Interaction Between Chicken (Gallus gallus) E2F1 and TBP Protein and Analysis of Their Gene Expression Characteristics in Tissues
TANG Hong1,2, WANG Yan-Bi1,2, ZHAO Cai-Qin1,2, WANG Xiao-Ya1,2, SHI Hai-Ying1,2, XING Jing-Ru1,2, DUAN Zhi-Qiang1,2*
1 Key Laboratory of Plateau Mountain Animal Genetics, Breeding and Reproduction, Ministry of Education/Guizhou Province Key Laboratory of Animal Genetics, Breeding and Reproduction, Guizhou University, Guiyang 550025, China; 2 College of Animal Science, Guizhou University, Guiyang 550025, China
Abstract：E2F transcription factor 1 (E2F1) and TATA-binding protein (TBP) jointly participate in the gene transcription regulation. To investigate the interaction between chicken (Gallus gallus) E2F1 and TBP proteins and their gene expression characteristics in tissues, in this study, the recombinant eukaryotic expression vectors of chicken E2F1 and TBP genes were constructed and then transfected into human embryonic kidney cells (HEK-293T). Indirect immunofluorescence and co-immunoprecipitation (Co-IP) assays were performed to verify their interaction. In addition, qPCR assay was used to examine the expression levels of E2F1 and TBP genes in different tissues at 14 embryonic days (E14 d) in chick embryos and 1-day-old (H1 d), 7-day-old (H7 d), 14-day-old (H14 d) after shelling. The results showed that the recombinant vectors pCMV-HA-E2F1 and pCMV-Myc-TBP of chicken were successfully constructed, and the recombinant proteins HA-E2F1 and Myc-TBP were correctly expressed and mainly localized in the nucleus in plasmid-transfected cells. The results of tissue characteristics analysis showed that chicken TBP and E2F1 genes were expressed in different chicken tissues at 4 stages, and the expression levels were significant different (P<0.05). At E14 d, the expression of TBP and E2F1 genes was higher in lung, but lower in brain; at H1 d, the 2 gens had high expression level in liver, but low in brain; at H7 d, they showed high expression in brain, but low in leg muscles; at H14 d, there were high expression in lung, liver and heart, but low in chest and leg muscles. Chicken E2F1 protein could interact with TBP protein in the nucleus, and the expression levels of E2F1 and TBP genes were relatively high in lung and liver at different developmental stages. The results provide references for further studying the mechanism of E2F1 and TBP protein interaction regulating the development of chicken lung and liver tissues.
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