The Expression Pattern Of Bcl6 And Regulation Of Bcl6a By Prdml In Medaka

Both Bcl6 and Prdml are transcription factors,play important roles in the regulation of gene expression,cell differentiation,and immune response.In this study,we focus on the expression pattern of bcl6 and the regulation of bcl6a by Prdm 1 in medaka.Medaka Bcl6a and Bcl6B both contain N-terminal BTB/POZ domain and C-terminal zinc finger motifs,which are evolutionarily conserved.We have analyzed the expression of bcl6a and bcl6B at different embryonic development stages by RT-PCR,Q-PCR and in situ hybridization.The results showed that bcl6a and bcl6B were expressed during embryonic development,implying their role in embryonic development.bcl6a was highly expressed at 4-cell stage,blastocyst and gastrulation stage.The expression of bcl6a decreased with cell cleavage,and low expression of bcl6a was found during later embryonic development.And bcl6B was slightly expressed at early embryonic development stages,increased at 1 day post-fertilization(dpf),and maintained high expression from ldpf to fry.These indicated the homologous genes bcl6a and bcl6B may play a role at differernt embryonic stages.Oryzias latipes bcl6a and bcl6B could be detected in many tissues by RT-PCR and Q-PCR.The results showed that the expression of bcl6a is high in the liver and kidney,and bcl6B was highly expressed in spleen.bcl6a and bcl6B were highly expressed in these immune organs hinted their roles in fish immune response.In order to explore the regulation of bcl6a by Prdml,we have got bcl6a promoter sequence by PCR.The sequence contained 3 Prdml binding sites by software analysis.We have constructed the truncated fragments containing diflferent binding sites.To test promoter activity,the CO and CIK cell lines were transfected with pGL3-bcl6a and different truncated fragments,the results showed pGL3-bcl6a,pGL3-388,pGL3-467,pGL3-788,pGL3-910 exhibited different extent activity.Then the CO was transfected with pGL3-bcl6a and different truncated fragments,and co-transfected with recombinant plasmid pCS-prdmla and pCS-prdmlb.We observed Prdmla and Prdmlb could inhibit bcl6a promoter activity.Binding site 1 and 2 play a major role during the inhibition process.This inhibition of bcl6a by Prdml may play an important role in the fish immune system.Furthermore,medaka abdominal cavity was injected with LPS and PolyI:C,and we have analyzed the change of bcl6a expression in liver after immune stimuli by Q-PCR.Compared with the control injected with PBS,The expression of bcl6a was elevated obviously in 6 h after injection of polyI:C,and also increased in 12h after injection of LPS.These results further suggested that medaka bcl6a may play some roles in immune system.