mitochondrial membrane fusion depends on mitochondrial ATP. Studies using fibroblasts transfected with mitochondrial matrix-targeted DsRed and photoactivatable GFP indicated suppressed fusion activity of the inner mitochondrial membrane. Overall, these observations provide insight into the pathologic potential of these newly described PiC variants and, more generally, indicate that heterozygosity of a deleterious variant may be sufficient to cause mitochondrial dysfunction and organismal pathology.