Molecular Characterization of A New IgZ3 Subclass from Common Carp (Cyprinus Carpio) and Comparative Expression Analysis of IgH Transcripts During Ontogeny of Larvae
Authors: Zhang Fumiao , Li Mojin , Lv Cui , Wei Guangcai , Wang Chang , Wang Yimeng , An Liguo , Yang Guiwen 
Published: December 2020

Abstract
Background Immunoglobulins distributing among systemic immune tissues and mucosal immune tissues play an important role in teleost to protect them from infections in a pathogen-rich aquatic environment. Teleost IgZ/IgT subclasses with different tissue expression patterns may have varied immune functions.Results In the present study, a novel secreted IgZ heavy chain gene was cloned and characterized in common carp (Cyprinus carpio) which was different from the reported IgZ1 and IgZ2 in tissue-specific expression profile. The obtained IgZ-like subclass was designated as CcIgZ3 which complete open reading frame contained 1650 bp encoding a protein of 549 amino acid residues. The phylogenetic analysis revealed that CcIgZ3 was grouped with carp IgZ2 and was in the same branch as other teleosts IgZ/IgT. Basal expression detection of IgH in healthy adult common carp showed that CcIgZ3 transcripts were widely expressed in systemic immune tissues and Mucosal-associated lymphoid tissues. It was expressed at a higher level in the head kidney, gill, and gonad, followed by spleen, hindgut, oral epithelium, liver, brain, muscle, foregut, and blood, but at a very low level in the skin. The transcript level of CcIgZ3 mRNA in isolated leukocytes from peripheral blood cells was significantly higher than that of isolated leukocytes from the spleen. Different groups of common carp were infected with Aeromonas hydrophila via intraperitoneal injection and immersion respectively. The qRT-PCR analysis demonstrated that a significant difference in CcIgZ3 mRNA level between immersion and injection groups existed in all the detected tissues including head kidney, spleen, liver, and hindgut, especially in hindgut CcIgZ3 mRNA level was higher in immersion than in the injection group. Different routes for Aeromonas hydrophila challenge to common carp had comparatively less effect on IgM response. Further study on the relative expression of the IgH gene during the ontogeny of common carp presented that the tissue-specific expression profile of CcIgZ3 was very different from the others. In the early larval development stage of common carp from 1 dpf to 31 dpf, the qRT-PCR analysis demonstrated that the CcIgZ3 mRNA level increased gradually with a similar dynamic tendency of IgZ1 and IgZ2 and IgM was the dominant Ig with obviously higher abundance. The results of tissue-specific expression of IgH at 65 dpf of common carp showed that CcIgZ3 was expressed at mucosal sites including both hindgut and gill but IgZ1 was preferentially expressed in hindgut and IgZ2 in gill. Except for qRT-PCR analysis, the detection of the CcIgZ3-expressing cells and IgM-expressing cells by in situ hybridization was performed. The results showed that CcIgZ3 and IgM transcripts could be detected in the spleen, gill, and hindgut of the common carp at 65dpf. Conclusions These results revealed that CcIgZ3 gene transcript occurred in the early development stage of common carp not only in systemic tissues but also in mucosal tissues. CcIgZ3 could be induced with significantly different expression profiles at immune tissues after the challenge by Aeromonas hydrophila immersion and intraperitoneal injection, which indicated that CcIgZ3 might play a more important role in mucosal immunity than in systemic immunity.