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.

Chez les procaryotes, les protéines sont synthétisées dans le cytoplasme avant d'être transportés vers différentes destinations, intra- ou extra-cellulaires. Les bactéries Gram-négatives ont mis au point une grande collection de mécanismes et systèmes, appelés systèmes de sécrétion bactérienne, pour sécréter des protéines à travers leur paroi cellulaire vers l'extérieur. Le système de sécrétion de type VI, identifié dans les années 2006-2008, est une nano-machine polyvalente répandue chez les bactéries pathogènes. Il y a de nombreuses preuves que T6SS injecte des protéines toxiques (effecteurs) directement dans les cellules eucaryotes et procaryotes pour les tuer. Pour empêcher la destruction cellules provenant de la même espèce, les bactéries possédant un T6SS produisent également des protéines d'immunité qui neutralisent les effets toxiques des effecteurs de leurs congénères. Le T6SS est formé de 13 composants de coeur (nommés TssA-M) en une structure souvent comparée à un "bactériophage inversé". La queue, semblable à celle de phages, a une forme tubulaire (la gaine et le tube interne) et polymérise à partir d'une plaque basale ancrée sur un complexe membranaire trans-périplasmique. La contraction de la gaine fournit l'énergie nécessaire pour propulser le tube intérieur à travers la paroi vers les cellules proies. Dans le cadre de ma thèse, je me suis impliqué dans la détermination de la structure et la dynamique de certains composants du T6SS de la d’Escerichia coli enteroaggrégatif (EAEC). Plusieurs structures ont été déterminées et analysées. Quatre articles ont été publiés et deux autres sont en préparation. / In prokaryotes, proteins are synthesized in the cytoplasm before being transported to various destinations, intra- or extra-cellular. Gram-negative bacteria have developed a large collection of mechanisms and systems, termed bacterial secretion systems, to secrete proteins through their cell wall to the exterior. The type VI secretion system, identified in years 2006-2008, is a versatile nano-machine prevalent in pathogenic bacteria. There have been many evidences that T6SS delivers toxic proteins directly into both eukaryotic and prokaryotic cells to kill them. To prevent killing of sibling cells (cells from the same species), T6SS+ cells produce also immunity proteins that neutralize the toxic effects of their cognate effectors. T6SS contains 13 core-components (TssA-M), assembling a structure often quoted as an “inverted bacteriophage”. A phage-like tubular tail (the sheath and the internal tube) polymerizes from a baseplate-like complex, anchored to the cell internal and outer membranes via a membrane anchored complex spanning the periplasm. Contraction of the sheath provides the necessary energy to propel the internal tube through the wall towards the prey cells. In the framework of my PhD, I became involved in determining the structure and dynamics of some components of the EAEC sci1 T6SS, mostly on the membrane and baseplate subcomplexes. Several structures have been determined and analysed. Four articles have been published and two other are in preparation.