The mechanism of gamete fusion remains largely unknown on molecular level despite its indisputable significance. A few molecules requisite for membrane interaction are known, among them IZUMO1 on sperm and tetraspanin CD9 on egg. A concept of a large multitprotein complex on both membranes forming a fusion machinery is recently emerging. CD9 is expected to play a role in organizing egg membrane order and to interact laterally with other factors. On the other hand, IZUMO1 contains one immunoglobulin (Ig) domain, which is known for its interaction with various molecules and thus is expected to have a binding partner on the egg membrane. Ig domains are known for their involvement in interactions with Fc receptors and Fc receptor-like molecules, which makes these a potential partner for sperm in gamete binding and fusion. Recent experiments identified Fc receptor-like 3 as a candidate binding partner for sperm by one-bead one-compound assay. The aim of presented study was to assess the localization of FCRL3 in mouse eggs, together with its potential co-localization with IZUMO1 on complexes of zona-free eggs with bound sperm. In this study, FCRL3 molecule has been found across the cytoplasm, possibly in the intracytoplasmic membrane compartments of the cell, as well as on the egg membrane. Its presence...

The Testis Specific Serine Kinase (Tssk) family of proteins is a large group of kinases that present high level of conservation within paralogs, as well as within species. In addition, in all reported cases as well as in the analysis of expressed sequence tags available in databases, this family of proteins presents a very strict pattern of either testicular or male-gonadal expression. This high level of conservation prompted the postulate that these kinases ought to be important for either testicular function or fertilization. In this work we attempt a biochemical characterization of one family member (Tssk6) in the mouse. We also analyze the male infertility phenotype presented by mice null for Tssk6 revealing its requirement for actin dynamics and the relocalization of proteins necessary for gamete fusion. In this analysis we described Tssk6 as the second protein known to date to be necessary in the sperm for gamete fusion to take place. We also examined a novel member of the Tssk family in the mouse as well as ortholog proteins in two invertebrates (C. elegans and D. melanogaster). Although our understanding of the function, activity and regulation of these kinases remains small, this work constitutes a significant advance towards the understanding of the identity of the Tssk family. The results that follow have far reaching effects that surpass the realm of the Tssk family. They influence the study of sperm biological processes like the changes in sperm cytoskeletal structures and the acrosome reaction. They also influence the field of developmental biology and scientist working in the molecular characterization of the process of gamete fusion and zygote formation. Lastly, the work here presented influences as well evolutionary developmental biology through the study of a highly conserved family of proteins that is essential for reproduction and could play a role in the process of speciation.

Sperm-­-egg membrane interaction and fusion is mediated by various molecules of the different protein network that are located on both egg and sperm membrane. So far, many proteins have been selected to be fusion candidates, some of them (Izumo1, CD9, Juno) were proven to be essential, whereas others were discovered to play an unsuspected new active role (CD46, tetraspanins). After the adhesion of sperm to an egg, Juno located on the oolema associates with monomeric Izumo1 on sperm membrane, which is results in Izumo1 dimerization following quick removal of Juno from the egg surface as described in mouse. It implies that additional receptor on the egg membrane is required to play a role in sperm-­-egg fusion. To find a human fusogenic receptor for IZUMO1 protein we used one-­-bead-­-one-­-compound (OBOC) assay, a random screening approach. A bead, fulfilling all the requirements when interacting with the human sperm, carried a peptide sequence showing homology with the conserved Ig domain of the human specific Fc receptor-­-like protein 3 (FcRL3). In general, the ...