1990-1994: Veterinary School, Maisons-Alfort, France.
June 1994: Diploma of Veterinary Studies.
1994-1995: Master degree (D. E. A. d'Endocrinologie et Interactions cellulaires).
1997: Doctorate of Veterinary Studies. Thesis entitled: "Myogenesis in vertebrates: embryological and molecular aspects."
1995-2000 : PhD in Prof. Margaret Buckingham's laboratory - Thesis entitled: "YAC attack of the transcriptional regulation of the myogenic determination factor Myf5 in the mouse embryo".
Research experience, relevant scientific techniques and skills acquired
February-March 1992: training in the laboratory of Prof. P. Tiollais: subcloning and sequencing.
March-April 1994: training in the laboratory of Prof. J-J. Panthier (Veterinary National School of Alfort, Maisons-Alfort, France): micro-injection of DNA in the pronucleus of mouse eggs and mice genotyping.
April-August 1994: training for obtention of the Diploma of Veterinary Studies in the laboratory of Prof. M. Buckingham (Pasteur Institute, Paris, France): subcloning, cell culture (transfection), transgenesis, mice breeding and genotyping (PCR, Southern blot).
September 1994-September 1995: predoctoral training in the same laboratory: YAC technology (isolation and manipulation).
October 1995-November 2000: doctoral training in the same laboratory: YAC manipulation and transgenesis; analysis of transgenic embryos: histology, in situ hybridisations, quantitative RT‑PCR.
November 2000-January 2001: post-doctoral training in the same laboratory
January 2001-December 2002: post-doctoral training in the laboratory of Prof. J. Mullins (The University of Edinburgh, Edinburgh, UK): BAC manipulation, transgenesis, in situ hybridisation, immunohistochemistry, immunoblotting, qRT-PCR, renal and cardiovascular phenotyping of mouse mutants.
(1) Analysis of salt and water homeostasis in the Ren-1d mouse mutants.
(2) Role of Cited2, an ubiquitous CREB/p300 cofactor, in adrenal development
December 2002-October 2013: Research Associate (CR2 then CR1, INSERM) in the laboratory of Prof. X. Jeunemaitre: Roles of WNK1 and WNK4 in cardiovascular and renal pathophysiology: BAC manipulation, generation of conditional mouse models of human hypertension syndromes, renal, cardiovascular and molecular phenotyping of mouse models.
Experimental analysis of Familial Hyperkalemic Hypertension, a rare form of human hypertension
Arterial hypertension is a complex trait caused by many environmental and genetic factors playing together and in interaction. This complexity has limited the identification of susceptibility genes for this major cardiovascular risk factor to controversial loci and candidate genes. Conversely, search for major genes implicated in rare Mendelian forms of the disease has been particularly successful. Up to now, the genes identified were corresponding to products already known or suspected to play a role in blood pressure regulation. The story initiated by Lifton’s group and ours is of another nature since it ended up with the identification of completely unsuspected genes. Familial hyperkalemic hypertension (FHHt) syndrome, also referred to as Gordon syndrome or pseudohypoaldosteronism type 2, is a rare inherited form of hypertension associated with hyperkalemia and hyperchloremic metabolic acidosis despite a normal glomerular filtration rate (OMIM no. 145260). The clinical data suggested that this syndrome could be due to an altered ionic transport in the renal distal tubule. At least four different genes are responsible for FHHt, suggesting a set of related disorders. In collaboration with Dr Lifton’s group, we have identified the genes corresponding to two of the four loci, responsible for the disease in seven families out of the 80 collected up to now.
These two genes, WNK1 and WNK4, belong to a new family of serine/threonine kinases, the WNK family for With No lysine (K), whose potential role in the maintenance of blood pressure was previoulsy unsuspected. The mutations found at the WNK1 locus in two FHHt kindreds correspond to large deletions of the first intron of the gene (22 and 41kb in a 60kb intron) and lead to the overexpression of WNK1. Our group has shown that WNK1 is a complex gene giving rise to two groups of isoforms: long ubiquitous isoforms (L-WNK1), containing the whole kinase domain, and a kidney-specific isoform (KS-WNK1), lacking the majority of the kinase domain and strongly expressed in the Distal Convoluted Tubule (DCT).
Several in vitro studies showed that WNK4 and L-WNK1 could activate or inhibit a number of ionic transporters and channels, such as the NaCl cotransporter NCC or the potassium channel ROMK. Furthermore, they showed that KS-WNK1 probably acts as a dominant-negative, preventing L-WNK1 action. These in vitro studies have been confirmed by the characterisation of two transgenic lines bearing a mutated WNK4. Overexpression of mutant WNK4 led to an increased activity of NCC, accompanied by hypertension and hyperkalemia. Inactivation of L-WNK1 led to embryonic death before E13 in L-WNK1-/- embryos and to a decrease in blood pressure in L-WNK1+/- mice that was not due to an ionic renal disturbance. This mouse model suggested that L-WNK1 could regulate blood pressure through its action in extra-renal tissues. We have shown that L-WNK1 is expressed in the entire vascular tree and in the heart. We hypothesised that this cardiovascular expression could participate in the early lethality observed in L-WNK1-/- embryos and the extra-renal origin of the decreased blood pressure observed in L-WNK1+/- adult mice.
The aim of my team's projects are to understand the molecular and pathophysiological mechanisms by which the deletion of WNK1 first intron leads to FHHt through the molecular and phenotypical analysis of several transgenic mouse lines.
We first showed that deletion of the first intron of mouse WNK1 leads to the misexpression of L-WNK1 and KS-WNK1 not only in the kidney but also in extra-renal tissues, especially in the cardiovascular system. These results suggest that WNK1 intron 1 contains one or several regulatory sequence(s) of WNK1 isoforms expression. Cross-species sequence comparison of WNK1 first intron and transient transfection assays allowed us to identify two intronic regulatory sequences of WNK1 expression.
As a first step towards the understanding of the role played by WNK1 in the regulation of blood pressure through its action on ionic transport in the kidney, we generated animals carrying a null-allele of KS-WNK1. These mice do not developp a hyperkalemic hypertension despite the activation of NCC. However, analysis of the expression of several sodium and potassium channels and transporters such as ENaC and ROMK suggested that sodium reabsorption and potassium secretion is modified in the DCT of KS-WNK1-/- mutant mice and that compensatory mechanisms are put in place in downstream nephron segments to prevent the development of hyperkalemic hypertension. Our results therefore suggest that FHHt could be a consequence of a dysregulation of ionic transport not only in the DCT, as suspected from the clinical data and suggested from the analysis of WNK4 transgenic mice, but also in the downstream nephron segments.
In conclusion, genetic and experimental analysis of Familial Hyperkalemic Hypertension, a very rare form of human hypertension, allowed the discovery of a new key player in blood pressure regulation, WNK1. Continuing the genetic analysis of non-WNK1 related FHHt cases could therefore lead to the identification of another unsuspected regulator of blood pressure. The original and unexpected results we obtained on the involvement of WNK1 in the regulation of vasoconstriction have modified our point of view on the pathophysiological mechanisms leading to FHHt in humans since they suggest that this syndrome could not only be a renal syndrome. In particular, they could explain why there is a delay between the apparition of the metabolic disorders and the hypertension in FHHt patients. In accordance with the clinical data, we have shown in vivo that the kidney-specific isoform of WNK1 is involved in the regulation of sodium and potassium balance in the DCT. However, in contradiction with the clinical data, our results suggest that FHHt is a pathology not only of the DCT but also of the downstream nephron segments. More largely, the new pathway for ionic transport regulation that we have contributed to identify could be affected in the more common forms of hypertension.