Sperm proteomics: road to male fertility and contraception

Abstract

Spermatozoa are highly specialized cells that can be easily obtained and purified. Mature spermatozoa are transcriptionally and translationally inactive and incapable of protein synthesis. In addition, spermatozoa contain relatively higher amounts of membrane proteins compared to other cells; therefore, they are very suitable for proteomic studies. Recently, the application of proteomic approaches such as the two-dimensional polyacrylamide gel electrophoresis, mass spectrometry, and differential in-gel electrophoresis has identified several sperm-specific proteins. These findings have provided a further understanding of protein functions involved in different sperm processes as well as of the differentiation of normal state from an abnormal one. In addition, studies on the sperm proteome have demonstrated the importance of spermatozoal posttranslational modifications and their ability to induce physiological changes responsible for fertilization. Large-scale proteomic studies to identify hundreds to thousands of sperm proteins will ultimately result in the development of novel biomarkers that may help to detect fertility, the state of complete contraception, and beyond. Eventually, these protein biomarkers will allow for a better diagnosis of sperm dysfunctions and aid in drug development. This paper reviews the recent scientific publications available from the PubMed database to address sperm proteomics and its potential application to characterize male fertility and contraception.

Figure 1

Schematic diagram showing the cellular, genetic, and chromatin changes during spermatogenesis. (a) The figure represents the cellular changes during spermatogenesis coupled with its genetic modifications. Following spermatogenesis, the male primordial germ cells, spermatogonia, first differentiate to primary spermatocytes and undergo genetic recombination to produce haploid round spermatids. The round spermatids then participate in another differentiation process to produce the mature spermatozoa. For successful fertilization, mammalian spermatozoa undergo first capacitation and then the acrosome reaction, which collectively allows spermatozoa to penetrate the zona pellucida and to fuse with the oocyte plasma membrane both in vivo and in vitro. (b) The figure shows chromatin changes that cause the transition of the mammalian nucleohistone to nucleoprotamine [17, 18]. In addition, the Donut-Loop model has been inserted at the end to represent the internal structure of the protamine-DNA fibers inside the toroid [17, 18]. The red color indicates histones and the blue color indicates DNA.

Figure 2

Schematic diagram showing the extraction and analysis of proteins from sperm cells. After collection and liquefaction of a whole semen sample, it can be used as it is or it can be subjected to density gradient centrifugation to separate mature and immature sperm populations. The recovered spermatozoa can be extracted and purified, after which the protein concentration can be determined in order to allow for an equal protein-loading gradient on electrophoretic gels. The further analysis of the sperm proteome is done from an aliquot of the sperm cells by using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) followed by excision of protein spots from the 2D-PAGE gels and matrix-assisted laser desorption/ionization-mass spectroscopy (MALDI-MS) or tandem MS (MS/MS) identification. Alternatively, the initial protein extract can be digested into peptides, which can be separated via liquid chromatography, followed by MS/MS analysis.

Figure 3

Diagram showing a mature human spermatozoon with its different parts. Previously published studies were used as a reference to summarize the list of proteins localized in different regions of the spermatozoon by immunolocalization. Proteins localized in acrosome, head, midpiece, and tail are mainly involved in capacitation and acrosomal reaction, spermatozoa-oocyte interaction (zona binding), energy production and metabolism, and motility and metabolism, respectively. The localization of a single protein at multiple sites in the sperm cell indicates its potential involvement in different physiological processes of the spermatozoon. The listed proteins include protein kinase A (PKA), phospholipase C zeta (PLC zeta), sperm head- and tail-associated protein (SHTAP), calicin, cylicins I and II, actin-capping proteins (ACP), enolase 1 (ENO1), cysteine-rich secretory protein 2 (CRISP2), arginine vasopressin receptor (AVPR), voltage-dependent anion channel (VDAC), glyceraldehyde 3-phosphate dehydrogenase (GAPDH), glutathione peroxidase 4 (GPX4), equatorial segment protein (ESP), heat shock protein 70 (HSP70), novel pyruvate kinase (PK-S), sperm acrosome membrane-associated protein 1 (SPACA1), izumo sperm-egg fusion 1 (IZUMO1), aquaporin 7 (AQPs), and the actin-related protein 2/3 complex (Arp2/3 complex).