There is an increasing trend towards advancing the understanding and development of ethylene oligomerization catalysts, both in academia and industry. The metal of choice in this chemistry is invariably chromium, which has shown great versatility in selective trimerization/tetramerization, non-selective oligomerization and polymerization of ethylene. While much success has been achieved in ethylene trimerization, the same con not be said about tetramerization catalysis. Aminophosphine based ligands have demonstrated their ability towards selective 1-octene production, however, the popular PNP catalyst is able to achieve only 70% selectivity. In order to explore the possibility of developing and enhancing the selectivity of chromium based ethylene tetramerization catalyst, this thesis work was undertaken. The ligand systems we chose for our work were bidentate aminophosphine based (PN(CH2)nNP), which has yielded interesting selective oligomerization. Subtle modifications were found to result in drastic changes in selectivity, from tetramerization (PN(CH2)3NP) to trimerization (PN(CH2)2NP). We managed to successfully develop the first truly selective (over 90%) 1-octene catalyst with polymer-free behavior. Further modifications on the ligand framework, where one atom of Si was used to link the two NP units, resulted in non-selective oligomerization, in which case we determined that the oxidation-state of chromium is a key player. We explored other modifications on our selective ligands in which one of the arms on the bidentate ligand was replaced with a base-donor amine, phosphine or pyridine, and resulted in interesting selectivity changes. The final modification that we tested was a novel N(CH2)2P ligand and found it to be a highly active, non-selective oligomerization catalyst.

There is an increasing trend towards advancing the understanding and development of ethylene oligomerization catalysts, both in academia and industry. The metal of choice in this chemistry is invariably chromium, which has shown great versatility in selective trimerization/tetramerization, non-selective oligomerization and polymerization of ethylene. While much success has been achieved in ethylene trimerization, the same con not be said about tetramerization catalysis. Aminophosphine based ligands have demonstrated their ability towards selective 1-octene production, however, the popular PNP catalyst is able to achieve only 70% selectivity. In order to explore the possibility of developing and enhancing the selectivity of chromium based ethylene tetramerization catalyst, this thesis work was undertaken. The ligand systems we chose for our work were bidentate aminophosphine based (PN(CH2)nNP), which has yielded interesting selective oligomerization. Subtle modifications were found to result in drastic changes in selectivity, from tetramerization (PN(CH2)3NP) to trimerization (PN(CH2)2NP). We managed to successfully develop the first truly selective (over 90%) 1-octene catalyst with polymer-free behavior. Further modifications on the ligand framework, where one atom of Si was used to link the two NP units, resulted in non-selective oligomerization, in which case we determined that the oxidation-state of chromium is a key player. We explored other modifications on our selective ligands in which one of the arms on the bidentate ligand was replaced with a base-donor amine, phosphine or pyridine, and resulted in interesting selectivity changes. The final modification that we tested was a novel N(CH2)2P ligand and found it to be a highly active, non-selective oligomerization catalyst.

There is an increasing trend towards advancing the understanding and development of ethylene oligomerization catalysts, both in academia and industry. The metal of choice in this chemistry is invariably chromium, which has shown great versatility in selective trimerization/tetramerization, non-selective oligomerization and polymerization of ethylene. While much success has been achieved in ethylene trimerization, the same con not be said about tetramerization catalysis. Aminophosphine based ligands have demonstrated their ability towards selective 1-octene production, however, the popular PNP catalyst is able to achieve only 70% selectivity. In order to explore the possibility of developing and enhancing the selectivity of chromium based ethylene tetramerization catalyst, this thesis work was undertaken. The ligand systems we chose for our work were bidentate aminophosphine based (PN(CH2)nNP), which has yielded interesting selective oligomerization. Subtle modifications were found to result in drastic changes in selectivity, from tetramerization (PN(CH2)3NP) to trimerization (PN(CH2)2NP). We managed to successfully develop the first truly selective (over 90%) 1-octene catalyst with polymer-free behavior. Further modifications on the ligand framework, where one atom of Si was used to link the two NP units, resulted in non-selective oligomerization, in which case we determined that the oxidation-state of chromium is a key player. We explored other modifications on our selective ligands in which one of the arms on the bidentate ligand was replaced with a base-donor amine, phosphine or pyridine, and resulted in interesting selectivity changes. The final modification that we tested was a novel N(CH2)2P ligand and found it to be a highly active, non-selective oligomerization catalyst.

La chaîne invariante (Ii) agit à titre de chaperon dans l’assemblage et le trafic des molécules du complexe majeur d’histocompatibilité de classe II (CMHII). Chez l’humain, les deux isoformes prédominantes, p33 et p35, diffèrent par la présence d’un motif di-arginine (RXR). Ce dernier permet la rétention de p35 au réticulum endoplasmique (RE) jusqu’à son masquage par une molécule de CMHII. La chaîne invariante forme des trimères auxquels s’associent successivement jusqu’à trois dimères αß de CMHII résultant en la formation de pentamères, heptamères et nonamères. Toutefois, la stœchiométrie exacte des complexes Ii-CMHII qui quittent le RE et le mécanisme permettant le masquage du motif RXR demeurent un sujet de débats. Dans un premier temps, nous avons examiné par une approche fonctionnelle la stœchiométrie des complexes formés autour de p33 et de p35. Nous avons observé que p35 engendre la formation de complexes nonamériques (αßIi)3 et permet l’incorporation de différents isotypes de CMHII autour d’un même trimère de p35 alors que p33 facilite la formation de pentamères (αß)1Ii3. Lors de l’étude du masquage du motif RxR par les CMHII, nous avons montré que son inactivation requiert une interaction directe (en cis) entre les sous-unités p35 et CMHII, résultant en une rétention des trimères de p35 insaturés au RE. Aussi, nous avons observé que contrairement aux complexes p33-CMHII, les complexes p35-CMHII sont retenus au RE lorsque coexprimés avec la protéine NleA de la bactérie Escherichia coli entérohémorragique. Comme l’expression de NleA interfère avec la formation des vésicules COPII responsable de l’export du RE, nous supposons que la sortie du RE des complexes p35-CMHII dépend des vésicules COPII alors que la sortie des complexes formés autour de l’isoforme p33 est indépendante de la formation de ces vésicules. La trimérisation d’Ii représente la toute première étape dans la formation des complexes Ii-CMHII. Deux domaines d’Ii permettent la formation de trimères; le domaine de trimérisation (TRIM) et le domaine transmembranaire (TM). Nous nous sommes intéressés à la nécessité de ces domaines dans la trimérisation de la chaîne et la formation subséquente de complexes avec les CMHII. Nous avons démontré que le domaine TRIM n’est pas essentiel à la trimérisation de la chaîne, à la formation de pentamères et de nonamères ainsi qu’au trafic adéquat de ces complexes Ii-CMHII dans la cellule. En absence des domaines TM d’Ii et des CMHII, nous avons observé la formation de complexes pseudo-nonamériques. Ceci suppose que la présence de ce domaine n’est pas un prérequis à la formation de nonamères. En conséquence, la présence d’un seul domaine de trimérisation de Ii est requise pour la formation de trimères et de complexes nonamériques. L’ensemble de nos résultats démontrent que la fonction de p35 n’est pas redondante à celle de p33. p35 influence de manière distincte le trafic des CMHII puisqu’il affecte la stœchiométrie des sous-unités incorporées aux complexes Ii- CMHII. / The invariant chain (Ii) assists in the folding and trafficking of MHC class II molecules (MHCII). Four different isoforms of the human Ii have been described (p33, p35, p41 and p43). The main isoforms, p33 and p35, differ by the presence of a di-arginine (RXR) endoplasmic reticulum (ER) retention motif in p35. This motif is inactivated upon binding of MHCII. In the ER, p33 and p35 assemble into trimers before associating with MHCII. The sequential binding of up to three MHCII αß dimers to Ii trimers results in the formation of pentamers, heptamers and nonamers. However, the exact stoichiometry of the Ii-MHCII complex and the mechanism allowing shielding of the ER retention motif remain a matter of debate. To shed light on these issues, we chose a functional approach to examine the stoichiometry of complexes formed around the p33 and p35 isoforms. We showed that p35 promotes formation of nonameric complexes (αßIi)3 while formation of pentameric complexes (αß)1Ii3 was observed for p33. We then showed that formation of nonameric complexes can result in the inclusion of distinct MHCII isotypes around a single trimeric p35 scaffold. When answering the question wetter masking of the p35 RXR motif by MHCII results in the formation of nonamers, we showed that the actual inactivation of motif requires a direct cis-interaction between p35 and the MHCII, precluding ER egress of unsaturated p35 trimers. Interestingly, as opposed to p33-MHCII complexes, p35-MHCII complexes remained in the ER when co-expressed with the NleA protein of enterohaemorrhagic Escherichia coli. Expression of this bacterial protein is thought to interfere with the formation of COPII vesicles, leading to the conjecture that p35-MHCII and p33-MHCII complexes exit the ER in a COPII-dependant and COPII-independent manner, respectively. The trimerization of Ii represents the very first step in the formation of Ii-MHCII complex. Two domains of Ii, the trimerization domain (TRIM) and the transmembrane (TM) domain have been shown to trigger the trimerization of the chain. We focused our attention on the requirement of the two trimerization domains in Ii self-association and in the formation of pentameric and nonameric complexes. We showed that the TRIM domain of Ii is not essential for the chain’s trimerization, formation of pentamers and nonamers and for proper traffic with MHCII molecules. In absence of the Ii and MHCII TM domains, we observed the formation of a nonamer-like structure hereby suggesting that the presence of this domain is not a prerequisite for nomamer complex formation. Consequently, our results showed that either Ii trimerization domains are sufficient for Ii trimer formation and nonameric complex trafficking. Taken together, our results demonstrate that the function of the p35 isoform is not redundant, influencing distinctively MHCII trafficking as the subunit stoichiometry of oligomeric Ii/MHCII complexes is affected by p35.

Indiana University-Purdue University Indianapolis (IUPUI) / Phosphatases of regenerating liver, consisting of PRL-1, PRL-2 and PRL-3, belong to a novel protein tyrosine phosphatases subfamily, whose overexpression promotes cell proliferation, migration and invasion and contributes to tumorigenesis and metastasis. However, although great efforts have been made to uncover the biological function of PRLs, limited knowledge is available on the underlying mechanism of PRLs’ actions, therapeutic value by targeting PRLs, as well as the physiological function of PRLs in vivo. To answer these questions, we first screened a phage display library and identified p115 RhoGAP as a novel PRL-1 binding partner. Mechanistically, we demonstrated that PRL-1 activates RhoA and ERK1/2 by decreasing the association between active RhoA with GAP domain of p115 RhoGAP, and displacing MEKK1 from the SH3 domain of p115 RhoGAP, respectively, leading to enhanced cell proliferation and migration. Secondly, structure-based virtual screening was employed to discover small molecule inhibitors blocking PRL-1 trimer formation which has been suggested to play an important role for PRL-1 mediated oncogenesis. We identified Cmpd-43 as a novel PRL-1 trimer disruptor. Structural study demonstrated the binding mode of PRL-1 with the trimer disruptor. Most importantly, cellular data revealed that Cmpd-43 inhibited PRL-1 induced cell proliferation and migration in breast cancer cell line MDA-MB-231 and lung cancer cell line H1299. Finally, in order to investigate the physiological function of PRLs, we generated mouse knockout models for Prl-1, Prl-2 and Prl-3. Although mice deficient for Prl-1 and Prl-3 were normally developed, Prl-2-null mice displayed growth retardation, impaired male reproductive ability and insufficient hematopoiesis. To further investigate the in vivo function of Prl-1, we generated Prl-1-/-/Prl-2+/- and Prl-1+/-/Prl-2-/- mice. Similar to Prl-2 deficient male mice, Prl-1-/-/Prl-2+/- males also have impaired spermatogenesis and reproductivity. More strikingly, Prl-1+/-/Prl-2-/- mice are completely infertile, suggesting that, in addition to PRL-2, PRL-1 also plays an important role in maintaining normal testis function. In summary, these studies demonstrated for the first time that PRL-1 activates ERK1/2 and RhoA through the novel interaction with p115 RhoGAP, targeting PRL-1 trimer interface is a novel anti-cancer therapeutic treatment and both PRL-1 and PRL-2 contribute to spermatogenesis and male mice reproductivity.