Proteomics of Spermatogenesis
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Charles Pineau 1 AuthorId : Author. Hide details. Abstract : The strikingly complex structural organization of the mammalian testis in vivo creates particular difficulties for studies of its organization, function and regulation. These difficulties are particularly pronounced for investigations of the molecular communication networks within the seminiferous tubules that govern spermatogenesis.
The use of classical molecular and cell biology approaches to unravel this complexity has proved problematic, due to difficulties in maintaining differentiated germ cells in vitro, in particular. The lack of a suitable testing ground has led to a greater reliance on high-quality proteomic and genomic analyses as a prelude to the in vitro antx1d in vivo testing of hypotheses.
In this study, we highlight the options currently available for research, as used in our laboratory, in which proteomic and integrative genomic strategies are applied to the study of spermatogenesis in mammals. We will comment on results providing insight into the molecular mechanisms underlying normal and pathological spermatogenesis and new perspectives for the treatment of male infertility in humans. Finally, we will discuss the relevance of our strategies and the unexpected potential and perspectives they offer to teams involved in the study of male reproduction, within the framework of the Human Proteome Project.
This work introduces some of the integrative genomic concepts and works used by our team to gain new insight into mammalian spermatogenesis, a remarkably sophisticated process. We demonstrate the relevance of these integrative approaches to understand the cellular cross talks established between the somatic Sertoli cells and the germ cell lineage, within the seminiferous epithelium.
Our work also contributes to new knowledge on the pathophysiology of testicular function, with promising clinical applications. Proteome definition: The proteome has been defined as the protein complement of the genome. However, the definition of proteome has changed since it was first defined by Wilkins et al. Today, the term 'proteome' has developed to be: "The proteome of an individual is defined by the sum and the time dynamics of all protein species occurring during the life-time of this individual".
This definition of proteome includes the protein expression of the individual protein, the isoforms of a protein and post-translational modifications of a protein However, no protein list was published.
Linking transcriptomics and proteomics in spermatogenesis
The only far-reaching human sperm proteome analysis available to date is work done by Baker et al. Additionally; 2-DE had been used in 8 studies to map human sperm proteome. To our best knowledge, no other techniques had been used for proteome profiling of human spermatozoa, including multidimensional protein identification technique MudPIT or combined fractional diagonal chromatography COFRADIC technique. A more extensive human sperm proteome could be obtained by combining different MS proteomics techniques. We have shown that different MS proteomics techniques are able to identify a unique set of proteins How many proteins are expressed in human sperm?
The near-complete sequencing of the human genome has yielded the total gene estimates that, at first glance, seem surprisingly low; of the order of open reading frames 27, However, when a gene is expressed it is subjected to alternative splicing mechanisms and post-translational modifications. It is estimated each gene could produce between 5 to 6 mRNAs by an alternative splicing mechanism and each of these mRNA species is in turn translated into proteins that are processed in various ways, generating on the order of 8—10 different modified forms of each polypeptide chain.
Defining each and every one of these proteins is what global collaborations, such as the Human Proteome Organization HUPO is set to undertake.
The question 'how many proteins, the most highly differentiated and unique cell type in the human body, the spermatozoa, contain? Of course, it is quite difficult to predict the size of the human spermatozoa proteome from the existing proteomics data, knowing the current limitation of MS proteomics technology 26, However, Baker et al 30 used the current proteomics data available from yeast proteome to predict the number of protein species of the human spermatozoa to be As Baker et al.
However, Baker et al. For any gene or protein list, DAVID software tools are able to identify enriched biological themes, particularly GO terms, discover enriched functional-related gene groups, visualize genes or proteins on BioCarta and KEGG pathway maps, explore gene or protein names in batch, link genedisease associations, etc.
Biological function of human sperm proteome: Table 1 shows the ten most important catalogue outputs for biological function analysis by DAVID software DAVID software was only able to catalogue of the submitted proteins. This means that biological functions of about proteins out of the collected human sperm proteome are still unknown.
DAVID categorized glucose catabolic processes and oxidative phosphorylation which is necessary for the homeostasis. In the table, we also find proteins belonging to spermatogenesis 3. Cellular component of human sperm proteome: Table 2 shows the top ten outputs of cellular localization of the collected human sperm proteome from DAVID software.
[PDF] Linking transcriptomics and proteomics in spermatogenesis. - Semantic Scholar
The software was able to map of the identified proteins. It is well known that the human sperm lost most of its cytoplasm during spermiogensis process. A large number of proteins were categorized to be from mitochondria. Mitochondrial protein is not astonishing since the neck of human sperm is rich in mitochondria.
Additionally, protein enriched parts belonging to the tail of human sperm were identified as cytoskeleton As it is shown in Table 2 no transmembrane proteins were categorized from the collected human sperm proteome which are important types of proteins for the oocyte and sperm interaction.
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This probably is caused by MS proteomics techniques used for the proteome mapping of human sperm. It is a well-known fact that the hydrophobic proteins, such as transmembrane proteins, rarely appear in gel-based techniques Using gel-free techniques, such as MudPIT, will improve the deeper coverage of human sperm proteome.
However, MudPIT is not a straightforward technique and it needs some expertise 38, Functional categorization of the collected human sperm proteome: The most statistically significant functional annotation by DAVID software were the actylated proteins However, the exact function of these large numbers of post-translational modifications is unknown personal communication with Baker M, author of the largest human sperm proteome published to date Observing the enrichment of fatty acid and pyruvat metabolism is not surprising since sperm is under hypoxic condition.
Sperm: a silent cell? Martinez-Heredia et al 21 identified transcrip-tion factor proteins in the proteome mapping of human sperm using 2-DE technique, in the pI range