SUMO1 in human sperm: new targets, role in motility and morphology and relationship with DNA damage

Abstract

In studies carried out previously, we demonstrated that small ubiquitin-like modifier 1 (SUMO1) is associated with poor sperm motility when evaluated with a protocol that reveals mostly SUMO1-ylated live sperm. Recently, with another protocol, it has been demonstrated that SUMO is expressed in most sperm and is related to poor morphology and motility, suggesting that sumoylation may have multiple roles depending on its localisation and targets. We show herein, by confocal microscopy and co-immunoprecipitation, that dynaminrelated protein 1 (DRP1), Ran GTPase-activating protein 1 (RanGAP1) and Topoisomerase IIa, SUMO1 targets in somatic and/or germ cells, are SUMO1-ylated in mature human spermatozoa. DRP1 co-localises with SUMO1 in the mid-piece, whereas RanGAP1 and Topoisomerase IIa in the post-acrosomal region of the head. Both SUMO1 expression and co-localisation with the three proteins were significantly higher in morphologically abnormal sperm, suggesting that sumoylation represents a marker of defective sperm. DRP1 sumoylation at the mid-piece level was higher in the sperm of asthenospermic men. As in somatic cells, DRP1 sumoylation is associated with mitochondrial alterations, this protein may represent the link between SUMO and poor motility. As SUMO pathways are involved in responses to DNA damage, another aim of our study was to investigate the relationship between sumoylation and sperm DNA fragmentation (SDF). By flow cytometry, we demonstrated that SUMO1-ylation and SDF are correlated (rZ0.4, P!0.02, nZ37) and most sumoylated sperm shows DNA damage in co-localisation analysis. When SDF was induced by stressful conditions (freezing and thawing and oxidative stress), SUMO1-ylation increased. Following freezing and thawing, SUMO1-Topoisomerase IIa co-localisation and co-immunoprecipitation increased, suggesting an involvement in the formation/repair of DNA breakage.

Sperm DNA fragmentation in cryopreserved samples from subjects with different cancers

“…Although ideal control subjects should be healthy normozoospermic men, the control group in the present study consisted of men who were cryopreserving samples because of oligozoospermia or for autoimmune pathologies, which may affect SDF. However, we recently evaluated PI brighter SDF levels in cryopreserved spermatozoa from normozoospermic men as part of a different project and found similar values to those observed in the two non-cancer groups in the present study (Marchiani et al 2014b).…”

Expression, Localization of SUMO-1, and Analyses of Potential SUMOylated Proteins in Bubalus bubalis Spermatozoa

“…Sumoylation of sperm proteins at various stages of spermatogenesis was previously demonstrated in other species, including humans and rodents (Rogers et al, 2004; Vigodner and Morris, 2005; Vigodner et al, 2006, 2013; Brown et al, 2008; Marchiani et al, 2011, 2014; Stanton et al, 2012). However, the role of SUMO in spermatogenesis is as yet undefined, with involvement in processes, including heterochromatin organization, DNA repair mechanisms, and regulation of gene expression, all being suggested (Rogers et al, 2004; Vigodner et al, 2006; Stanton et al, 2012).…”

“…The Bull spermatozoa were frozen in liquid nitrogen and, for the experiments, were thawed out in a water bath at 37°C and were separated by percoll density gradient centrifugation, as described previously (Marchiani et al, 2014). …”

“…While the role of SUMO in spermatogenesis remains undefined, it is suggested to be involved in heterochromatin stability, DNA repair mechanisms and the regulation of gene expression (Rogers et al, 2004; Vigodner et al, 2006; Stanton et al, 2012; Marchiani et al, 2014). SUMO1 expression is demonstrated in a high percentage of live, mature human sperm (Marchiani et al, 2011), However, excessive levels of sumoylation may be linked to an abnormal sperm morphology and motility (Vigodner et al, 2013; Marchiani et al, 2014) and to sperm DNA fragmentation (Marchiani et al, 2014). The localization of SUMO in sperm and its target proteins defined its promising role in different circumstances.…”

“…The localization of SUMO in sperm and its target proteins defined its promising role in different circumstances. Targets that have been identified in mature human sperm include the mitochondrial protein DRP1, the microtubule-organizing protein RanGAP1 and topoisomerase IIa, which is involved in chromatin remodeling (Marchiani et al, 2014). More studies should be conducted on the proteomic profile of sperm, particularly post-transcriptional protein modifications such as SUMOylation, which might give new insights to resolving infertility issues.…”