D-dopachrome tautomerase from Japanese sea bass(Lateolabrax japonicus) is a chemokine-like cytokine and functional homolog of macrophage migration inhibitory factor

D-dopachrome tautomerase(DDT),a member of the macrophage migration inhibitory factor(MIF)protein superfamily,is a newly described cytokine with chemokine-like characteristics.However,research on fish DDT remains limited.In this study,we identified a DDT homolog(LjDDT)from the Japanese sea bass,Lateolabrax japonicus.Sequence analysis showed that LjDDT had typical sequence features of known DDT and MIF homologs and was most closely related to DDT of rock bream(Oplegnathus fasciatus).LjDDT transcripts were detected in all tested tissues of healthy Japanese sea bass,with the highest expression found in the liver.Upon infection with Vibrio harveyi,LjDDT transcripts were significantly down-regulated in the three tested tissues,including the liver,spleen,and head kidney.Recombinant LjDDT(rLjDDT)and the corresponding antibody(anti-rLjDDT)were subsequently prepared.The administration of 100μg/g anti-rLjDDT had a statistically significant protective effect on the survival of V.harveyi-infected fish.Moreover,rLjDDT was able to induce the migration of monocytes/macrophages(MO/MФ)and lymphocytes both in vitro and in vivo,but without significant influence on the migration of neutrophils.rLjDDT exhibited chemotactic activity for lipopolysaccharide(LPS)-stimulated M1-type MO/MΦin vitro,but not for cAMP-stimulated M2-type MO/MΦ.Furthermore,the knockdown of LjCD74,but not LjCXCR4,significantly down-regulated the rLjDDT-enhanced migration of MO/MΦand relieved the rLjMIF-inhibited migration of MO/MΦ.These results indicate that LjCD74 may be the major chemotactic receptor of LjDDT and LjMIF in Japanese sea bass MO/MΦ.Combined rLjDDT+rLjMIF treatment had no significant effect on the migration of MsiRNA,LjCD74si-,or LjCXCR4sitreated MO/MΦcompared to the control group,suggesting that the roles of LjDDT and LjMIF may be antagonistic.In conclusion,our study demonstrates for the first time that DDT may play a role in the immune responses of fish against bacterial infection through chemotactic recruitment of MO/MΦvia mediation of CD74 as an antagonist of MIF.

The Characteristics of vasa Gene from Japanese Sea Bass(Lateolabrax japonicas) and Its Response to the External Hormones

The RNA helicase Vasa is an important regulator of primordial germ cell development. Its function in mature fish, espe- cially the hormone-related differences in maturing male fish has seldom been documented. In this study, a full length cDNA sequence of the vasa gene was cloned from Japanese sea bass, Lateolabraxjaponicas, and it was namedjsb-vasa. Homology analysis showed thatjsb-vasa was closely related to its teleost homologs. The spatial distribution ofjsb-vasa indicated that it was only highly ex- pressed in testis, showing its germ cell-specific expression pattern. During the testicular development cycle, jsb-vasa was highly expressed during early period of spermatogenesis, and reduced when spermatogenesis advanced. In addition, the jsb-vasa gene ex- pression was significantly inhibited at 6 h, 12 h and 24 h after injecting hCG （human ehorionic gonadotropin） and GnRHa （Gonad- otropin-releasing hormone analogue）, indicating thatjsb-vasa gene may play an important role in spermatogenesis of Japanese sea bass, and be under the regulation of external sex hormones.