Expression of mammalian Onzin and Fungal Cadmium Resistance 1 in S. cerevisiae suggests ancestral functions of PLAC8 proteins in regulating mitochondrial metabolism and DNA damage repair

Protein domains are structurally and functionally distinct units responsible for particular protein functions or interactions. Although protein domains contribute to the overall protein function(s) and can be used for protein classification, about 20% of protein domains are currently annotated as “domains of an unknown function” (DUFs). DUF 614, a cysteine-rich domain better known as PLAC8 (Placenta-Specific Gene 8), occurs in proteins found in the majority of Eukaryotes. PLAC8-containing proteins play important yet diverse roles in different organisms, such as control of cell proliferation in animals and plants or heavy metal resistance in plants and fungi. For example, Onzin from Mus musculus is a key regulator of cell proliferation, whereas FCR1 from the ascomycete Oidiodendron maius confers cadmium resistance. Onzin and FCR1 are small, single-domain PLAC8 proteins and we hypothesized that, despite their apparently different role, a common molecular function of these proteins may be linked to the PLAC8 domain. To address this hypothesis, we compared these two PLAC8-containing proteins by heterologous expression in the PLAC8-free yeast Saccharomyces cerevisiae . When expressed in yeast, both Onzin and FCR1 improved cadmium resistance, reduced cadmium-induced DNA mutagenesis, localized in the nucleus and induced similar transcriptional changes. Our results support the hypothesis of a common ancestral function of the PLAC8 domain that may link some mitochondrial biosynthetic pathways (i.e. leucine biosynthesis and Fe-S cluster biogenesis) with the control of DNA damage, thus opening new perspectives to understand the role of this protein domain in the cellular biology of Eukaryotes. Author Summary Protein domains are the functional units of proteins and typically have distinct structure and function. However, many widely distributed protein domains are currently annotated as “domains of unknown function” (DUFs). We have focused on DUF 614, a protein domain found in many Eukaryotes and better known as PLAC8 (Placenta-Specific Gene 8). The functional role of DUF 614 is unclear because PLAC8 proteins seem to play important yet different roles in taxonomically distant organisms such as animals, plants and fungi. We used S. cerevisiae to test whether these apparently different functions, namely in cell proliferation and metal tolerance, respectively reported for the murine Onzin and the fungal FCR1, are mediated by the same molecular mechanisms. Our data demonstrate that the two PLAC8 proteins induced the same growth phenotype and transcriptional changes in S. cerevisiae . In particular, they both induced the biosynthesis of the amino acid leucine and of the iron-sulfur cluster, one of the most ancient protein cofactors. These similarities support the hypothesis of an ancestral function of the DUF 164 domain, whereas the transcriptomic data open new perspectives to understand the role of PLAC8-proteins in Eukaryotes.