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Expression of Hr-Erf Gene during Ascidian Embryogenesis
Jung Eun Kim1, Won Young Lee1, Gil Jung Kim1
1Department of Marine Molecular Biotechnology, Gangneung-Wonju National University, Gangneung 210-702, Republic of Korea.
Abstract:
FGF9/16/20 signaling pathway specify the developmental fates of notochord, mesenchyme, and neural cells in ascidian embryos. Although a conserved Ras/MEK/Erk/Ets pathway is known to be involved in this signaling, the detailed mechanisms of regulation of FGF signaling pathway have remained largely elusive. In this study, we have isolated Hr-Erf, an ascidian orthologue of vertebrate Erf, to elucidate interactions of transcription factors involved in FGF signaling of the ascidian embryo. The Hr-Erf cDNA encompassed 3110 nucleotides including sequence encoded a predicted polypeptide of 760 amino acids. The polypeptide had the Ets DNA-binding domain in its N-terminal region. In adult animals, Hr-Erf mRNA was predominantly detected in muscle, and at lower levels in ganglion, gills, gonad, hepatopancreas, and stomach by quantitative real-time PCR (QPCR) method. During embryogenesis, Hr-Erf mRNA was detected from eggs to early developmental stage embryos, whereas the transcript levels were decreased after neurula stage. Similar to the QPCR results, maternal transcripts of Hr-Erf was detected in the fertilized eggs by whole-mount in situ hybridization. Maternal mRNA of Hr-Erf was gradually lost from the neurula stage. Zygotic expression of Hr-Erf started in most blastomeres at the 8-cell stage. At gastrula stage, Hr-Erf was specifically expressed in the precursor cells of brain and mesenchyme. When MEK inhibitor was treated, embryos resulted in loss of Hr-Erf expression in mesenchyme cells, and in excess of Hr-Erf in a-line neural cells. These results suggest that zygotic Hr-Erf products are involved in specification of mesenchyme and neural cells.
Insights
We identified Hr-Erf, a key transcription factor in ascidian embryos, crucial for specifying mesenchyme and neural cell fates via the FGF signaling pathway. Its expression patterns reveal its role in early development.
Area of Science:
- Developmental Biology
- Molecular Biology
- Genetics
Background:
- Fibroblast Growth Factor (FGF) signaling pathways regulate cell fate determination in ascidian embryos, including notochord, mesenchyme, and neural cells.
- While the Ras/MEK/Erk/Ets pathway is implicated, the precise regulatory mechanisms of FGF signaling remain unclear.
Purpose of the Study:
- To isolate and characterize Hr-Erf, an ascidian ortholog of vertebrate Erf, to investigate its role in FGF signaling during ascidian embryogenesis.
- To elucidate the interactions of transcription factors within the FGF signaling pathway in ascidian embryos.
Main Methods:
- Isolation and sequencing of Hr-Erf cDNA.
- Quantitative real-time PCR (QPCR) for analyzing Hr-Erf mRNA expression in adult tissues and during embryogenesis.
- Whole-mount in situ hybridization to visualize maternal and zygotic Hr-Erf mRNA distribution.
- Treatment with a MEK inhibitor to observe effects on Hr-Erf expression patterns.
Main Results:
- Hr-Erf cDNA encodes a 760-amino acid polypeptide with an Ets DNA-binding domain.
- Hr-Erf mRNA is predominantly found in adult muscle, with lower levels in other tissues.
- Maternal Hr-Erf transcripts are present in fertilized eggs and decrease after the neurula stage.
- Zygotic Hr-Erf expression begins at the 8-cell stage, with specific expression in brain and mesenchyme precursors during gastrulation.
- MEK inhibition disrupts Hr-Erf expression in mesenchyme and neural cells.
Conclusions:
- Zygotic Hr-Erf plays a significant role in the specification of mesenchyme and neural cells in ascidian embryos.
- The study provides insights into the regulatory mechanisms of FGF signaling and transcription factor interactions during early development.
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