Earticle

초록

영어

Calreticulin (CRT) is a multifunctional ubiquitous protein that is widely presented in all cells in eukaryotes except erythrocytes. CRT is well known for diverse cellular functions such as endoplasmic reticulum (ER)- specialized protein quality control during protein synthesis and folding, in-vivo Ca2+ homeostasis, antigen presentation, phagocytosis, wound-healing, proliferation, adhesion, and migration of cells. In the current study, we identified CRT from Hippocampus abdominalis (HaCRT) and analyzed expression profiles and functional properties. The cDNA sequence of HaCRT was identified with an open reading frame of 1226 bp. The molecular weight of HaCRT was estimated as 49 kDa. The in-silico study revealed conserved sequence arrangements such as two CRT signature motifs (5′-KHEQSIDCGGGYVKVF-3′ and 5′ -LMFGPDICG-3′ ), triplicate repeats (5′ -IKDPEAKKPEDWD- 3′ , 5′-IPDPDDTKPEDWD-3′ , 5′-IPDPDAKKPDDWD-3′ ), signal peptide and an ER-targeting 5′ - KDEL-3′ sequence of HaCRT. Close sequence similarity of HaCRT was observed with Hippocampus comes from phylogenetic analysis and pairwise sequence comparison. From quantitative polymerase chain reaction (qPCR) results, HaCRT was ubiquitously distributed in all tested tissues and expression levels of HaCRT were significantly modulated in blood, liver and gill tissues after stimulation with Streptococcus iniae, Edwardsiella tarda, polyinosinic:polycytidylic acid, and lipopolysaccharides. Bacterial- and pathogen-associated molecular patternsbinding activities were observed with recombinant HaCRT (rHaCRT). The treatment of murine macrophages with rHaCRT induced the expression of immune genes, such as tumor necrosis factor-α (TNF-α), interleukin 6 (IL- 6), inducible nitric oxide synthase (iNOS), and interleukin-1β (IL-1β). Furthermore, rHaCRT exhibited woundhealing ability. Based on the results from the above study, we suggest that HaCRT play an indispensable role in the immunity of big-belly seahorses by recognition and elimination of pathogens as well as the tissue repairing process.

목차

ABSTRACT
1. Introduction
2. Materials and methods
2.1. Identification and bioinformatics analysis of HaCRT sequences
2.2. Experimental fish rearing and tissue isolation
2.3. Challenge experiment
2.4. RNA isolation and cDNA synthesis
2.5. Transcriptional analysis of HaCRT
2.6. Expression vector construction
2.7. rHaCRT overexpression and purification
2.8. PAMPs- and bacterial-binding activity of rHaCRT
2.9. Cell culture
3. Results
3.1. Identification and sequence analysis of HaCRT
3.2. Tissue-specific mRNA transcription of HaCRT
3.3. Transcription profile of HaCRT upon immune challenge
3.4. rHaCRT purification and SDS-PAGE
3.5. PAMPs- and bacterial-binding ability of rHaCRT by ELISA
3.6. Expression pattern of inflammatory genes in murine macrophages inresponse to rHaCRT stimulation
3.7. Wound-healing activity of rHaCRT
4. Discussion
5. Conclusion
CRediT authorship contribution statement
Acknowledgement
Appendix A. Supplementary data
References

키워드

저자

• Sarithaa Sellaththurai [ Department of Marine Life Sciences & Fish Vaccine Research Center, Jeju National University, Jeju Self-Governing Province, 63243, Republic of Korea, Marine Science Institute, Jeju National University, Jeju Self-Governing Province, 63243, Republic of Korea ]
• W.K.M. Omeka [ Department of Marine Life Sciences & Fish Vaccine Research Center, Jeju National University, Jeju Self-Governing Province, 63243, Republic of Korea, Marine Science Institute, Jeju National University, Jeju Self-Governing Province, 63243, Republic of Korea ]
• Kishanthini Nadarajapillai [ Department of Marine Life Sciences & Fish Vaccine Research Center, Jeju National University, Jeju Self-Governing Province, 63243, Republic of Korea, Marine Science Institute, Jeju National University, Jeju Self-Governing Province, 63243, Republic of Korea ]
• K.A.S.N. Shanaka [ Department of Marine Life Sciences & Fish Vaccine Research Center, Jeju National University, Jeju Self-Governing Province, 63243, Republic of Korea, Marine Science Institute, Jeju National University, Jeju Self-Governing Province, 63243, Republic of Korea ]
• Sumi Jung [ Department of Marine Life Sciences & Fish Vaccine Research Center, Jeju National University, Jeju Self-Governing Province, 63243, Republic of Korea, Marine Science Institute, Jeju National University, Jeju Self-Governing Province, 63243, Republic of Korea ]
• Sukkyoung Lee [ Department of Marine Life Sciences & Fish Vaccine Research Center, Jeju National University, Jeju Self-Governing Province, 63243, Republic of Korea ]
• Jehee Lee [ Department of Marine Life Sciences & Fish Vaccine Research Center, Jeju National University, Jeju Self-Governing Province, 63243, Republic of Korea, Marine Science Institute, Jeju National University, Jeju Self-Governing Province, 63243, Republic of Korea ] Corresponding Author

참고문헌

발행기관

간행물

표지보기 관심저널 등록

• 간행물명

  해양과학연구소 연구논문집 [Bulletin of the Marine Science Institute]

• 간기

  폐간

• pISSN

  1225-5734

• 수록기간

  1977~2021

• 십진분류

  KDC 454 DDC 551