A widely employed germ cell marker is an ancient disordered protein with reproductive functions in diverse eukaryotes

Abstract

The advent of sexual reproduction and the evolution of a dedicated germline in multicellular organisms are critical landmarks in eukaryotic evolution. We report an ancient family of GCNA (germ cell nuclear antigen) proteins that arose in the earliest eukaryotes, and feature a rapidly evolving intrinsically disordered region (IDR). Phylogenetic analysis reveals that GCNA proteins emerged before the major eukaryotic lineages diverged; GCNA predates the origin of a dedicated germline by a billion years. Gcna gene expression is enriched in reproductive cells across eukarya -either just prior to or during meiosis in single-celled eukaryotes, and in stem cells and germ cells of diverse multicellular animals. Studies of Gcna-mutant C. elegans and mice indicate that GCNA has functioned in reproduction for at least 600 million years. Homology to IDRcontaining proteins implicated in DNA damage repair suggests that GCNA proteins may protect the genomic integrity of cells carrying a heritable genome.

SUMO ylation promotes protective responses to DNA ‐protein crosslinks

“…The findings above suggested that ACRC might have a role in processing DPCs in a SUMO‐driven manner. However, while homologs of ACRC are encoded by many eukaryotic genomes (Carmell et al , ), we found that it was absent or expressed at levels below the limit of detection in a range of human primary and cancer cell lines (Bekker‐Jensen et al , ; our unpublished observations), in agreement with recent work suggesting that expression of ACRC orthologs is enhanced in germ and stem cells (Carmell et al , ). Notably, ectopic expression of ACRC in U2OS cells reduced cellular fitness in a SIM‐dependent manner and delayed the clearance of DNMT1 DPCs (Fig EV3E and F), suggesting that a potential involvement of ACRC in SUMO‐mediated responses to DPCs could be restricted to highly specific cellular settings while enforcing its expression outside this context may interfere with other DPC‐processing mechanisms.…”

“…The work described here suggests an involvement of SUMO-binding ACRC/ GCNA family SprT proteases in such processes, and we provide evidence that in C. elegans germ and early embryonic cells, GCNA-1 has an important role in protecting against the lethality of DPC-inducing agents including formaldehyde in a manner that involves the integrity of its protease domain and is epistatic with SUMO. However, it has been shown that GCNA proteases are predominantly expressed in germ and stem cells (Carmell et al, 2016), supported by our findings, suggesting that unlike SPRTN, the involvement of this family of SprT proteases in cellular DPC responses is, in all likelihood, highly tissue-restricted. While we have not yet been able to experimentally verify that GCNA-type proteins are active proteases, this seems warranted by our finding that point mutation of the highly conserved putative catalytic glutamic acid residue in the SprT domain of C. elegans GCNA-1 sensitizes worms to formaldehyde to the same extent as gcna-1 lossof-function.…”

“…The findings above suggested that ACRC might have a role in processing DPCs in a SUMO-driven manner. However, while homologs of ACRC are encoded by many eukaryotic genomes (Carmell et al, 2016), we found that it was absent or expressed at levels below the limit of detection in a range of human primary and cancer cell lines (Bekker-Jensen et al, 2017; our unpublished observations), in agreement with recent work suggesting that expression of ACRC orthologs is enhanced in germ and stem cells (Carmell et al, 2016).…”

“…We then generated a gcna-1 loss of function (lof) allele by knocking in a~6.6 kb gfp-containing selection cassette between the gcna-1 promoter and coding sequence, simultaneously generating a transcriptional gfp reporter (Figs 5C and EV4A; Dickinson et al, 2015). Inspection of gcna-1 promoter-driven GFP expression confirmed that GCNA-1 is mainly expressed in germ cells and early embryonic, proliferating cells but not in post-mitotic tissues (Figs 5D and EV4B;Carmell et al, 2016). Interestingly, using assays probing for survival of germ and early embryonic cells, we found that gcna-1 loss of function led to elevated formaldehyde sensitivity ( Fig 5E).…”

The testicular soma of Tsc22d3 knockout mice supports spermatogenesis and germline transmission from spermatogonial stem cell lines upon transplantation

“…The mouse vasa homologue (MVH) is expressed in the cytoplasm of germ cells; Vasa (also known as Ddx4) is an ATP-dependent RNA helicase that is highly conserved among vertebrates and invertebrates (Castrillon, Quade, Wang, Quigley, & Crum, 2000;Gustafson & Wessel, 2010). The antigen detected by the TRA98 antibody is the Gcna1 gene product (Carmell et al, 2016), and resides in the nucleus of testis germ cells (Inoue, Onohara, & Yokota, 2011). CD146, also called melanoma cell adhesion molecule (MCAM), is a transmembrane glycoprotein used as an SSC cell surface marker (Kanatsu- Shinohara, Morimoto, & Shinohara, 2012).…”