Combinatorial regulation by maternal transcription factors during activation of the endoderm gene regulatory network
- Author(s): Paraiso, Kitt
- Advisor(s): Cho, Ken W.Y.
- et al.
Zygotic gene activation (ZGA) is the process in the earliest stages of animal development when maternal transcription factors activate the expression of the first zygotic genes. In most metazoans, this process is coupled with specification of the three germ layers. In Xenopus, these two processes are initiated by the function of maternal transcription factors (TFs), which are localized in the egg and asymmetrically distributed to different cells of the embryo during cleavage stages. In this thesis, I uncover the combinatorial function of vegetally-localized TFs that are involved in the activation of the endodermal fate.
By gain- and loss-of-function approaches, I elucidate a novel function of the TF Otx1 in specification of the endoderm. Together with the well-known endodermal TF Vegt, Otx1 activates endodermal genes, while inhibiting mesodermal genes in the presumptive endoderm. By chromatin immunoprecipitation followed by qPCR or sequencing, I show that Vegt and Otx1 co-bind in the chromatin with the ubiquitously expressed Foxh1 prior to ZGA. These maternal TFs initiate the emergence of cis-regulatory regions by docking to clusters of consensus DNA binding motifs. The binding of their assembly pre-marks regions of active cis-regulation, which are subsequently decorated with epigenetic enhancer marks and then bound by zygotically-active endodermal TFs.
In addition to co-regulation between maternal TFs, by curating evidence to generate the Xenopus endoderm gene regulatory network (GRN), I find evidence that maternal TFs co-regulate downstream target genes with zygotic TFs. Analysis of the Xenopus endodermal GRN suggests that feed-forward loops, where a maternal TF activates a primary zygotic gene encoding a TF, and combinatorially these TFs activate a secondary zygotic gene. From ChIP-seq analysis, the chromatin binding of Vegt and the zygotically active Smad2/3 support this finding. While the functional role of this network motif is unknown, computational modeling suggests that this motif could regulate the multiple waves of zygotic transcription during ZGA.
My findings synthesize the roles of maternal TFs in relation to each other, as well as to the roles of zygotic TFs, using cis-regulatory and gene regulatory network perspectives. The use of multiple approaches provides a broader picture of transcriptional regulation and cell specification during this critical process of ZGA.