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The p53 family of transcription factors, which comprises the products of the TP53, TP63 and TP73 genes, is at the hub of different signalling pathways that determine the fate of a cell. In vitro, the p53 family posses a tumour suppressive function. However, in vivo, although p53-deficient mice develop spontaneous tumours, p73 and p63 KO animals show defects in neuronal and epidermal development, respectively. The ASPP family of proteins also consists of three members: ASPP 1, ASPP2 and iASPP. They are evolutionarily conserved binding partners and specific regulators of p53-, p63- and p73- mediated apoptosis in vitro. In contrast to ASPPl and ASPP2, the biological significance of iASPP in vivo and the possible interaction with p53 family members remains unclear, and it is the subject of this study. To investigate the role of iASPP in vivo, a transgenic mouse model was generated in which iASPP expression is controlled by the Cre/loxP recombination system. Deletion of iASPP resulted in the development of a cardiocutaneous disorder which displayed features of Arrhythmogenic Right Ventricular Cardiomyopathy (ARVC), defects in hair follicle position and impaired epithelial stratification. iASPP loss resulted in a sudden, premature and arrhythmic mode of death of all iASPP mutant mice. iASPP deficiency induced p53-dependent apoptosis in embryonic hearts and dilation of the right ventricle, however, the inactivation of p53 alleles only rescued the fibro-fatty deposits present in iASPP KO hearts. Mechanistically, iASPP locates at the polar ends of cardiomyocytes where it matches the location of other proteins known to be involved in the etiology of ARVC and in maintaining the integrity of intercalated discs. Loss of iASPP also resulted in increased differentiation of primary keratinocytes both in vitro and in vivo. Consistent with this, iASPP bound p63 and inhibited the transcriptional activity of both T Ap63a and T Ap63α and ΔNp63 in vitro, and regulated the expression level ofp63-regulated genes such as envoplakin. These results demonstrate that iASPP prevents cardiocutaneous disorder through its ability to inhibit p53-induced apoptosis and to influence the integrity of intercalated disc. Moreover, iASPP is also an important regulator of p63 and is involved in controlling epithelial stratification in vivo.

Long-distance signaling is an important process in the development of Arabidopsis thaliana. A leucine-rich repeat receptor-like kinase (LRR-RLK), XYLEM INTERMIXED WITH PHLOEM1 a.k.a. C-TERMINALLY ENCODED PEPTIDE RECEPTOR 1 (XIP1/CEPR1), functions in vascular development and has recently been implicated in nitrogen sensing and response. Previous results indicate that XIP1/CEPR1 also interacts with multiple proteins involved in sugar metabolism and transport as well as other metabolic proteins, which indicates a possible role for XIP1/CEPR1 in mediating sugar transport. xip1-1 seeds, which grow slowly in the cold in comparison to Columbia wild-type plants, were previously EMS mutagenized and screened for suppressors of the cold-sensitive phenotype. One of these suppressors, 9-12, maps to the lower region of chromosome V and several possible causative EMS-like mutations have been identified that may link XIP1/CEPR1 to a more general vascular transport role.

by Kin-mang Lau. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1994. / Includes bibliographical references (leaves 148-167). / Chapter I --- ABSTRACT --- p.1 / Chapter II --- INTRODUCTION --- p.3 / Chapter II. 1 --- Epidemiology of prostate cancer --- p.3 / Chapter II.2 --- Anatomy of the human prostate --- p.13 / Chapter II.3 --- Pathology of prostate diseases --- p.14 / Chapter II.3.1 --- Prostatic Hyperplasia / Chapter II.3.2 --- Atypical Hyperplasia / Chapter II.3.3 --- Prostatic Carcinoma / Chapter II.4 --- Tumour Suppressor Gene - Human p53 --- p.19 / Chapter II.4.1 --- General aspects / Chapter II.4.2 --- Human p53 gene - Historical perspectives / Chapter II.4.3 --- Human p53 gene - Structure / Chapter II.4.4 --- Human p53 protein - Structure / Chapter II.4.5 --- Wild-type p53 protein - Biochemical functions / Chapter II.4.6 --- Wild-type p53 protein - Biological function / Chapter II.4.7 --- Regulation of p53 function / Chapter II.4.8 --- p53 mutations in Human cancers / Chapter II.4.9 --- Properties of mutant p53 protein / Chapter II.5 --- Human Papillomavirus (HPV) --- p.38 / Chapter II.5.1. --- Virion Structure / Chapter II.5.2 --- Classification / Chapter II.5.3. --- Papillomaviruses in Human Cancers / Chapter II.5.4. --- Relationship between p53 alteration and HPV infection / Chapter II.6 --- p53 alteration and Prostate cancers --- p.42 / Chapter II.6.1. --- Cytogenetic studies / Chapter II.6.2. --- Hybridization analysis / Chapter II.6.3. --- p53 alterations and Prostatic cell lines / Chapter III. --- OBJECTIVES OF STUDY --- p.46 / Chapter IV --- MATERIALS & METHODS --- p.47 / Chapter IV.1 --- Patients and Materials --- p.47 / Chapter IV.2 --- Histological Grading --- p.47 / Chapter IV.2.1 --- Gleason grading / Chapter IV.2.2 --- W.H.O. grading (Mostofi) / Chapter IV.3 --- Staging of Prostatic carcinoma --- p.48 / Chapter IV.4 --- Collection of Blood and Tissue samples --- p.49 / Chapter IV.5 --- Immunohistochemical studies of Prostatic lesions --- p.50 / Chapter IV.5.1 --- Antibodies used / Chapter IV.5.2 --- Methods in frozen sections / Chapter IV.5.3 --- Methods in paraffin sections / Chapter IV.5.4 --- Controls / Chapter IV.5.5 --- Immunohistochemical evaluation / Chapter IV.6 --- Extraction of DNA from tissues and blood samples --- p.52 / Chapter IV.6.1 --- Extraction of genomic DNA from blood / Chapter IV.6.2 --- Extraction of genomic DNA from tissue / Chapter IV.7 --- Hybridization analysis --- p.54 / Chapter IV.7.1 --- Preparation of Cloned Probe DNA / Chapter IV.7.2 --- Transformation of CaCl2-treated competent cell / Chapter IV.7.3 --- Cultures of Transformants / Chapter IV.7.4 --- Isolationof plasmid DNA from transformant cultures / Chapter IV.7.5 --- Purification of DNA Probe by electroelution / Chapter IV.7.6 --- Radioactive labelling of DNA Probes / Chapter IV.7.7 --- Purification of radioactive labelled DNA Probes / Chapter IV.7.8 --- Southern Blotting Technique / Chapter IV.7.9 --- Hybridization of DNA Blots with labelled DNA Probe / Chapter IV.8 --- Polymerase Chain Reaction - Single Stranded Conformational Polymorphism (PCR-SSCP) --- p.63 / Chapter IV.8.1 --- 5'-radioactive labelling of primer / Chapter IV.8.2 --- Amplification of target sequence by PCR / Chapter IV.8.3 --- Nondenaturing Polyacrylamide Gel Electrophoresis / Chapter IV.8.4 --- Direct DNA sequencing of PCR products with p53 mutation / Chapter IV.8.5 --- Controls / Chapter IV.9 --- PCR method for detection of Human Papillomavirus (HPV) --- p.71 / Chapter IV.9.1 --- PCR-Amplification / Chapter IV.9.2 --- DNA alkali Blotting Technique / Chapter IV.9.3 --- Preparation of Radioactive labelled Oligoprobes / Chapter IV.9.4 --- Hybridization of DNA Blots with radioactive labelled Oligoprobes / Chapter IV.9.5 --- Controls / Chapter IV.9.6 --- Sensitivity of HPV 18 detection by PCR / Chapter V --- RESULTS --- p.76 / Chapter V.1 --- Grading and Staging of patients with prostatic carcinoma --- p.76 / Chapter V.2 --- Immunohistochemistry in prostatic lesions --- p.80 / Chapter V.3 --- Results of hybridization analysis --- p.81 / Chapter V.4 --- PCR-SSCP findings in prostatic hyperplasia and carcinoma --- p.97 / Chapter V.5 --- PCR detection of HPV in human prostate --- p.110 / Chapter VI --- DISCUSSION --- p.125 / Chapter VII --- CONCLUSION --- p.146 / Chapter VIII --- REFERENCES --- p.148 / Chapter IX --- APPENDIX --- p.168 / Chapter X --- ACKNOWLEDGEMENT --- p.172

Behçet’s disease (BD) is a chronic, multi systemic, recurrent vasculitis disease of unknown aetiology. The clinical manifestations are composed of relapsing episodes of recurrent oral ulcers, uveitis, skin lesions and genital ulcers along with musculoskeletal and neurological involvement. Pro-inflammatory cytokines are a key feature of the disease but the triggers for their induction are not well understood and/or controversial. Many cytokines (including IFNγ, IL-12, IL-23, IL-10 and IL-6) activate the JAK-STAT signalling pathway which is negatively regulated by Suppressor of Cytokine Signalling (SOCS) proteins. Therefore, it was hypothesised that SOCS proteins may be dysregulated in BD. The expression of SOCS 1-3 mRNA and protein was studied in peripheral blood mononuclear cells (PBMCs), Neutrophils and buccal mucosal cells (BMC) of BD patients and compared with healthy controls (HC) and recurrent aphthous stomatitis (RAS) patients. SOCS 1 and 3 were significantly upregulated in PBMCs of BD patients compared with HC (p=0.0149; p=0.0007) and there were subtle differences between expression in relapsed and symptom free BD (quiet BD). SOCS1 and SOCS 3 also significantly upregulated in BMC from oral ulcers of BD compared with HC (both at p=0.0001). Cytokines were examined in serum, saliva and culture supernatants from stimulated PBMCs. IL-6 were significantly upregulated in the saliva of relapsed BD patients compared with HC (p=0.0104) and the capacity for IL-10 secretion from BD was compromised. Phosphorylation of STATs, transcription factors RORγt, T-bet and 48 protein kinases were investigated using a novel PhosphFlow method and by microarray analysis. STATs were upregulated in BD and seven novel kinase proteins showed differential phosphorylation in BD. Conclusion: SOCS 1-3 expression has changed in BD patients with differences in PBMC and Neutrophil expression between the SOCS proteins. Phosphorylation of STATs and several kinases show up-regulation in BD and seven kinases with altered phosphorylation states in BD were identified as novel targets for future investigation.