| dc.description.abstract | In early embryonic development, the maternal-to-zygotic transition (MZT) is a crucial process, which involves two main events, the zygotic genome activation (ZGA) and the degradation of maternal components. In Drosophila embryos, the transcription factor Zelda (Zld) is known as the key activator to initiate ZGA. During MZT, Zelda serves multiple roles, mainly (1) as a strong activator that directly triggers ZGA, (2) as a pioneer factor, which enhances chromatin accessibility, thus facilitating the binding of the other transcription factors and promoting transcription. Previous analyses using Chromatin Immunoprecipitation- Sequencing (ChIP-seq) and Micrococcal Nuclease-Sequencing (MNase-seq) in early Drosophila embryos, have confirmed its role as a pioneer factor in regulating the chromatin landscape. Currently, we asked whether Zelda has the potential to reprogram the transcriptome in differentiated Drosophila cells. We were able to use the Bac-to-Bac expression system to ectopically express Zelda in the Drosophila late-stage embryonic cells, S2 cells. RNA-seq data revealed that Zelda could re-activate approximately 75% of early genes, along with markers of adult stem cells, indicating a potential for reprogramming. In my study, we asked whether Zelda is able to remodel chromatin landscape in S2 cells.
I first optimized the yield of recombinant baculovirus and the efficiency of Zelda expression in S2 cells, by attaining approximately 100-fold increase in infectivity. Secondly, I conducted ATAC-seq (Assay for Transposase-Accessible Chromatin using sequencing) to examine the effects of Zelda on chromatin in S2 cells, and investigated its correlation to the expression and Zelda binding profiles from previous lab results and literatures. Overall, 62,414 and 28,275 open regions were recovered in the control group and experimental group, respectively. The effects of Zelda on chromatin could be categorized into three classes. (1) Regions that were originally more closed became more open and concentrated in the presence of Zelda. The openness was positively correlated with the strength of Zelda binding and associated with up-regulated genes nearby the ATAC peaks. This suggested that Zelda has the ability to increase chromatin accessibility in S2 cells. (2) Regions that were originally open and associated with gene expression became closed within Zelda expression. Zelda may bind to these regions due to their high-accessibility. Intriguingly, Zelda binding led to more closed chromatin and down-regulated genes. Whether it is the direct effect of Zelda requires further investigation. (3) Regions that were unchanged with or without the presence of Zelda. Further analysis of the differences among the three classes may provide more insights of Zelda function and define Zelda-responsive features of chromatin. | en_US |