Proteins of the conserved Mep-Amt-Rh superfamily, including mammalian Rhesus factors,<p>mediate ammonium transport. Ammonium is an important nitrogen source for the<p>biosynthesis of amino acids for instance but its accumulation is also known as cytotoxic in<p>animals. Nevertheless, the controlled disposal of ammonium in urine plays a critical role in<p>the regulation of the acid-base homeostasis. Alteration in ammonium transport via human Rh<p>proteins could have clinical outcomes. In this work, we addressed aspects of structurefunction<p>analysis of altered human Rhesus proteins using a heterologous expression system<p>and further characterized aspects of the patho-physiological roles of Rh proteins using<p>knockout mice models available in the laboratory.<p>Using a yeast-based expression assay, we characterized human Rh variants resulting from non<p>synonymous single nucleotide polymorphisms (nsSNPs) with known or unknown clinical<p>phenotypes. The HsRhAG variants (I61R, F65S) associated to overhydrated hereditary<p>stomatocytosis (OHSt), a disease affecting erythrocytes, proved affected in intrinsic<p>bidirectional ammonium transport, suggesting altered ammonium transport as a potential<p>hallmark of the disease. Moreover, these variants showed trans-dominant negative effects on<p>the activity of their native HsRhAG counterpart, suggesting altered cooperation of the<p>subunits in “heteromeric” transport complexes. On the other hand, we revealed that the<p>R202C variant of HsRhCG, the orthologue of mouse Rhcg required for optimal urinary<p>ammonium excretion and blood pH control, shows an impaired inherent ammonium transport<p>activity. HsRhCGR202C may potentially confer susceptibility to disorders leading to metabolic<p>acidosis for instance. <p>MmRhcg has been shown to be expressed in the male mice epididymal tract, its absence<p>leading to a more acidic luminal fluid and to a reduced male fertility. Using mice<p>models, we further investigated the role of Rhcg and Rhbg proteins in the male<p>reproductive function. <p> / Doctorat en Sciences / info:eu-repo/semantics/nonPublished

Les reins jouent un rôle majeur dans la régulation de l'homéostasie acide-base. La production et l'excrétion rénale d'ammonium assurent environ deux tiers de l'excrétion nette d'acides dans les conditions normales et jusqu'à 90% en situation d'acidose métabolique. Bien que l'excrétion d'ammonium soit un processus finement régulé et capital au maintien de l'équilibre acido-basique, à ce jour aucun transporteur spécifique d'ammonium n'a été décrit chez les mammifères. Les protéines Rhésus Rhcg et Rhbg, membres distants de la famille des transporteurs d'ammonium Mep/Amt, semblaient de bons candidats à cette fonction. L'invalidation du gène Rhcg puis celle de Rhbg ont donc été entreprises chez la souris afin d'estimer leur rôle potentiel dans le transport de l'ammonium. / Doctorat en sciences, Spécialisation biologie moléculaire / info:eu-repo/semantics/nonPublished