This dissertation, "Study of the in Vivo Role of TSPYL2 in Transgenic Mice" by Kin-wang, Chan, 陳健宏, was obtained from The University of Hong Kong (Pokfulam, Hong Kong) and is being sold pursuant to Creative Commons: Attribution 3.0 Hong Kong License. The content of this dissertation has not been altered in any way. We have altered the formatting in order to facilitate the ease of printing and reading of the dissertation. All rights not granted by the above license are retained by the author. Abstract: Abstract of thesis entitled Study of the in vivo role of TSPYL2 in transgenic mice Submitted by Chan Kin Wang for the degree of Doctor of Philosophy at the University of Hong Kong in January 2007 Ventricular septal defect (VSD) is the most common congenital heart defect at birth. Our group has revealed that TSPYL2 was up-regulated in heart biopsies from patients with VSD by RNA arbitrarily-primed PCR. Analysis on full length cDNA sequence of TSPYL2 revealed a nucleosome assembly protein (NAP) domain. NAP domain- containing proteins have diversified cellular and developmental functions. Studies have shown that TSPYL2 can arrest cell growth in HeLa cells and interact with other proteins in neurons as a transcriptional complex. My research aims to delineate the in vivo functions of TSPYL2. In order to investigate the functions of TSPYL2 in heart development, the transgenic approach was taken. Transgenic mice were generated with promoters reported to be heart-specific: human BNP promoter and rat Mlc-2v promoter. However, four lines of transgenic mice were generated with brain specific expression of TSPYL2 and two transgenic lines were generated with transgene expression in the heart. None of the heterozygous transgenic mice showed any obvious developmental or physiological defect at all stages examined. In severe VSD, due to the higher systolic pressure in the left ventricle, blood flow through the hole to the right ventricle during systole. Due to the inefficient cardiac output from the left ventricle, extra work is required for the defective heart to meet the physiological needs and thus volume-overload hypertrophic stress is resulted. To investigate the functions of TSPYL2 during the hypertrophic stress, mice were induced to cardiac hypertrophy with 5-day daily injections of isoproterenol. In two transgenic lines that expressed TSPYL2 in the heart, the hypertrophic response upon isoproterenol induction was significantly reduced, suggesting a protective role of TSPYL2 expression during the hypertrophic stress. Although the expression of endogenous Tspyl2 was not significantly up-regulated upon this acute induction of hypertrophy, the positive electromobility shift assay results for binding by Mef-2, a hypertrophic response effector, in Tspyl2 promoter and its up-regulated expression during the neonatal stage of rapid cardiomyocte hypertrophic growth suggested the potential regulation of Tspyl2 by hypertrophic stimulation. This led to the hypothesis that TSPYL2 is up-regulated in VSD samples as a protective measure against the hypertrophic stress induced by the haemodynamic inefficiency in severe VSD cases. In one transgenic line, M2, all homozygous transgenic mice died from dehydration at weaning due to inability to drink from the water bottle. They could be rescued by feeding with hydrated food for ten days at weaning age. Their breeding ability was affected by the behavioral problems. The homozygous males did not mate and the homozygous females gave birth to babies with abrasions all over the bodies and died soon after birth. The site of transgene insertion was located by inverse PCR. As confirmed by RT-PCR, the insertion disrupted the expression of leucine-rich repeats containing 7 (Lrrc7). Lrrc7 is normally expressed in the brain and interacts with many neurologically important proteins, such as NR2B, CaMKII, α-actinin, etc. Therefore, ablation of Lrrc7 expression may