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The effect of short term seawater exposure on the metabolism of Arctic char (Salvelinus alpinus)
s117
P28-2
P29-2
Cloning of an eyestalk neuropeptide gene encoding for the putative gonad-inhibiting hormone in the shrimp, Metapenaeus ensis Gu P-L and Chan S-M Department of Zoology, University of Hong Kong, Hong Kong SAR China
Characterization of the major egg yolk polypeptides in the sand shrimp, Metapenaeus ensb Chan S-M and Wong LS Department of Zoology, University of Hong Kong, Hong Kong SAR, China
We report the cloning and characterization of an additional eyestalk cDNA that shows characteristics of the crustacean CHHMIWGIH tily. It consists of an open reading frame of 879 bp and is encoded by a pre-prohormone of 105 amino acids. The signal peptide and the mature peptide consist of 27 and 78 amino acid residues respectively. It shows the highest amino acid sequence homology to the gonadinhibiting hormone (GIH) of the lobster Hommus Initial study of the genomic organization americanus. suggests that there is a close relationship and a high degree of similarity between thii gene and other neuropeptides of the same shriip. Unlike the MIH and CHH-like gene, the shrimp GIH is also expressed in the CNS besides the eyestalk. GIH transcript is most abundant in the juvenile shrimp and newly spawned mature females. Recombinant protein for the GIH was produced for antibody production Initial study by in vivo injection of the and bioassay. recombinant protein into juvenile shrimp indicates that the GIH has little molt inhibitory effect. Future studies on the gonad inhibitory effect will be performed in the reproductive season of the shriip. Supported by a Hong Kong University institutional
grant.
P29-3
P29-1
The effect of short term seawater exposure on the metabolism of Arctic char (Salvelinus al’inus). Bvstriansky Js, Wright PA and Ballantyne JS Department of Zoology, University of Guelph, Guelph, Qntario, Canada NlG 2Wl. The metabolic effects of a short term (96 hours) transfer of Arctic char (Salvelinus alpims) to seawater (34!%0)was monitored through the measurement of several plasma and tissue parameters. Exposure to seawater induced an initial increase in plasma ion (Na+, Cr) and osmolality levels which peaked at 48 hours, and subsequently declined as osmoregulatory processes were up-regulated. Enzymes of several key metabolic pathways were measured in tissues directly involved in osmoregulation (e.g. gill) and in support tissues (e.g. liver, muscle). Concurrent with the decline in plasma ion levels, there was an increase in the activities of several gill enzymes (e.g. citrate synthase), indicating an increased metabolic cost of osmoregulation in seawater. Seawater exposure also induced increases in oxidative, amino acid and lipid metabolism of the liver and muscle. Alterations in amino acid metabolism of these tissues were correlated with changes in tissue and plasma free amino acid content. Changes in liver and muscle metabolism indicate their role as supportive tissues to the osmoregulatory organs. These findings outline the initial changes in metabolism associated
Although the extra-ovarian and intra-ovarian sources of vitellogenin have been postulated in crustaceans, the site for vitellogenesis in shrimp has not been determined. To study vitellogenesis of the sand shrimp, Idefupenueus ensis, major egg yolk proteins in the ovary of the shrimp was analyzed by SDS-PAGE. Two major polypeptides (70 kDa and 170 kDa) have been identified that are related to the ovarian maturation The amount of these two polypeptides increased cycle. rapidly during the early stage of vitellogenesis and constituted over 80% of the total ovarian protein content at the late vitellogenic stage. Antibodies to the 170 kDa and 70 kDa polypeptides were synthesized and used to characterize vitellogenin of the shrimp. To determine the site for the synthesis of these two polypeptides, total RNAs from the ovary, hepatopancreas and sub-epidermal adipose tissues of vitellogenic females were used to produce a population of mixed proteins by an in vitro translation kit. Western blot analysis using the above antibodies indicated that the 170 kDa polypeptide appeared to arise from the exogenous sources whereas the 70 kDa polypeptide is produced by the ovary. Both antibodies were also used to isolate clones fi-om the expression library generated from these two tissues.
with seawater
adaptation
of Arctic char.
Differential gene expression during larval development of silver sea bream. Deane EE’, Kelly SP’, Larsen SW, Collins PM* and Woo NYS’ ‘Department of Biology, The Chinese University of Hong Kong, Shatin, NT., Hong Kong, and *Department of Ecology, Evolution & Marine Biology, University of California, Santa Barbara, USA. Genes are differentially expressed during the course of larval development. However, the identification and characterization of differentially expressed genes in developing fish has received little attention. Here we report on the differential expression of genes in the developing silver sea bream (5” s&u) larvae. We injected mature female fish with human chorionic gonadotropin and then the eggs were stripped firom female and fertilized. Larvae were allowed to develop in seawater and fed first with a microencapsulated diet (up to Day 7) followed by a mixture of rotifers and artemia Larvae were collected and total whole body RNA was extracted from pools of larvae. In order to identify diffkrentially expressed genes we used the technique of RNA arbitrarily primed polymerase chain reaction (RAP-PCR). Single strand cDNA was synthesized from RNA using reverse transcriptase with an 18 base pair arbitrary primer and then PCR amplified with combinations of arbitrary primers. We identified 15 cDNA clones representing genes that are induced or repressed between Day 1 and Day 21 of development. The size of these cDNA clones ranged from 200 to 800 base pairs and their sequences were analyzed using the BLAST program. Presently we are attempting to isolate more differentially expressed genes during larval development
P28-2
P29-2
Cloning of an eyestalk neuropeptide gene encoding for the putative gonad-inhibiting hormone in the shrimp, Metapenaeus ensis Gu P-L and Chan S-M Department of Zoology, University of Hong Kong, Hong Kong SAR China
Characterization of the major egg yolk polypeptides in the sand shrimp, Metapenaeus ensb Chan S-M and Wong LS Department of Zoology, University of Hong Kong, Hong Kong SAR, China
We report the cloning and characterization of an additional eyestalk cDNA that shows characteristics of the crustacean CHHMIWGIH tily. It consists of an open reading frame of 879 bp and is encoded by a pre-prohormone of 105 amino acids. The signal peptide and the mature peptide consist of 27 and 78 amino acid residues respectively. It shows the highest amino acid sequence homology to the gonadinhibiting hormone (GIH) of the lobster Hommus Initial study of the genomic organization americanus. suggests that there is a close relationship and a high degree of similarity between thii gene and other neuropeptides of the same shriip. Unlike the MIH and CHH-like gene, the shrimp GIH is also expressed in the CNS besides the eyestalk. GIH transcript is most abundant in the juvenile shrimp and newly spawned mature females. Recombinant protein for the GIH was produced for antibody production Initial study by in vivo injection of the and bioassay. recombinant protein into juvenile shrimp indicates that the GIH has little molt inhibitory effect. Future studies on the gonad inhibitory effect will be performed in the reproductive season of the shriip. Supported by a Hong Kong University institutional
grant.
P29-3
P29-1
The effect of short term seawater exposure on the metabolism of Arctic char (Salvelinus al’inus). Bvstriansky Js, Wright PA and Ballantyne JS Department of Zoology, University of Guelph, Guelph, Qntario, Canada NlG 2Wl. The metabolic effects of a short term (96 hours) transfer of Arctic char (Salvelinus alpims) to seawater (34!%0)was monitored through the measurement of several plasma and tissue parameters. Exposure to seawater induced an initial increase in plasma ion (Na+, Cr) and osmolality levels which peaked at 48 hours, and subsequently declined as osmoregulatory processes were up-regulated. Enzymes of several key metabolic pathways were measured in tissues directly involved in osmoregulation (e.g. gill) and in support tissues (e.g. liver, muscle). Concurrent with the decline in plasma ion levels, there was an increase in the activities of several gill enzymes (e.g. citrate synthase), indicating an increased metabolic cost of osmoregulation in seawater. Seawater exposure also induced increases in oxidative, amino acid and lipid metabolism of the liver and muscle. Alterations in amino acid metabolism of these tissues were correlated with changes in tissue and plasma free amino acid content. Changes in liver and muscle metabolism indicate their role as supportive tissues to the osmoregulatory organs. These findings outline the initial changes in metabolism associated
Although the extra-ovarian and intra-ovarian sources of vitellogenin have been postulated in crustaceans, the site for vitellogenesis in shrimp has not been determined. To study vitellogenesis of the sand shrimp, Idefupenueus ensis, major egg yolk proteins in the ovary of the shrimp was analyzed by SDS-PAGE. Two major polypeptides (70 kDa and 170 kDa) have been identified that are related to the ovarian maturation The amount of these two polypeptides increased cycle. rapidly during the early stage of vitellogenesis and constituted over 80% of the total ovarian protein content at the late vitellogenic stage. Antibodies to the 170 kDa and 70 kDa polypeptides were synthesized and used to characterize vitellogenin of the shrimp. To determine the site for the synthesis of these two polypeptides, total RNAs from the ovary, hepatopancreas and sub-epidermal adipose tissues of vitellogenic females were used to produce a population of mixed proteins by an in vitro translation kit. Western blot analysis using the above antibodies indicated that the 170 kDa polypeptide appeared to arise from the exogenous sources whereas the 70 kDa polypeptide is produced by the ovary. Both antibodies were also used to isolate clones fi-om the expression library generated from these two tissues.
with seawater
adaptation
of Arctic char.
Differential gene expression during larval development of silver sea bream. Deane EE’, Kelly SP’, Larsen SW, Collins PM* and Woo NYS’ ‘Department of Biology, The Chinese University of Hong Kong, Shatin, NT., Hong Kong, and *Department of Ecology, Evolution & Marine Biology, University of California, Santa Barbara, USA. Genes are differentially expressed during the course of larval development. However, the identification and characterization of differentially expressed genes in developing fish has received little attention. Here we report on the differential expression of genes in the developing silver sea bream (5” s&u) larvae. We injected mature female fish with human chorionic gonadotropin and then the eggs were stripped firom female and fertilized. Larvae were allowed to develop in seawater and fed first with a microencapsulated diet (up to Day 7) followed by a mixture of rotifers and artemia Larvae were collected and total whole body RNA was extracted from pools of larvae. In order to identify diffkrentially expressed genes we used the technique of RNA arbitrarily primed polymerase chain reaction (RAP-PCR). Single strand cDNA was synthesized from RNA using reverse transcriptase with an 18 base pair arbitrary primer and then PCR amplified with combinations of arbitrary primers. We identified 15 cDNA clones representing genes that are induced or repressed between Day 1 and Day 21 of development. The size of these cDNA clones ranged from 200 to 800 base pairs and their sequences were analyzed using the BLAST program. Presently we are attempting to isolate more differentially expressed genes during larval development