東京海洋大学博士学位論文 平成27年度(2015) 応用生命科学 課程博士甲第369号指導教員: 近藤　秀裕全文公表年月日: 2018-07-09東京海洋大学201

Kittipong Thanasaksiri

東京海洋大学博士(海洋科学)東京海洋大学博士学位論文 平成27年度(2015) 応用生命科学 課程博士甲第369号指導教員: 近藤　秀裕全文公表年月日: 2018-07-09doctoral thesi

Kittipong, Thanasaksiri

キティポン, タナサクシリ

Open Access Collection of International and Scholarly Papers

Studies on the effects of temperature on thegene expression profiles of immune-relatedgenes in Japanese flounder Paralichthysolivaceus学位名 博士(海洋科学)学位授与機関 東京海洋大学学位授与年度 2015学位授与番号 12614博甲第369号URL http://id.nii.ac.jp/1342/00001213/[課程博士・論文博士共通]  博士学位論文内容要約 Summary  Viral pathogens have been causing economic losses in mariculture worldwide. Many researchers have emphasized on study the fish immune system to prevent the viral diseases. Temperature is one of the most important influences on the immune system of fish. Several studies have been shown that temperature changes modulated immune-related gene expression. However, gene expression changes in response to virus and temperature changes in teleosts have not been comprehensively analyzed. Hence, the objective of this research is to study the effect of temperature on gene expression profiling. Type I interferon (type I IFN) system is an important innate immune response to induce antiviral state. There is evidence that temperatures influenced on the expression profiles of Mx gene, a marker of type I IFN expression. Thus, in this first study, gene expression profiling of poly (I:C)-treated Japanese flounder, Paralichthys olivaceus, under different temperatures was investigated using microarray analysis. The response was analyzed in spleen tissue at 3 and 24 h post injection (hpi) at 15 °C and 25 °C. A large number of genes in fish treated with poly (I:C) at 25 °C were expressed at 3 hpi, whereas the expression profiles at 24 hpi appeared to be similar to those of the controls. Cluster analysis of the different expression profiles showed three distinct groups of up-regulated genes in fish reared at 15 °C. These were early (3 hpi), early-to-late (3 and 24 hpi), and late (24 hpi) up-regulated genes. These genes included type I IFN-related genes and inflammatory genes. Among the up-regulated genes, most of the type I IFN-related genes played early-to-late- and late-responding genes at 15 °C but early-responding genes at 25 °C. Thus, several up-regulated genes in these groups from the microarray result were further verified by qPCR. These results indicate that the type I IFN-related gene expressions of P. olivaceus treated with poly (I:C) can be influenced by temperature. Suppressor of cytokine signaling (SOCS) family members play crucial roles in regulating cytokine action and have now been identified in several fish species. Of several up-regulated genes from microarray result, SOCS1 gene showed different expression profiles between two temperatures and expression of SOCS1 gene seemed to be related with the expression patterns of interferon regulatory genes. In the second study, SOCS family members of Japanese flounder were identified and analyzed. Using the EST sequences obtained from next generation sequencing of Japanese flounder, candidate ESTs were identified. The coding sequences of the predicted SOCS members were obtained using 3′- and/or 5′-RACE PCR. Phylogenetic analysis revealed that there are 8 members of SOCS family in Japanese flounder: CISH, SOCS1a, SOCS1b, SOCS3a, SOCS3b, SOCS5, SOCS6, SOCS9. SOCS1a had 32.3 % identity to SOCS1b, whereas SOCS3a had 45.2% identity to SOCS3b. The expression of all SOCS genes was detectable in all the tissues examined. Stimulation of Japanese flounder reared at 15 or 25 °C with poly (I:C) had effect on CISH, SOCS1 and SOCS3 genes expression. All SOCS genes were significant expressed after FKC of E. tarda injection. These results indicated that all SOCS genes might be involved in immune responses and CISH, SOCS1 and SOCS3 genes may have distinct functions to the other SOCS members.  Nucleotide-binding and oligomerization domain (NOD)-like receptor (NLR) family represents a group of intracellular sensors of the innate immune system. NLR family CARD domain containing 5 (NLRC5) is a 専 攻 Major Applied Marine Bioscience 氏 名 Name Kittipong Thanasaksiri 論文題目 Title Studies on the effects of temperature on the gene expression profiles of immune-related genes in Japanese flounder Paralichthys olivaceus （ヒラメ Paralichthys olivaceus における免疫関連遺伝子群の発現動態に及ぼす水温の影響に関する解析） member of the highly conserved of NLR family. Previous study in mice showed that NLRC5 negatively regulates the NF-ᴋB and type I interferon signaling pathways. Furthermore, based on our microarray result, the NLRC5 was differentially up-regulated between two temperatures. In the third study, Japanese flounder NLRC5 gene was identified and analyzed. The predicted open reading frame (ORF) of Japanese flounder NLRC5 encoded a protein of 1,842 aa with twelve leucine-rich repeat domains (LRRs) at the C-terminus. NLRC5 expression was observed in all tissues studied. Upon poly (I:C) treatment, expression of NLRC5 gene was detected at 24 hpi at 15 °C and at 3 hpi at 25 °C. Significant up-regulation of NLRC5 gene was observed at 12 hpi after formalin-killed cells of Edwardsiella tarda but not Streptococcus iniae treatments. These results indicated that NLRC5 gene involves in fish immune responses. These results can be concluded that temperature modified and affected the time course changes of gene expression patterns. Furthermore, at high temperature (25 °C), response occurred earlier than low temperature (15 °C). Negative regulators of type I IFN signaling (SOCS1 and NLRC5) might be involved in the regulation of gene expressions modulated by temperatures.   

Studies on the effects of temperature on the gene expression profiles of immune-related genes in Japanese flounder Paralichthys olivaceus

https://oacis.repo.nii.ac.jp/?action=repository_action_common_download&item_id=1223&item_no=1&attribute_id=20&file_no=3

ヒラメ Paralichthys olivaceus における免疫関連遺伝子群の発現動態に及ぼす水温の影響に関する解析

Abstract

Similar works

Full text

Available Versions

Open Access Collection of International and Scholarly Papers