Thèse numérisée par la Direction des bibliothèques de l'Université de Montréal.

5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB) was normally recognised as a Cl- channel inhibitor, but its specificity is in question, since an inhibitory effect against K⁺ channels has been reported. To identify the significance of the molecules structural components, NPPB and related compounds, such as 2-(3-phenylpropylamino) benzoic acid (PPAB), 5- nitro-2-heptylamino benzoic acid (HANB) and 2-nitro-5-heptylamino benzoic acid (HANB-2) were synthesised by reductive amination using various aldehydes and amines. Using internodal cells of the giant green Characean algae, Nitella hookeri, the effects of NPPB and related compounds on cytoplasmic streaming and turgor regulation were determined. Previous experiments stated that cytoplasmic streaming was sensitive to NPPB, PPAB and HANB with IC₅₀ values of 24µmol/L, 455µmol/L, and 6.4mmol/L, respectively. In this report, the IC₅₀ values of purchased NPPB and niflumic acid were found to be 88.65µmol/L and 121.82µmol/L, respectively. Although the IC₅₀ value of purchased NPPB showed a slight difference from that of synthesised NPPB, the results of the cytoplasmic streaming experiment indicated the possibility of this analysis to be a simple assay system for analysing the effects of structural modification to ion channel inhibitors on their biological activity. Moreover, NPPB and PPAB seem to stimulate regulation of turgor pressure under hyperosmotic shock, which can be explained by a blockage of K⁺ efflux during osmotic stress leading to faster recovery of turgor regulation. Additionally, the results of cytosolic free Ca²⁺ analysis using aequorin technology also suggested that the possibility of this analysis to be used as a more direct measure of the inhibitory effect, while the cytoplasmic streaming analysis is a more indirect method. The preliminary results from this research suggest the significance of the simple assay systems for analysing the effects of structural modification ion channel inhibitors, which can be used for future study regarding ion channel structures.

5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB) was normally recognised as a Cl- channel inhibitor, but its specificity is in question, since an inhibitory effect against K⁺ channels has been reported. To identify the significance of the molecules structural components, NPPB and related compounds, such as 2-(3-phenylpropylamino) benzoic acid (PPAB), 5- nitro-2-heptylamino benzoic acid (HANB) and 2-nitro-5-heptylamino benzoic acid (HANB-2) were synthesised by reductive amination using various aldehydes and amines. Using internodal cells of the giant green Characean algae, Nitella hookeri, the effects of NPPB and related compounds on cytoplasmic streaming and turgor regulation were determined. Previous experiments stated that cytoplasmic streaming was sensitive to NPPB, PPAB and HANB with IC₅₀ values of 24µmol/L, 455µmol/L, and 6.4mmol/L, respectively. In this report, the IC₅₀ values of purchased NPPB and niflumic acid were found to be 88.65µmol/L and 121.82µmol/L, respectively. Although the IC₅₀ value of purchased NPPB showed a slight difference from that of synthesised NPPB, the results of the cytoplasmic streaming experiment indicated the possibility of this analysis to be a simple assay system for analysing the effects of structural modification to ion channel inhibitors on their biological activity. Moreover, NPPB and PPAB seem to stimulate regulation of turgor pressure under hyperosmotic shock, which can be explained by a blockage of K⁺ efflux during osmotic stress leading to faster recovery of turgor regulation. Additionally, the results of cytosolic free Ca²⁺ analysis using aequorin technology also suggested that the possibility of this analysis to be used as a more direct measure of the inhibitory effect, while the cytoplasmic streaming analysis is a more indirect method. The preliminary results from this research suggest the significance of the simple assay systems for analysing the effects of structural modification ion channel inhibitors, which can be used for future study regarding ion channel structures.

Thesis (M. Sc.)--University of Canterbury, 2006. / Typescript (photocopy). Includes bibliographical references (leaves 105-110). Also available via the World Wide Web.

Uremic pruritus (UP) arises in end-stage renal disease (ESRD) and is not relieved by proper dialysis. While the pathogenesis of UP is not well understood, UP responds poorly to anti-histamines. We performed a case-control study to test if cutaneous protease-mediated, non-histamine itch is augmented in UP, and if UP is associated with altered epidermal and/or papillary dermal innervation. We recruited 12 hemodialysis subjects with ESRD-specific itch (cases) (Visual Analogue Scale (VAS)-average itch in the preceding week, 78/100), and 13 age- and sex-matched hemodialysis subjects without pruritus (controls) (VAS- average itch in the preceding week, 0/100; p<0.0001 cases vs. controls). Cowhage spicule-induced itch was induced in the back where all subjects exhibited itch, and the entire duration of itch was measured with the general Labeled Magnitude Scale. Subsequently, a punch biopsy was taken from this sensory-tested skin and multi-label immunohistochemistry was performed to measure epidermal and papillary dermal innervation. In cases vs. controls, cowhage-induced area under the curve (AUC) for itch was significantly larger (median, 25%–75%: 175.4, 101.0–252.2 vs. 42.4, 24.0–160; p=0.04) as was perceived peak itch intensity (53.6, 53.3–78.9 vs. 34.2, 20.9–55.6; p=0.02). Cases showed a significant reduction in papillary dermal nerve length (PDNL)/mm epidermis (2295, 1659–2970 vs. 2909, 2228–3523; p=0.003), resulting from the loss of papillary dermal (PD)-calcitonin gene related peptide (CGRP) (+) nerves (p<0.0001), with preservation of %PD-substance P (+) nerves (p=0.1) and intraepidermal nerve fiber density (p=0.1). VAS-average itch in the preceding week negatively correlated with PDNL/mm epidermis (correlation coefficient (CC)=-0.53, p=0.003) and %PD-CGRP (+) nerves (CC=-0.37, p=0.03). Cowhage-induced AUC-itch negatively correlated with %PD-CGRP nerves only in cases (CC=-0.40, p=0.02). Our data suggest augmented protease-dependent signaling contributes to UP and indicate a mechanism for how PD-CGRP (+) nerve loss contributes to UP and augmented cowhage-itch: loss of an afferent skin-derived itch-inhibition signal to the spinal cord dorsal horn. / 2016-10-02T00:00:00Z

In this study, four anion transporter (AT) blockers, DIDS (4, 4′-diisothiocyanatostilbene-2, 2′-disulfonic acid), 9-AC (anthracene-9-carboxylic acid), NPPB (5-nitro-2-(3-phenylpropylamino) benzoic acid), and IAA-94 (indanyloxy acetic acid) were selected to evaluate ATs as potential target sites for insect and nematode control. All the AT blockers showed slowly developing toxicity against second-stage larvae of <i>Meloidogyne incognita</i> (Kofoid and White 1919) Chitwood 1949 and adults of <i>Caenorhabditis elegans</i> Maupas 1900 but not against third-stage larvae of <i>Heterorhabditis bacteriophora</i> Poinar 1975 even at 200 ppm. Symptoms of AT blocker toxicity observed in <i>C. elegans</i> adults were increased pharyngeal muscle contractions and decreased locomotion. Exposure of <i>C. elegans</i> as fourth-stage larvae to double-stranded RNA (dsRNA) of <i>ceclc-1</i> and <i>ceclc-2</i> (VGCC genes coding for CeClC-1 and CeClC-2, respectively) either alone or together for 24 h decreased their expression in F1 progeny in a time-dependent manner. Reduction in expression of <i>ceclc-2</i> alone or together with <i>ceclc-1</i> significantly increased pharyngeal contractions and decreased locomotion in significantly higher percentage of F1 progeny. The above findings suggested AT blockers nematicidal activity primarily comes from inhibition of CeClC-2 channels, while inhibition of CeClC-1 channels may enhance this activity. All the AT blockers showed slowly developing toxicity against adults of a susceptible strain (Oregon-R) of <i>Drosophila melanogaster</i> Meigen 1830, while DIDS, was equally toxic to dieldrin-resistant rdl flies. All AT blockers, except 9-AC, at 100 µM showed significant excitatory effect on desheathed central nervous system (CNS) of third-instar larvae of <i>Drosophila</i>, while DIDS showed a modest excitatory effect on ascending peripheral nerves. Feeding adult flies on 10% sugar solution mixed with 100 ppm of DIDS for 6 h decreased the midgut pH by 2 units approximately. All the AT blockers inhibited the growth of larvae (in weight), increased the developmental time, and decreased survival when <i>Ostrinia nubilalis</i> (Hübner 1796) second-instar larvae were fed for seven days. All the AT blockers decreased the midgut alkalinity and inhibited chloride ion transport from midgut lumen into epithelia in fifth-instar larvae when fed for 3 h on treated diet. Positive correlations observed among growth, midgut alkalinity, and midgut chloride transport in AT blocker-fed larvae suggested that inhibition of chloride/bicarbonate exchangers by AT blockers may have contributed to midgut alkalinity decrease affecting the digestion and resulting in observed lethal and sublethal effects. / Ph. D.

Made available in DSpace on 2014-06-12T15:54:28Z (GMT). No. of bitstreams: 2 arquivo6489_1.pdf: 1208262 bytes, checksum: c1b40f981f11c014c773e3922d544d81 (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) Previous issue date: 2011 / Conselho Nacional de Desenvolvimento Científico e Tecnológico / A caracterização de canais iônicos em células tronco é importante pelo fato destas estruturas atuarem no controle da proliferação e diferenciação destas células. Há poucos trabalhos focados nos canais iônicos presentes nas membranas plasmáticas das células tronco e nenhum deles trata das células tronco mesenquimais da geléia de Wharton do cordão umbilical humano (whMSC). O cordão umbilical é uma fonte importante de células tronco para tão esperada terapia celular. Estes fatos motivaram o estudo dos canais iônicos das whMSC como objetivo maior do presente trabalho. Foi utilizada a técnica de patch-clamp na configuração whole-cell e a atuação de inibidores específicos para identificar os canais iônicos presentes nas whMSC. Os inibidores escolhidos foram: tetraetilamonio (TEA) e 4-aminopiridina (4-AP) como inibidores de canais de potássio dependentes de voltagem e de Ca2+, Glibenclamida (GB) um inibidor de canais de potássio dependentes de ATP, canais aniônicos dependentes de volume celular e CFTR e 5-nitro-2-(3-phenylpropylamino) ácido benzóico (NPPB) um potente bloqueador dos canais de cloreto, mas que também é capaz de bloquear os canais de potássio dependentes de Ca2+ de condutância intermediária. Os resultados mostraram que as correntes iônicas ativadas por voltagem das whMSC podem ser bloqueadas consideravelmente por TEA (10 mM), 4-AP (5 mM) e NPPB (100 &#956;M) indicando a presença dos canais de K+ dependentes de voltagem e de Ca2+ e de canais de Cl- dependentes de voltagem conforme descrito para células tronco de outras origens. Utilizando GB (100 &#956;M) observamos um aumento (estimulação) significativo da corrente. Este é um resultado inédito, pois estudos anteriores demonstram que o GB atua inibindo a corrente iônica transmembrana. Em todos os casos, com todos os inibidores testados, os efeitos foram reversíveis. Estes resultados demonstram que as membranas citoplasmáticas das whMSC possuem canais de K+ dependentes de voltagem e de Ca2+ e canais de Cl- dependentes de voltagem. O surpreendente efeito da glibenclamida indica a presença de canais iônicos desconhecidos nas membranas das whMSC

Le cœur des vertébrés est un organe modulaire qui requiert le " patterning " complexe des champs morphogénétiques cardiogènes et la convergence coordonnée des diverses sous-populations de progéniteurs cardiogéniques. Au moins 7 facteurs de transcription de la famille T-box coopèrent au sein de ces nombreuses sous-populations de progéniteurs cardiogéniques afin de réguler la morphogenèse et l’agencement de multiples structures le long de l’ébauche cardiaque, ce qui explique que les mutations humaines de ces gènes engendrent diverses malformations congénitales cardiaques (MCCs). L’un de ces gènes T-box, Tbx5, dont l’haploinsuffisance génère le syndrome de Holt-Oram (SHO), intervient dans une grande variété de réseaux de régulation géniques (RRGs) qui orchestrent la morphogenèse des oreillettes, du ventricule gauche, de la valve mitrale, des septums inter-auriculaire et inter-ventriculaire, ainsi que du système de conduction cardiaque. La diversité des RRGs impliqués dans la formation de ces structures cardiaques suggère que Tbx5 détient une profusion de fonctions qui ne seront identifiables qu’en répertoriant ses activités moléculaires dans chaque lignée cardiaque examinée isolément. Afin d’aborder cette problématique, une ablation génétique de Tbx5 dans l’endocarde a été réalisée. Cette expérience a démontré le rôle crucial de Tbx5 dans la survie des cellules endocardiques bordant le septum primum et des cardiomyocytes au sein de cette structure embryonnaire qui contribuera à la morphogenèse du septum inter-auriculaire. En outre, cette étude a révélé l’existence d’une communication croisée entre la sous-population de cellules endocardiques Tbx5+ et le myocarde au niveau du septum primum, afin d’assurer la survie des cardiomyocytes, et ultimement de garantir la maturation du septum inter-auriculaire. Nos résultats confirment aussi l’importance de l’interdépendance génétique (Tbx5 et Gata4 ainsi que Tbx5 et Nos3) entre différents loci dans la morphogenèse de la cloison inter-auriculaire, et particulièrement de l’influence que peut avoir l’environnement sur la pénétrance et l’expressivité des communications inter-auriculaires (CIAs) dans le SHO. En outre, puisque les fonctions d’un gène dépendent ordinairement des différents isoformes qu’il peut générer, une deuxième étude a focalisé davantage sur l’aspect transcriptionnel de Tbx5. Cette approche a mené à la découverte de 6 transcrits alternatifs exhibant des fonctions à la fois communes et divergentes. La caractérisation de 2 de ces isoformes a révélé le rôle de l’isoforme long (Tbx5_v1) dans la régulation de la croissance des cardiomyocytes durant la cardiogénèse, tandis que l’isoforme court (Tbx5_v2), préférentiellement exprimé dans le cœur mature, réprime la croissance cellulaire. Il est donc entièrement concevable que les mutations de TBX5 entraînant une troncation de la région C-terminale accroissent la concentration d’une protéine mutée qui, à l’instar de Tbx5_v2, interfère avec la croissance de certaines structures cardiaques. En revanche, la divergence de fonctions de ces isoformes, caractérisée par les disparités de localisation subcellulaire et de d’interaction avec d’autres cofacteurs cardiaques, suggère que les mutations affectant davantage un isoforme favoriseraient l’émergence d’un type particulier de MCC. Finalement, un dernier objectif était d’identifier le ou les mécanisme(s) moléculaire(s) par le(s)quel(s) Tbx5 régule son principal gène cible, Nppa, et d’en extraire les indices qui éclairciraient sa fonction transcriptionnelle. Cet objectif nécessitait dans un premier lieu d’identifier les différents modules cis-régulateurs (MCRs) coordonnant la régulation transcriptionnelle de Nppa et Nppb, deux gènes natriurétiques dont l’organisation en tandem et le profil d’expression durant la cardiogénèse sont conservés dans la majorité des vertébrés. L’approche d’empreinte phylogénétique employée pour scanner le locus Nppb/Nppa a permis d’identifier trois MCRs conservés entre diverses espèces de mammifères, dont un (US3) est spécifique aux euthériens. Cette étude a corroboré que la régulation de l’expression du tandem génique Nppb/Nppa requérait l’activité transcriptionnelle d’enhancers en complément aux promoteurs de Nppa et Nppb. La concordance quasiment parfaite entre les profils d’expression de Tbx5 et de ces deux gènes natriurétiques chez les mammifères, suggère que le gradient d’expression ventriculaire de Tbx5 est interprété par le recrutement de ce facteur au niveau des différents enhancers identifiés. En somme, les études présentées dans cette thèse ont permis de clarifier la profusion de fonctions cardiaques que possède Tbx5. Certaines de ces fonctions émanent de l’épissage alternatif de Tbx5, qui favorise la synthèse d’isoformes dotés de propriétés spécifiques. Les diverses interactions combinatoires entre ces isoformes et d’autres facteurs cardiaques au sein des diverses sous-populations de progéniteurs cardiogènes contribuent à l’émergence de RRGs cardiaques divergents. / The vertebrate heart is a modular organ, which requires the complex patterning of the morphogenetic heart fields and the coordinated convergence of the diverse subpopulations of cardiogenic progenitors. At least 7 transcription factors of the T-box family cooperate within these numerous subpopulations of cardiogenic progenitors to regulate the morphogenesis and the layout of multiple structures along the primordial heart tube, which explains that the human mutations of these genes induce various congenital heart defects (CHDs). One of these T-box genes, Tbx5, whose haploinsufficiency generates the Holt-Oram syndrome (HOS), intervenes in a wide variety of gene regulatory networks (GRNs) that orchestrate the morphogenesis of the atria, the left ventricle, the mitral valve, the inter-atrial and inter-ventricular septa, as well as the cardiac conduction system. The diversity of GRNs involved in the formation of these cardiac structures suggests that Tbx5 holds a profusion of functions which will be identifiable only by indexing its molecular activities in each separately examined cardiac lineage. To address this problem, a conditional knockout of Tbx5 in the endocardium was generated. This experiment demonstrated a crucial role of Tbx5 in the survival of the endocardial cells lining the septum primum and the cardiomyocytes within this embryonic structure, which will contribute to the morphogenesis of the inter-atrial septum. Moreover, this study revealed a crosstalk between the Tbx5-positive endocardial cells subpopulation and the myocardium at the level of the septum primum to ensure the survival of cardiomyocytes, and ultimately to guarantee the maturation of the inter-atrial septum. Our results also confirmed the importance of genetic interdependence (Tbx5 and Gata4 as well as Tbx5 and Nos3) between different loci in the morphogenesis of the inter-atrial septum, and particularly the influence that the environment can have on the penetrance and the expressivity of atrial septal defects (ASDs) in the HOS. Besides, since the functions of a gene usually depend on the different isoforms it can generate, a second study focused more on the transcriptional aspect of Tbx5. This approach led to the discovery of 6 alternative transcripts exhibiting both common and specific functions. The characterization of 2 of these isoforms revealed the role of the long isoform (Tbx5_v1) in the regulation of cardiomyocytes growth during cardiogenesis, whereas the short isoform (Tbx5_v2), preferentially expressed in the mature heart, represses cell growth. It is thus entirely conceivable that TBX5 mutations leading to a C-terminal truncation increase the concentration of a mutated protein, which, like Tbx5_v2, interferes with the growth of certain cardiac structures. On the other hand, the divergence of functions of these isoforms, characterized by the disparities of subcellular localization and interaction with other cardiac cofactors, suggests that mutations affecting more one isoform would favor the emergence of a particular type of CHD. Finally, a last objective was to identify one or several molecular mechanism(s) by which Tbx5 regulates its main target gene, Nppa, and to extract clues that might clarify its transcriptional function. This objective required in a first place to identify the various cis-regulatory modules (CRMs) coordinating the transcriptional regulation of Nppa and Nppb, two natriuretic genes whose tandem organization and expression pattern during cardiogenesis are preserved in most vertebrates. The phylogenetic footprint approach employed to scan the Nppb/Nppa locus allowed the identification of three CRMs evolutionary conserved between different mammals species, one of which (US3) is specific to eutherians. This study confirmed that the regulation of the tandem genes Nppb/Nppa required the transcriptional activity of enhancers in complement to Nppa and Nppb promoters. The almost perfect concordance between the expression profiles of Tbx5 and these two natriuretic genes in mammals, suggests that the ventricular expression gradient of Tbx5 is interpreted by the recruitment of this factor to the identified enhancers. Altogether, the studies presented in this thesis allowed clarifying the profusion of Tbx5 cardiac functions. Some of these functions emanate from the alternative splicing of Tbx5, which favors the synthesis of isoforms endowed with specific properties. The diverse combinatorial interactions between these isoforms and other cardiac factors within the various cardiogenic progenitor subpopulations contribute to the emergence of distinct cardiac RRGs.

Le cœur des vertébrés est un organe modulaire qui requiert le " patterning " complexe des champs morphogénétiques cardiogènes et la convergence coordonnée des diverses sous-populations de progéniteurs cardiogéniques. Au moins 7 facteurs de transcription de la famille T-box coopèrent au sein de ces nombreuses sous-populations de progéniteurs cardiogéniques afin de réguler la morphogenèse et l’agencement de multiples structures le long de l’ébauche cardiaque, ce qui explique que les mutations humaines de ces gènes engendrent diverses malformations congénitales cardiaques (MCCs). L’un de ces gènes T-box, Tbx5, dont l’haploinsuffisance génère le syndrome de Holt-Oram (SHO), intervient dans une grande variété de réseaux de régulation géniques (RRGs) qui orchestrent la morphogenèse des oreillettes, du ventricule gauche, de la valve mitrale, des septums inter-auriculaire et inter-ventriculaire, ainsi que du système de conduction cardiaque. La diversité des RRGs impliqués dans la formation de ces structures cardiaques suggère que Tbx5 détient une profusion de fonctions qui ne seront identifiables qu’en répertoriant ses activités moléculaires dans chaque lignée cardiaque examinée isolément. Afin d’aborder cette problématique, une ablation génétique de Tbx5 dans l’endocarde a été réalisée. Cette expérience a démontré le rôle crucial de Tbx5 dans la survie des cellules endocardiques bordant le septum primum et des cardiomyocytes au sein de cette structure embryonnaire qui contribuera à la morphogenèse du septum inter-auriculaire. En outre, cette étude a révélé l’existence d’une communication croisée entre la sous-population de cellules endocardiques Tbx5+ et le myocarde au niveau du septum primum, afin d’assurer la survie des cardiomyocytes, et ultimement de garantir la maturation du septum inter-auriculaire. Nos résultats confirment aussi l’importance de l’interdépendance génétique (Tbx5 et Gata4 ainsi que Tbx5 et Nos3) entre différents loci dans la morphogenèse de la cloison inter-auriculaire, et particulièrement de l’influence que peut avoir l’environnement sur la pénétrance et l’expressivité des communications inter-auriculaires (CIAs) dans le SHO. En outre, puisque les fonctions d’un gène dépendent ordinairement des différents isoformes qu’il peut générer, une deuxième étude a focalisé davantage sur l’aspect transcriptionnel de Tbx5. Cette approche a mené à la découverte de 6 transcrits alternatifs exhibant des fonctions à la fois communes et divergentes. La caractérisation de 2 de ces isoformes a révélé le rôle de l’isoforme long (Tbx5_v1) dans la régulation de la croissance des cardiomyocytes durant la cardiogénèse, tandis que l’isoforme court (Tbx5_v2), préférentiellement exprimé dans le cœur mature, réprime la croissance cellulaire. Il est donc entièrement concevable que les mutations de TBX5 entraînant une troncation de la région C-terminale accroissent la concentration d’une protéine mutée qui, à l’instar de Tbx5_v2, interfère avec la croissance de certaines structures cardiaques. En revanche, la divergence de fonctions de ces isoformes, caractérisée par les disparités de localisation subcellulaire et de d’interaction avec d’autres cofacteurs cardiaques, suggère que les mutations affectant davantage un isoforme favoriseraient l’émergence d’un type particulier de MCC. Finalement, un dernier objectif était d’identifier le ou les mécanisme(s) moléculaire(s) par le(s)quel(s) Tbx5 régule son principal gène cible, Nppa, et d’en extraire les indices qui éclairciraient sa fonction transcriptionnelle. Cet objectif nécessitait dans un premier lieu d’identifier les différents modules cis-régulateurs (MCRs) coordonnant la régulation transcriptionnelle de Nppa et Nppb, deux gènes natriurétiques dont l’organisation en tandem et le profil d’expression durant la cardiogénèse sont conservés dans la majorité des vertébrés. L’approche d’empreinte phylogénétique employée pour scanner le locus Nppb/Nppa a permis d’identifier trois MCRs conservés entre diverses espèces de mammifères, dont un (US3) est spécifique aux euthériens. Cette étude a corroboré que la régulation de l’expression du tandem génique Nppb/Nppa requérait l’activité transcriptionnelle d’enhancers en complément aux promoteurs de Nppa et Nppb. La concordance quasiment parfaite entre les profils d’expression de Tbx5 et de ces deux gènes natriurétiques chez les mammifères, suggère que le gradient d’expression ventriculaire de Tbx5 est interprété par le recrutement de ce facteur au niveau des différents enhancers identifiés. En somme, les études présentées dans cette thèse ont permis de clarifier la profusion de fonctions cardiaques que possède Tbx5. Certaines de ces fonctions émanent de l’épissage alternatif de Tbx5, qui favorise la synthèse d’isoformes dotés de propriétés spécifiques. Les diverses interactions combinatoires entre ces isoformes et d’autres facteurs cardiaques au sein des diverses sous-populations de progéniteurs cardiogènes contribuent à l’émergence de RRGs cardiaques divergents. / The vertebrate heart is a modular organ, which requires the complex patterning of the morphogenetic heart fields and the coordinated convergence of the diverse subpopulations of cardiogenic progenitors. At least 7 transcription factors of the T-box family cooperate within these numerous subpopulations of cardiogenic progenitors to regulate the morphogenesis and the layout of multiple structures along the primordial heart tube, which explains that the human mutations of these genes induce various congenital heart defects (CHDs). One of these T-box genes, Tbx5, whose haploinsufficiency generates the Holt-Oram syndrome (HOS), intervenes in a wide variety of gene regulatory networks (GRNs) that orchestrate the morphogenesis of the atria, the left ventricle, the mitral valve, the inter-atrial and inter-ventricular septa, as well as the cardiac conduction system. The diversity of GRNs involved in the formation of these cardiac structures suggests that Tbx5 holds a profusion of functions which will be identifiable only by indexing its molecular activities in each separately examined cardiac lineage. To address this problem, a conditional knockout of Tbx5 in the endocardium was generated. This experiment demonstrated a crucial role of Tbx5 in the survival of the endocardial cells lining the septum primum and the cardiomyocytes within this embryonic structure, which will contribute to the morphogenesis of the inter-atrial septum. Moreover, this study revealed a crosstalk between the Tbx5-positive endocardial cells subpopulation and the myocardium at the level of the septum primum to ensure the survival of cardiomyocytes, and ultimately to guarantee the maturation of the inter-atrial septum. Our results also confirmed the importance of genetic interdependence (Tbx5 and Gata4 as well as Tbx5 and Nos3) between different loci in the morphogenesis of the inter-atrial septum, and particularly the influence that the environment can have on the penetrance and the expressivity of atrial septal defects (ASDs) in the HOS. Besides, since the functions of a gene usually depend on the different isoforms it can generate, a second study focused more on the transcriptional aspect of Tbx5. This approach led to the discovery of 6 alternative transcripts exhibiting both common and specific functions. The characterization of 2 of these isoforms revealed the role of the long isoform (Tbx5_v1) in the regulation of cardiomyocytes growth during cardiogenesis, whereas the short isoform (Tbx5_v2), preferentially expressed in the mature heart, represses cell growth. It is thus entirely conceivable that TBX5 mutations leading to a C-terminal truncation increase the concentration of a mutated protein, which, like Tbx5_v2, interferes with the growth of certain cardiac structures. On the other hand, the divergence of functions of these isoforms, characterized by the disparities of subcellular localization and interaction with other cardiac cofactors, suggests that mutations affecting more one isoform would favor the emergence of a particular type of CHD. Finally, a last objective was to identify one or several molecular mechanism(s) by which Tbx5 regulates its main target gene, Nppa, and to extract clues that might clarify its transcriptional function. This objective required in a first place to identify the various cis-regulatory modules (CRMs) coordinating the transcriptional regulation of Nppa and Nppb, two natriuretic genes whose tandem organization and expression pattern during cardiogenesis are preserved in most vertebrates. The phylogenetic footprint approach employed to scan the Nppb/Nppa locus allowed the identification of three CRMs evolutionary conserved between different mammals species, one of which (US3) is specific to eutherians. This study confirmed that the regulation of the tandem genes Nppb/Nppa required the transcriptional activity of enhancers in complement to Nppa and Nppb promoters. The almost perfect concordance between the expression profiles of Tbx5 and these two natriuretic genes in mammals, suggests that the ventricular expression gradient of Tbx5 is interpreted by the recruitment of this factor to the identified enhancers. Altogether, the studies presented in this thesis allowed clarifying the profusion of Tbx5 cardiac functions. Some of these functions emanate from the alternative splicing of Tbx5, which favors the synthesis of isoforms endowed with specific properties. The diverse combinatorial interactions between these isoforms and other cardiac factors within the various cardiogenic progenitor subpopulations contribute to the emergence of distinct cardiac RRGs.