Expression Analysis of Heavy-Chain-Only Antibodies in Cloudy Catshark and Japanese Bullhead Shark
Reo Uemura,
Susumu Tanimura,
Nao Yamaguchi,
Ryuichi Kuroiwa,
Gabriel Takashi Andrés Tsutsumi,
Toshiaki Fujikawa,
Kiyoshi Soyano,
Kohsuke Takeda,
Yoshimasa Tanaka
Affiliations
Reo Uemura
Department of Cell Regulation, Graduate School of Biomedical Sciences, Nagasaki University, Bunkyo-machi 1-14, Nagasaki 852-8521, Japan
Susumu Tanimura
Department of Cell Regulation, Graduate School of Biomedical Sciences, Nagasaki University, Bunkyo-machi 1-14, Nagasaki 852-8521, Japan
Nao Yamaguchi
Department of Cell Regulation, Graduate School of Biomedical Sciences, Nagasaki University, Bunkyo-machi 1-14, Nagasaki 852-8521, Japan
Ryuichi Kuroiwa
Department of Cell Regulation, Graduate School of Biomedical Sciences, Nagasaki University, Bunkyo-machi 1-14, Nagasaki 852-8521, Japan
Gabriel Takashi Andrés Tsutsumi
Department of Cell Regulation, Graduate School of Biomedical Sciences, Nagasaki University, Bunkyo-machi 1-14, Nagasaki 852-8521, Japan
Toshiaki Fujikawa
Institute for East China Sea Research, Organization for Marine Science and Technology, Nagasaki University, 1551-7, Taira-machi, Nagasaki 851-2213, Japan
Kiyoshi Soyano
Institute for East China Sea Research, Organization for Marine Science and Technology, Nagasaki University, 1551-7, Taira-machi, Nagasaki 851-2213, Japan
Kohsuke Takeda
Department of Cell Regulation, Graduate School of Biomedical Sciences, Nagasaki University, Bunkyo-machi 1-14, Nagasaki 852-8521, Japan
Yoshimasa Tanaka
Center for Medical Innovation, Nagasaki University, Sakamoto 1-7-1, Nagasaki 852-8588, Japan
Heavy chain-only antibodies in sharks are called immunoglobulin new antigen receptors (IgNAR), consisting of one variable region (VNAR) and five constant regions (C1-C5). The variable region of IgNAR can be expressed as a monomer composed of a single domain, which has antigen specificity and is thus gaining attention as a next-generation antibody drug modality. In this study, we analyzed IgNAR of the cloudy catshark and Japanese bullhead shark, small demersal sharks available in the coastal waters of Japan. By analyzing the IgNAR gene sequence and comparing it with the constant regions of five other known shark species, high homology was observed in the C4 region. Consequently, we expressed the recombinant protein of the C4 domain from the cloudy catshark in E. coli, immunized rats, and produced antibodies. The obtained antiserum and mAbs recognized the C4 recombinant protein of the cloudy catshark, but reacted minimally with the plasma of non-immunized cloudy catsharks and instead reacted with the plasma of Japanese bullhead sharks. The results of this study imply that the protein expression levels of IgNAR in cloudy catsharks may be relatively lower compared to those in Japanese bullhead sharks, however, this interpretation remains to be determined through further studies.
