NME6 belongs to the nucleoside diphosphate kinase (NDPK/NME/Nm23) family of enzymes that catalyze the transfer of the terminal phosphate from nucleoside triphosphates, to nucleoside diphosphates. During recent years, research revealed a growing number of NDPK/NME molecular functions related to human pathologies. The family consists of ten members divided in two groups. Group I members, NME1-NME4, are highly homologous among themselves and with orthologues in other Metazoans. They possess the NDPK activity in the hexameric form. The evolutionary older NME5-NME9 (Group II), display lower mutual homology and remain scarcely studied. Our group provided important information on NME6, an unusual NDP kinase and resolved existing discrepancies. We positioned the NME6 in the mitochondrial matrix, despite the absence of a canonical mitochondrial targeting sequence. Contrary to Group I members, NME6 prefers the monomeric state therefore lacking the NDP kinase activity. Physical interaction was confirmed with RCC1L (WBSCR16), a protein involved in mitoribosome assembly and translation. The overexpression of NME6 reduced the mitochondrial respiration capacity. Thus, our research revealed unexpected properties of NME6 and the NME Group II proteins, and opened new, important research avenues. In this project we will use standard and state-of-the art methods to 1) describe the non-canonical import of NME6 to mitochondria, 2) define its functioning hub within the mitochondrial protein expression, 3) provide biophysical answers to its oligomeric state and enzymatic activity and 4) explore interaction interfaces with its protein partners. We are confident that our research will not only answer questions about the biology of the NDPK/NME family but contribute to the understanding of the non-canonical mitochondrial import in general, the control of oxidative phosphorylation and the mechanisms of maturation of mitochondrial RNA and mitoribosome assembly, which remains poorly understood.

Within this project we aim to:

A)      describe the non-canonical import mechanism of NME6 to mitochondria

B)      narrow down the possible NME6 functions in mitochondrial respiration, mRNA maturation and mitoribosomal assembly

C)      clarify the structure-function relationships of NME6