Exploring mitochondrial calcium signalling in human disease: focus on Parkinson’s disease

PINK1 is a mitochondrial Ser/Thr kinase involved in mitophagy. Its mutated forms are associated to genetic forms of Parkinson’s disease. In this work, we have shown the identification and characterization of this protein’s role on mitochondrial calcium (Ca2+) handling. More specifically, we obtained pieces of evidence supporting PINK1 regulation of the mitochondrial calcium uniporter (MCU) complex with an involvement of its kinase activity. Cells overexpressing PINK1 showed a decreased mitochondrial Ca2+ uptake and the co-expression of PINK1 with MICU1, a regulator of the MCU complex promoting mitochondrial Ca2+ influx, caused a reduction of MICU1 protein expression levels. Firstly, we verified the physical interaction between PINK1 and MICU1 by coimmunoprecipitation experiments. Then, to understand how PINK1 could affect MICU1 activity, we generated non phosphorylable and phosphomimetic mutants of MICU1 at the T256 residue, a putative phosporylation site for PINK1, identified by alignment between the ubiquitin sequence, that isphosphorylated by PINK1, and the MICU1 sequence. Cells overexpressing MICU1 phosphomimetic mutant T256E showed a decreased mitochondrial Ca2+ transient respect to cells overexpressing MICU1 WT. Also, we observed a more rapid turnover of MICU1 T256E mutant, that can explain the lower mitochondrial Ca2+ transient of cells overexpressing this mutant. MICU1 T256 phosphomimetic mutation mimic the effects of PINK1 on MICU1 WT. Using the artificial mutant of PINK1 lacking of its kinase activity, PINK1 KDD, and the Parkinson’s disease related mutant of PINK1 (PINK1 W437*), that has a reduced kinase activity, we found that these two mutant had only a partial effect on mitochondrial calcium uptake respect to PINK1 wt, supporting the high relevance of PINK1 kinase activity in the regulation of mitochondrial calcium influx.