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Vegetally localised Vrtn functions as a novel repressor to modulate bmp2b transcription during dorsoventral patterning in zebrafish

ABSTRACT: The vegetal pole cytoplasm represents a crucial source of maternal dorsal determinants for patterning the dorsoventral axis of the early embryo. Removal of the vegetal yolk in the zebrafish fertilised egg before the completion of the first cleavage results in embryonic ventralisation, but removal of this part at the two-cell stage leads to embryonic dorsalisation. How this is achieved remains unknown. Here, we report a novel mode of maternal regulation of BMP signaling during dorsoventral patterning in zebrafish. We identify Vrtn as a novel vegetally localised maternal factor with dorsalising activity and rapid transport towards the animal pole region after fertilisation. Co-injection of vrtn mRNA with vegetal RNAs from different cleavage stages suggests the presence of putative vegetally localised Vrtn antagonists with slower animal pole transport. Thus, vegetal ablation at the two-cell stage could remove most of the Vrtn antagonists, and allows Vrtn to produce the dorsalising effect. Mechanistically, Vrtn binds a bmp2b regulatory sequence and acts as a repressor to inhibit its zygotic transcription. Analysis of maternal-zygotic vrtn mutants further shows that Vrtn is required to constrain excessive bmp2b expression in the margin. Our work unveils a novel maternal mechanism regulating zygotic BMP gradient in dorsoventral patterning.

ORGANISM(S): Danio rerio

PROVIDER: GSE103090 | GEO | 2017/08/26

SECONDARY ACCESSION(S): PRJNA400171

REPOSITORIES: GEO

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Project description:In all animals, the initial events of embryogenesis are controlled by maternal gene products that are deposited into the developing oocyte. At some point after fertilization, control of embryogenesis is transferred to the zygotic genome in a process called the maternal to zygotic transition (MZT). During this time maternal RNAs are degraded and zygotic RNAs are transcribed1. A long standing question has been, what factors regulate these events? The recent findings that microRNAs and Smaugs mediate maternal transcript degradation brought new life to this old problem2,3, however, the transcription factors that activate zygotic gene expression remained elusive. A clue came from the finding that many early zygotic genes in Drosophila share a cis-regulatory heptamer motif, CAGGTAG and related sequences, collectively referred to as TAG-team sites4,5. We asked whether there was a dedicated transcription factor that interacts with these sites to activate early genes. Here we report the discovery of a zinc-finger protein, Zelda (Zld) that binds specifically to TAG-team sites, and is capable of activating transcription in transient transfection assays. Mutant embryos lacking zld are defective in the cellularization process, and fail to activate the transcription of many early zygotic genes involved in cellularization, sex determination, and dorsoventral patterning. Global expression profiling confirmed that Zld plays a key role in the activation of the early zygotic genome, and suggests that Zld may also play a role in maternal RNA degradation during the MZT since many RNAs are up-regulated in the absence of Zld. Experiment Overall Design: Total RNA samples were extracted from three replicate collections of 1-2 hr yw and M- zld embryos by TRIzol (invitrogen). A portion of the collected embryos was fixed and stained with DAPI; 90% were in nuclear cycles 8 to13. cDNA was prepared using the GeneChipÂ® HT One-Cycle cDNA Synthesis Kit (Manufactured by Invitrogen for Affymetrix) and labeled with the BioArrayâ¢ HighYieldâ¢ RNA Transcript Labeling Kit (Enzo). Labeled probes were hybridized to Drosophila Genome 2 Affymetrix arrays and processed by a GeneChip Fluidics Station 400. Data were acquired and normalized by the GeneChipÂ® Scanner 3000 and processed by the Affymetrix GeneChip Operating Software (GCOS). t-test analysis was performed on the data from three biological replicates.