In-depth proteomic analysis of the human sperm reveals complex protein compositions

Abstract

The male gamete (sperm) can fertilize an egg, and pass the male genetic information to the offspring. It has long been thought that sperm had a simple protein composition. Efforts have been made to identify the sperm proteome in different species, and only about 1000 proteins were reported. However, with advanced mass spectrometry and an optimized proteomics platform, we successfully identified 4675 human sperm proteins, of which 227 were testis-specific. This large number of identified proteins indicates the complex composition and function of human sperm. Comparison with the sperm transcriptome reveals little overlap, which shows the importance of future studies of sperm at the protein level. Interestingly, many signaling pathways, such as the IL-6, insulin and TGF-beta receptor signaling pathways, were found to be overrepresented. In addition, we found that 500 proteins were annotated as targets of known drugs. Three of four drugs studied were found to affect sperm movement. This in-depth human sperm proteome will be a rich resource for further studies of sperm function, and will provide candidate targets for the development of male contraceptive drugs.

Introduction

Approximately one in six couples experience difficulty in conceiving a child. Male infertility accounts for about half the cases in which assisted reproductive techniques are recommended [1]. Over 85% of infertile male can actually produce sperm [2]; however, for some reason, those sperm are often unable to fertilize an egg.

As the only cell performing its function outside the male human body, sperm is a highly specialized cell with distinct morphological and compositional differences compared with other somatic and germ cells [3]. It was long believed that the function of sperm was only to deliver the paternal genomes to the egg. However, recent studies have shown that sperm can deliver a complex set of RNAs to the egg [4]. In addition, the entire cell, including the midpiece and tail, enters the egg in most species [5]. Current studies have suggested that sperm defects can disrupt embryo development, even if the genome carried by the cells is perfectly normal [6]. Thus, characterization of the protein composition of sperm can help better understand sperm function.

The recently proposed Chromosome-Centric Human Proteome Project (C-HPP) aims to define the full set of proteins encoded in each chromosome. The initial goal of the C-HPP is to identify at least one representative protein encoded by each of the approximately 20,300 human genes [7]. The genes and proteins had tissue-dependent expression. According to previous analyses, the human testis, the male gonad producing sperm, contains the largest number of tissue-specific genes across the 31 human tissues [8]. Thus, in order to characterize all the proteins, proteins in the testis and/or testicular cells should be carefully studied. The in-depth proteomic analysis of human sperm will produce data important for C-HPP.

To date, some efforts have been made to identify the human sperm proteome, and in fact, a list of 1056 proteins has been reported [9]. Additionally, Johnston et al. [10] claimed identification of 1760 proteins in human sperm, but this protein list is not available. Efforts to identify the sperm proteome in other species, including drosophila and mammals, such as rat and mouse [11], [12], have also been made, and the numbers of proteins identified were all around or below one thousand. In the present study, using the advanced mass spectrometry and an optimized proteomics platform, we successfully identified 4675 unique proteins from human sperm, which showed the complex composition of human sperm.