Genomic localization and immune response of IgT in grass carp (Ctenopharyngodon idellus)

Teleost fish have three types of immunoglobulins (Ig): IgM, IgD, and IgT, of which IgT is unique to teleost fish. Though grass carp (Ctenopharyngodon idella) is one of the most farmed freshwater fish species worldwide, the genomic localization of the heavy chain gene of IgT (IGHT) and the composition and usage of the heavy chain variable region (VH) genes remain unknown. In this study, tissue expression pattern analysis revealed that grass carp IgT is constitutively expressed in various tissues, with higher expression levels in the spleen, kidney, and skin. After immersion immunization with Flavobacterium columnare, the expression of grass carp IgT in the skin increased dramatically both at the mRNA and protein levels. Subsequently, we amplified the genomic DNA of IGHT and localized it on chromosome III, approximately 28 kbp upstream of the heavy chain constant region (CH) gene of IgM3. A total of 93 IgT VH genes were predicted and localized on chromosome III. Sequence analysis showed that these VH genes can be divided into 12 subgroups, including VH1, VH2, VH3, VH4, VH5, VH6, VH8, VH9, VH10, VH11, VH12, and VH13. High-throughput sequencing revealed that IgT preferentially utilizes the V12-2 gene, and the frequency of V12-2 gene usage increased dramatically after F. columnare infection. The results of this study expand our understanding of the immune response of teleost IgT to bacterial infection at the genomic, mRNA, and protein levels. In addition, the identification and analysis of VH genes have laid an important foundation for identifying pathogen specific IgT.