The B cell population is one of the key components of the adaptive immune system, which protects the host from a tremendous variety of pathogens by producing antibodies. B cells develop from hematopoietic stem cells through a pathway known as B lymphopoiesis. This is a process accompanied by intensive gene expression reprogramming. By the end, genes appropriate for the B lineage are activated and those that are not are continuously repressed. The regulation of lineage gene expression is conferred by a network of transcriptional regulators. Although some key components have been defined, more factors, especially those orchestrating the repression of non-B lineage genes, remain to be identified. Chemically induced mutagenesis is a potent way of identifying genes with critical biological functions. Injection of n-ethyl-n-nitrosourea, a mutagen, has generated a unique point mutation in the mouse Gon4-like (Gon4l) gene that specifically causes a loss of peripheral B cells while maintaining the T cell population. The mutation is therefore named Justy for Just T cells. The goal of this thesis project is to analyze the Justy mice and provide insights into the mechanisms underlying the regulation of B lymphopoiesis. The work presented here demonstrates that the protein encoded by Gon4l is essential for early B lymphopoiesis, which is likely through the repression of non-B lineage genes. Gon4l protein contains conserved domains implicated in transcriptional repression and associates in a complex with the transcriptional repression mediators Yin Yang 1 and Sin3a/HDAC1, after these proteins are transiently expressed in cell lines. When bound to DNA, Gon4l is capable of repressing a nearby promoter and this function correlates with its ability to form a complex. Therefore, these results suggest that Gon4l may function as a transcriptional regulator by employing its associated co-factors in the identified complex. Lastly, a wide spectrum of tumors developed in Justy mice, indicating that Gon4l can also act as a tumor suppressor.

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Diffuse large B cell lymphoma (DLBCL) is the most common non-Hodgkin’s lymphoma. Patients who fail conventional therapy (~50%) have a poor prognosis and few treatment options. It is essential to understand the underlying biological processes, the progression of the disease, and utilize this information to develop new therapeutics. DLBCL patients with high C-MYC expression have a poor prognosis and new therapeutics for these patients are needed. This thesis describes work testing the hypothesis that JQ1, which can indirectly inhibit C-MYC in some tumors, can be used as an effective treatment for DLBCL. Some tumors have an unknown mechanism causing high C-MYC expression, leading me to investigate the underlying mechanisms. YY1 is a transcriptional regulator of c- Myc and has been implicated in DLBCL and as a potential regulator of the germinal center (GC) reaction. DLBCL arises from GC cells or post-GC cells. I tested the hypothesis that YY1 regulates the GC reaction. SMURF2 is an E3-ubiquitin ligase for YY1 and a tumor suppressor for DLBCL. I was interested in examining the mechanism underlying the suppression of DLBCL by SMURF2 leading to the hypothesis that SMURF2 regulates the GC. This thesis shows JQ1 leads to cell death and cellular senescence in human DLBCL cells. I conclude that BRD4 inhibition by JQ1 or derivatives could provide a new therapeutic avenue for DLBCL patients. I also show loss of YY1 perturbs the GC by decreasing the dark zone and increasing apoptosis. Finally I show modulation of SMURF2 does not affect the GC, suggesting SMURF2 utilizes a different mechanism to act as a tumor suppressor and may not modulate YY1 in the context of the GC.

La NADH:ubiquinone oxydoréductase (complexe I) est le plus gros complexe enzymatique du système mitochondrial d'oxydation phosphorylante (43 sous-unités chez l'homme). Très peu de données sont disponibles concernant les mécanismes régulant l'expression de ces protéines. <br />Cette étude a été initiée par l'étude des promoteurs de deux gènes du complexe I mitochondrial humain. Les résultats montrent que le gène NDUFS8 qui code pour la sous-unité 23 kDa (TYKY) est transcrit sous le contrôle des facteurs de transcription YY1 et Sp1 tandis que gène NDUFS7 codant pour la sous-unité 20 kDa (PSST) est régulé par NRF-1 et Sp1. <br />Dans la deuxième partie de ce travail, une méthode d'analyse du protéome mitochondrial humain par électrophorèse bidimensionnelle a été développée. Le but est d'aborder de manière globale et sans a priori l'expression des protéines du complexe I : déterminer qui est régulé et comment en réponse à un stimulus déterminé? <br />Des cartes de référence ont été développées à partir de mitochondries extraites de placenta humain en utilisant deux types de gradient de pH : l'un est adapté aux protéines acides et neutres, l'autre aux protéines basiques. Sur ces cartes, 85 protéines mitochondriales ont été identifiées par spectrométrie de masse dont 17 sous-unités du complexe I. Cette technique d'analyse protéomique a ensuite été utilisée pour étudier la régulation de l'expression des protéines mitochondriales par le fer. Sur le plan technique, les premiers résultats sont encourageants : les gels d'électrophorèse bidimensionnelle préparés avec des mitochondries extraites de cellules en culture sont de bonne qualité et des variations reproductibles de l'expression de sous-unités du complexe I et d'autres protéines mitochondriales ont pu être détectées. Sur le plan fondamental, les données obtenues sont préliminaires. Il sera nécessaire de réaliser de nouvelles expériences pour confirmer les premières observations et analyser la cinétique des variations détectées.