Characterization of Mud Loach (Misgurnus mizolepis) Apolipoprotein A-I: cDNA Cloning, Molecular Phylogeny and Expression Analysis

미꾸라지(Misgurnus mizolepis) Apolipoprotein A-I cDNA의 구조, 분자계통 및 발현 특징 분석

  • Lee, Youn-Ho (National Fisheries Research and Development Institute) ;
  • Noh, Jae-Koo (National Fisheries Research and Development Institute) ;
  • Kim, Keun-Yong (Department of Aquaculture, Pukyong National University) ;
  • Cho, Young-Sun (Department of Aquaculture, Pukyong National University) ;
  • Nam, Yoon-Kwon (Department of Aquaculture, Pukyong National University) ;
  • Kim, Dong-Soo (Department of Aquaculture, Pukyong National University)
  • Published : 2007.02.25


Full length complementary DNA encoding apolipoprotein A-I (apoA-I) was isolated and characterized in mud loach (Misgurnus mizolepis). Mud loach apoA-I cDNA encoding 24 bp of 5'-untranslated region (UTR), 762 bp of single open reading frame (ORF) consists of 254 amino acids and 293 bp of 3'-UTR excluding stop codon and poly (A+) tail. Two overlapping polyadenylation signals (AATAAAATAAA) was found 9 bp prior to the poly (A+) tail. Mud loach apoA-I represented considerable homology to those from other teleost species at amino acid level with conserving common features of vertebrate apoA-I. Molecular phylogenetic analysis inferred the phylogenetic hypothesis that was generally in accordance with the previous taxonomic relationship. Apolipoprotein A-I mRNA was detected in various tissues, but the mRNA levels were quite varied depending on tissues based on semi-quantitative RT-PCR. Liver and brain showed the significantly higher levels of apoA-I transcripts than other tissues. mRNA expression of apoA-I was quite low in very early stage of embryonic development, however dramatically enhanced from 8 hours post fertilization. This increased mRNA level was retained consistently up to 14 days post hatching.

우리나라 주요 담수어종인 미꾸라지(Misgurnus mizolepis)로 부터 apolipoprotein A-I (apoA-I) cDNA를 분리하고 그 구조, 분자 계통 및 발현 특징을 분석하였다. 미꾸라지 apoA-I cDNA는 254개의 아미노산을 암호화하고 있는 762 bp의 ORF를 포함하고 있었으며 아울러 24 bp의 5'UTR 및 293 bp의 3'UTR(종결 코돈 및 poly A tail 제외)를 갖고 있었다. 미꾸라지 apoA-I은 여타 척추동물 apoA-I과의 다중배열 시 염기서열에서는 많은 차이를 나타내었지만 단백질의 구조적 특징은 높은 상동성을 보였고, 또한 척추동물의 apoA-I들과의 분자계통을 분석한 결과, 종래 알려진 분류학적 위치와 비교적 잘 일치하였다. 미꾸라지 apoA-I mRNA는 RT-PCR 분석을 통해 간 및 뇌 조직에서 분석한 다른 조직보다 유의적으로 높게 발현하는 것으로 나타났고, 특히 간에서 가장 높은 발현을 보였다. 수정시부터 부화 후 14일까지 초기 발생 및 치어에서의 apoA-I mRNA 발현을 조사한 결과 수정 8시간째부터 급격한 발현의 증가가 시작되어 이후 지속적으로 높은 발현 수준을 유지하였다.



  1. Babin, P., C. Thisse, M. Durliat, M. Andre, M.-A. Akimenko and B. Thisse, 1997. Both apolipoprotein E and A-I genes are present in a nonmammalian vertebreate and are highly expressed during embryonic development. Proc. Natl. Acad. Sci. USA, 94, 8622-8627
  2. Babin, P. J., 1987. Plasma lipoprotein and apolipoprotein distribution as a function of density in the rainbow trout (Salmo gairdneri). Biochem. J., 246, 425-429
  3. Boguski, M. S., N. Elshourbagy, J. M. Taylor and J. I. Gordon, 1985. Comparative analysis of repeated sequences in rat apolipoproteins A-I, A-IV, and E. Proc. Natl. Acad. Sci. USA, 82, 992-996
  4. Chen, W.Y., J.A.C. John, C.-H. Lin, H.-F. Lin, S.-C. Wu, C.-H. Lin, C.-Y. Chang, 2004. Expression of metallothionein gene during embryonic and early larval development in zebrafish. Aquat. Toxicol., 69, 215-227
  5. Chenna, R., H. Sugawara, T. Koike, R. Lopez, T. J. Gibson, D. G. Higgins and J. D. Thompson, 2003. Multiple sequence alignment with the clustal series of programs. Nucleic Acids Res., 31, 3497-3500
  6. Christoffels, A., E. G. L. Koh, J.-M. Chia, S. Brenner, S. Aparicio and B. Venkatesh, 2004. Fugu genome analysis provides evidence for a whole-genome duplication early during the evolution of ray-finned fishes. Mol. Biol. Evol., 21, 1146-1151
  7. Delcuve, G. P., J. M. Sun and J. R. Davie, 1992. Expression of rainbow trout apolipoprotein A-I genes in liver and hepatocellular carcinoma. J. Lipid Res., 33, 251-262
  8. Frank, P. G and Y. L. Marcel, 2000. Apolipoprotein A-I: structurefunction relationships. J. Lipid Res., 41, 853-872
  9. Hall, T. A., 1999. BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symp. Ser., 41, 95-98
  10. Jaillon, O., J. M. Aury, F. Brunet, J. L. Petit, N. Stange-Thomann, E. Mauceli, L. Bouneau, C. Fischer, et al., 2004. Genome duplication in the teleost fish Tetraodon nigroviridis reveals the early vertebrate proto-karyotype. Nature, 431, 946-957
  11. Khuseyinova, N. and W. Koenig, 2006. Apolipoprotein A-I and risk for cardiovascular diseases. Curr. Atheroscler Rep., 8, 365-373
  12. Kim, D. S., J. Y. Jo and T. Y. Lee, 1994. Induction of triploidy in mud loach (Misgurnus mizolepis) and its effect on gonadal development and growth. Aquaculture, 120, 263-270
  13. Kondo, H., I. Kawazoe, M. Nakaya, K. Kikuchi, K. Aida and S. Watabe, 2001. The novel sequences of major plasma apolipoproteins in the eel Anguilla japonica. Biochim. Biophys. Acta, 1531, 132-142
  14. Kondo, H., K. Morinaga, R. Misaki, M. Nakaya and S. Watabe, 2005. Characterization of the pufferfish Takifugu rubripes apolipoprotein multigene family. Gene, 346, 257-266
  15. Kumar, S., K. Tamura and M. Nei, 2004. MEGA3: Integrated software for molecular evolutionary genetics analysis and sequence alignment. Brief. Bioinform., 5, 150-163
  16. Li, W.-H., M. Tanimura, C.-C. Luo, S. Datta and L. Chan, 1988. The apolipoprotein multigene family: biosynthesis, structure, structure-function relationships, and evolution. J. Lipid Res., 29, 245-271
  17. Llewellyn, L., V. P. Ramsurn, T. Wigham, G. E. Sweeney and D. M. Power, 1998. Cloning, characterization and expression of the apolipoprotein A-I gene in the sea bream (Sparus aurata). Biochim. Biophys. Acta, 1442, 399-404
  18. Nam, Y. K., 2005. Tailoring fish genome and transgenic manipulation as exemplified by mud loach (Misgurnus mizolepis). Fish Genet. Breed. Sci., 35, 113-121
  19. Wallaert, C. and P. J. Babin, 1994. Age-related, sex-related, and seasonal changes of plasma lipoprotein concentrations in trout. J. Lipid Res., 35, 1619-1633