Sham, Wai Yan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references (leaves 173-205). / Abstracts in English and Chinese. / Declaration of Originality --- p.i / Thesis / Assessment Committee --- p.ii / Abstract --- p.iii / 摘要 --- p.vi / Acknowledgements --- p.viii / Table of Contents --- p.x / List of Figures --- p.xiv / List of Tables --- p.xvii / List of Abbreviations --- p.xix / Chapter Chapter I --- Introduction --- p.1 / Chapter 1.1 --- Habitats and diversity of fish --- p.1 / Chapter 1.2 --- Stress accompanied with osmotic changes --- p.2 / Chapter 1.3 --- Physiological consequences associated with osmoregulation --- p.4 / Chapter 1.3.1 --- Energy cost (energy metabolism) --- p.4 / Chapter 1.3.2 --- Food intake --- p.5 / Chapter 1.3.3 --- Reproduction and development --- p.5 / Chapter 1.3.4 --- Growth --- p.6 / Chapter 1.3.5 --- Immunity --- p.6 / Chapter 1.4 --- Osmoregulatory systems in teleosts --- p.7 / Chapter 1.4.1 --- Ion transport --- p.7 / Chapter 1.4.1.1 --- Mechanism in freshwater --- p.8 / Chapter 1.4.1.2 --- Mechanism in seawater --- p.8 / Chapter 1.4.2 --- Water channel --- p.11 / Chapter 1.4.3 --- Accumulation of compatible osmolytes --- p.11 / Chapter 1.5 --- Kinetics on ion transporters --- p.12 / Chapter 1.5.1 --- Na+K+ATPase --- p.12 / Chapter 1.5.2 --- Other ion transporters --- p.14 / Chapter 1.5.3 --- Other factors related to controlling ion transport --- p.15 / Chapter 1.6 --- Pleiotropic hormones involved in osmoregulation --- p.17 / Chapter 1.6.1 --- Growth hormone/ insulin-like factor-1 axis in seawater adaptation --- p.17 / Chapter 1.6.2 --- Prolactin in freshwater adaptation --- p.19 / Chapter 1.6.3 --- "Dual osmoregulatory role of cortisol, the major corticosteroid in fish" --- p.20 / Chapter 1.7 --- "Growth hormone receptor (GHR), prolactin receptor (PRLR) and signaling pathways" --- p.22 / Chapter 1.8 --- Involvement of sex hormones on osmoregulation --- p.25 / Chapter 1.8.1 --- Estrogens --- p.25 / Chapter 1.8.2 --- Testosterone --- p.26 / Chapter 1.9 --- Biological actions of GH through GHR and PRL through PRLR --- p.27 / Chapter 1.9.1 --- Regulation of GHR --- p.27 / Chapter 1.9.2 --- Regulation of PRLR --- p.28 / Chapter 1.9.3 --- Promoter characterization --- p.30 / Chapter 1.10 --- Adaptation to tonicity-ORE/OREBP pathway --- p.32 / Chapter 1.11 --- Animal model --- p.35 / Chapter 1.12 --- Rationale and objectives of the project --- p.35 / Chapter Chapter II --- Materials and methods --- p.38 / Chapter 2.1 --- Animals --- p.38 / Chapter 2.2 --- In vivo experiments --- p.38 / Chapter 2.3 --- In vitro experiments --- p.38 / Chapter 2.4 --- RNA preparation --- p.39 / Chapter 2.5 --- RNA integrity - formaldehyde agarose gel electrophoresis of mRNA --- p.40 / Chapter 2.6 --- Reverse transcription polymerase chain reaction (RT-PCR) --- p.40 / Chapter 2.7 --- Polymerase chain reaction PCR --- p.41 / Chapter 2.8 --- Quantitative Real-time PCR --- p.41 / Chapter 2.9 --- Cell culture --- p.43 / Chapter 2.10 --- Bioinformatics search --- p.43 / Chapter 2.11 --- Plasmid constructions --- p.43 / Chapter 2.12 --- Transient transfection --- p.47 / Chapter 2.13 --- Luciferase assay --- p.47 / Chapter 2.14 --- Site-directed mutagenesis of sbPRLRl and sbPRLR2 gene promoters --- p.48 / Chapter 2.14.1 --- One-round PCR amplification --- p.48 / Chapter 2.14.2 --- Sequential PCR amplification --- p.48 / Chapter 2.15 --- Nuclear extract preparation for electrophoretic mobility shift assays (EMSA) --- p.52 / Chapter 2.16 --- Electrophoretic mobility shift assays (EMSA) --- p.53 / Chapter 2.16.1 --- Preparation of annealed complementary oligonucleotides --- p.53 / Chapter 2.16.2 --- Preparation of radiolabeled oligonucleotides --- p.54 / Chapter 2.16.3 --- Preparation of non-denaturing polyacrylamide gel --- p.54 / Chapter 2.16.4 --- Electrophoretic mobility shift assay --- p.54 / Chapter 2.17 --- Data analysis --- p.56 / Chapter Chapter III --- Result: In vivo gene expression studies --- p.57 / Chapter 3.1 --- Aim of the chapter --- p.57 / Chapter 3.2 --- Results --- p.57 / Chapter 3.2.1 --- Validation of real-time PCR --- p.57 / Chapter 3.2.2 --- "Transcript levels of sbPRLRl, sbPRLR2, sbNKA α- and sbNKA β-subunit after transfer of fish from seawater to hypersaline water" --- p.61 / Chapter 3.2.2.1 --- Gill --- p.61 / Chapter 3.2.2.2 --- Kidney --- p.64 / Chapter 3.3 --- Discussion --- p.67 / Chapter 3.3.1 --- sbPRLR gene expression after seawater-to-hypersaline water transfer --- p.67 / Chapter 3.3.2 --- sbNKA α- and sbNKA β-subunit expression after seawater-to-hypersaline water transfer --- p.71 / Chapter 3.3.3 --- Overall summary --- p.76 / Chapter Chapter IV --- Result: In vitro gene expression studies --- p.77 / Chapter 4.1 --- Aim of the chapter --- p.77 / Chapter 4.2 --- Results --- p.77 / Chapter 4.2.1 --- Gill organ culture --- p.77 / Chapter 4.2.2 --- Kidney explants --- p.81 / Chapter 4.3 --- Discussion --- p.84 / Chapter Chapter V --- Result: Studies on the gene promoter of sbPRLRs under osmotic change --- p.88 / Chapter 5.1 --- Aim of the chapter / Chapter 5.2 --- Results --- p.88 / Chapter 5.2.1 --- The 5´ة-UTR region of the two sbPRLR genes contain several osmotic response elements --- p.88 / Chapter 5.2.2 --- Promoter activities of the 5´ة-flanking regions of the two sbPRLR genes under osmotic stress --- p.91 / Chapter 5.2.3 --- Identification of a DNA region required for the responses to NaCl --- p.97 / Chapter 5.2.4 --- Interaction of the osmotic response region and transcription factors --- p.104 / Chapter 5.2.4.1 --- Identification of DNA-binding activity in nuclear extract --- p.104 / Chapter 5.2.4.2 --- OSTF1 transactivates the sbPRLR gene promoters --- p.107 / Chapter 5.3 --- Discussion --- p.109 / Chapter Chapter VI --- Result: Studies on the gene promoters of sbPRLR under hormonal challenge --- p.125 / Chapter 6.1 --- Aim of the chapter --- p.125 / Chapter 6.2 --- Results --- p.125 / Chapter 6.2.1 --- The 5´ة-UTR region of the two sbPRLR genes contains several hormone response elements --- p.125 / Chapter 6.2.2 --- Identification of a DNA region required for the responses to steroid hormones --- p.126 / Chapter 6.2.2.1 --- E2-dependent regulation of sbPRLR 1 and sbPRLR2 genes use imperfect EREs --- p.126 / Chapter 6.2.2.2 --- An ARE motif in sbPRLR2 gene promoter --- p.132 / Chapter 6.2.2.3 --- An imperfect and a novel GRE present on the sbPRLRl and sbPRLR2 gene promoters respectively --- p.138 / Chapter 6.2.3 --- Interaction of the hormone response region and transcription factors --- p.144 / Chapter 6.2.3.1 --- The ER interacts with the putative ERE motifs in vitro --- p.144 / Chapter 6.2.3.2 --- The AR interacts with the putative ARE motif in vitro --- p.145 / Chapter 6.2.3.3 --- The GR interacts with the putative GRE motifs in vitro --- p.145 / Chapter 6.3 --- Discussion --- p.152 / Chapter 6.3.1. --- Sex steroids --- p.152 / Chapter 6.3.1.1 --- Estrogens --- p.152 / Chapter 6.3.1.2 --- Androgens --- p.157 / Chapter 6.3.2. --- Interrenal hormone ´ؤ cortisol --- p.163 / Chapter Chapter VII --- General conclusion and perspectives --- p.170 / References --- p.173

Thesis (Ph.D.)--Murdoch University, 2003. / Thesis submitted to the Division of Science and Engineering. Bibliography: leaves 195-212.

Thesis (Ph. D.)--Murdoch University, 1999. / Includes bibliographical references (leaves 116-130).

Thesis (M.Sc.) -- University of Adelaide, Dept. of Zoology, 1972.

Thesis (Ph.D.)--Murdoch University, 2004. / Thesis submitted to the Division of Health Sciences. Bibliography: leaves 63-76.

Thesis (Ph. D.)--Murdoch University, 2003. / Title from PDF title page (viewed Mar. 6, 2005). Includes bibliographical references (p. 195-212).

Kong Pui Shan. / Thesis submitted in: November 2004. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 164-185). / Abstracts in English and Chinese. / Table of Contents --- p.i / List of Figures and Tables --- p.iii / List of Abbreviations --- p.viii / Chapter Chapter 1: --- Introduction --- p.1 / Chapter 1.1 --- Endocrine system and the pituitary gland in teleosts --- p.1 / Chapter 1.2 --- The biology of black seabream (Acanthopagrus schlegeli) --- p.4 / Chapter 1.3 --- Seasonal variation of gene expression in black seabream pituitary --- p.7 / Chapter 1.4 --- Expressed sequence tag (EST) analysis --- p.9 / Chapter 1.5 --- Basic Local Alignment Search Tool (BLAST) --- p.11 / Chapter 1.6 --- Aim of this study --- p.14 / Chapter Chapter 2: --- Materials and Methods --- p.15 / Chapter 2.1 --- Preliminary EST analysis of black seabream pituitary cDNA libraries --- p.15 / Chapter 2.2 --- Larger scale EST analysis of black seabream pituitary cDNA libraries --- p.19 / Chapter 2.3 --- "Northern blot analysis of gene expression levels of growth hormone (GH), prolactin (PRL), somatolactin (SL) and P-actin in adult female black seabream pituitaries" --- p.23 / Chapter 2.4 --- Real time-PCR analysis of seasonal expression of growth hormone family genes in black seabream pituitary --- p.25 / Chapter 2.5 --- Dot blot (miniarray) analysis of seasonal expression of different genes expressed in black seabream pituitary --- p.28 / Chapter Chapter 3: --- Preliminary EST Analysis of Black Seabream Pituitary cDNA Libraries --- p.30 / Chapter 3.1 --- Results and discussion --- p.30 / Chapter 3.1.1 --- EST analysis of black seabream pituitary cDNA libraries --- p.30 / Chapter 3.1.2 --- Full-length sequencing of different hormone genes of black seabream --- p.47 / Chapter Chapter 4: --- Larger Scale EST Analysis of Black Seabream Pituitary cDNA Libraries --- p.64 / Chapter 4.1 --- Results and discussion --- p.64 / Chapter 4.1.1 --- EST analysis of black seabream pituitary cDNA libraries --- p.64 / Chapter 4.1.2 --- Genes first time found in fish --- p.91 / Chapter Chapter 5: --- Analysis of Black Seabream Pituitary Gene Expressions in Pituitary Collected from Different Months --- p.103 / Chapter 5.1 --- Introduction --- p.103 / Chapter 5.2 --- Northern blot analysis of seasonal variation of growth hormone family genes --- p.105 / Chapter 5.3 --- Seasonal analysis of growth hormone family genes by real time-PCR --- p.110 / Chapter 5.4 --- Sexual maturity of black seabream throughout the reproductive cycle --- p.118 / Chapter 5.5 --- Seasonal analysis of black seabream pituitary genes by dot blot study --- p.119 / Chapter 5.6 --- Conclusion --- p.153 / Chapter Chapter 6: --- General Discussion --- p.154 / References --- p.164

Thesis (Ph.D.)--Murdoch University, 2009. / Thesis submitted to the Faculty of Sustainability, Environmental and Life Sciences. Includes bibliographical references.

Fung Chun Kit. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 117-139). / Abstracts in English and Chinese. / Declaration of Originality --- p.i / Acknowledgements --- p.ii / Abstract --- p.iii / 摘要 --- p.v / List of figures --- p.vi / List of tables --- p.viii / List of abbreviations --- p.ix / Chapter Chapter I --- General introduction --- p.1 / Chapter 1.1 --- Different fish habitats with various salinities --- p.1 / Chapter 1.2 --- Osmotic challenges faced by teleosts --- p.2 / Chapter 1.3 --- High ionic strength results in DNA damage and excess water gain --- p.3 / Chapter 1.4 --- Osmoregulatory organs and mechanisms for osmotic balance --- p.4 / Chapter 1.5 --- Different tolerance to salinity changes --- p.8 / Chapter 1.6 --- Effective communication among osmoregulatory organs --- p.9 / Chapter 1.7 --- Introduction to GH and PRL --- p.9 / Chapter 1.8 --- Structure of the GHR and PRLR --- p.10 / Chapter 1.9 --- Hypoosmoregulatory action of GH/IGF-I axis in teleosts --- p.11 / Chapter 1.10 --- Hyperosmoregulatory action of PRL in teleosts --- p.11 / Chapter Chapter II --- Research rationale --- p.13 / Chapter 2.1 --- Physiological importance of osmoregulation in fish --- p.13 / Chapter 2.1.1 --- Energy metabolism --- p.13 / Chapter 2.1.2 --- Growth --- p.14 / Chapter 2.1.3 --- Immunity --- p.14 / Chapter 2.1.4 --- Reproduction --- p.15 / Chapter 2.2 --- Aquaculture importance --- p.15 / Chapter 2.3 --- Unknown molecular regulatory mechanism of hormones during salinity changes in fish --- p.16 / Chapter 2.4 --- Animal model --- p.17 / Chapter Chapter III --- In vivo studies of sbGHR and sbPRLR expression in osmoregulatory organs in response to salinity changes --- p.18 / Chapter 3.1 --- Introduction --- p.18 / Chapter 3.1.1 --- Dynamic change of GH level during salinity changes --- p.18 / Chapter 3.1.2 --- Dynamic change of PRL level during salinity changes --- p.19 / Chapter 3.1.3 --- In vitro studies of GH and PRL release from teleost pituitary in response to extracellular osmolality changes --- p.20 / Chapter 3.1.4 --- Biological actions of GH and PRL through the GHR and PRLR --- p.21 / Chapter 3.2 --- Materials and methods --- p.23 / Chapter 3.3 --- Results --- p.28 / Chapter 3.4 --- Discussion --- p.36 / Chapter 3.4.1 --- Plasma osmolality change during salinity changes --- p.36 / Chapter 3.4.2 --- Gene expression after HSW exposure --- p.38 / Chapter 3.4.3 --- Ionic mediators of the gene expression --- p.43 / Chapter 3.4.4 --- Gene expression after BW exposure --- p.44 / Chapter 3.4.5 --- Dynamic changes of the GHR and PRLR in response to salinity changes --- p.45 / Chapter 3.4.6 --- Regulation of the gene expression in response to salinity changes --- p.46 / Chapter Chapter IV --- Gene expression of sbGHR in liver during salinity changes --- p.49 / Chapter 4.1 --- Introduction --- p.49 / Chapter 4.1.1 --- Responses of the somatotropic axis to salinity changes in fish --- p.49 / Chapter 4.2 --- Materials and methods --- p.52 / Chapter 4.3 --- Results --- p.56 / Chapter 4.4 --- Discussion --- p.60 / Chapter 4.4.1 --- Inhibition of GHR and IGF-I gene expression in liver during HSW exposure --- p.60 / Chapter 4.4.2 --- Downregulation of GHR gene expression by hyperosmotic stress --- p.62 / Chapter 4.4.3 --- Growth retardation of fish during hyperosmotic environment --- p.64 / Chapter Chapter V --- Gene expression studies of sbPRLR in gill organ culture --- p.68 / Chapter 5.1 --- Introduction --- p.68 / Chapter 5.1.1 --- Functions of gill in fish osmoregulation --- p.68 / Chapter 5.1.2 --- Gill culture as a model for osmoregulation studies --- p.69 / Chapter 5.2 --- Materials and methods --- p.70 / Chapter 5.3 --- Results --- p.71 / Chapter 5.4 --- Discussion --- p.73 / Chapter Chapter VI --- Regulation of gene expression of sbGHR in liver during hyperosmotic stress: promoter studies --- p.75 / Chapter 6.1 --- Introduction --- p.75 / Chapter 6.1.1 --- What is a promoter? --- p.75 / Chapter 6.1.2 --- Promoter studies of GHR gene --- p.76 / Chapter 6.2 --- Materials and methods --- p.78 / Chapter 6.3 --- Results --- p.85 / Chapter 6.4 --- Discussion --- p.104 / Chapter Chapter VII --- General discussion and future perspectives --- p.111 / References --- p.117

Finally, the isolation and cloning of black sea bream SL receptor using PCR cloning and protein pull down assay were also attempted. Based on the PCR cloning results, the phylogenetic analysis of nonsalmonids fish GHR1 and SLR protein sequence, the GHR1 data of tissue distribution and effects of environmental salinity and fasting in tilapia, along with the results of far western blot, black sea bream GHR1 is probably a receptor for SL, however there is also a SL specific receptor in black sea bream. / In hormone treated primary cell culture of nonspawning black sea bream pituitary, 10-8 M E2 significantly increases SL mRNA level but 10-10 M, 10-9 M, 10-8 M of E2 inhibit GH mRNA level in female black seabream; 10-8 M E2 also inhibits SL and GH mRNA expression in bisexual black sea bream; 10-8 M MT inhibits SL mRNA expression in male black sea bream but any concentration of MT detected shows no significant effect on GH mRNA level. / Key words. somatolactin (SL), monthly changes, SL promoter, pit-1 and SL receptor / Somatolactin, SL, is a novel member of GH family of pituitary hormone only found in fish. It is considered to be a member of the GH gene family after gene duplication. Two types of SL, SL alpha and SL beta were identified, and SL 13 seems only in fresh water fish, such as goldfish, catfish, rainbow trout, eel and zebrafish. Black sea bream is a marine fish, and there is only SL alpha found from sequencing of over 100 SL cDNA clones. / The cDNAs encoding for transcription factor pit-1 variants were cloned and the transactivation of these Pit-1 isoforms on SL gene promoter were studied. Three variants of Pit-1 are first identified in fish. Pit-1b and Pit-1c can enhance SL promoter activity in Hepa-T1 cells respectively to about 2 fold and 12 fold, but pit-1a failed to activate the SL gene it in the same cells. All the three pit-1s of black sea bream couldn't reverse the inhibition of SL promoter in GH3 cells. The data suggest that N terminal 60 amino acid residues are critical in transactiation on SL promoter and SL promoter activity is possibly limited to fish SL secreting cells. / The SL gene promoter was obtained for gene regulation studies aiming to search for possible regulatory elements controlling the transcription of SL gene in black seabream. SL gene promoter is active in HepaT1 cells, but is inhibited in GH3 cells. Seven putative pit-1 response elements were confirmed with EMSA and super shift assay. / To study the physiological function of SL in black seabream, we initiated a study of monthly expressions of SL mRNA and gonadal somatic index (GSI) to determine whether SL is related to reproduction in black seabream, with GH mRNA levels were also detected for comparison. The results imply that function of SL is possibly related to early development of testis, while GH probably plays some roles in testis and ovary maturation. / by Tian, Jing. / "October 2007." / Source: Dissertation Abstracts International, Volume: 69-08, Section: B, page: 4574. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2007. / Includes bibliographical references (p. 156-170). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.