DEAD-Box Protein Ddx46 Is Required for the Development of the Digestive Organs and Brain in Zebrafish

Shunya Hozumi,Atsushi Kuroiwa,Akio Yoshizawa,Ryo Hirabayashi,Tohru Ishitani,Motoyuki Itoh,Yutaka Kikuchi,Makiko Tsutsumi,Mitsuko Ogata,Laszlo Orban

doi:10.1371/journal.pone.0033675

Shunya Hozumi, Atsushi Kuroiwa + Show 8 more

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https://doi.org/10.1371/journal.pone.0033675

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Journal: PloS one	Publication Date: Mar 19, 2012
Citations: 60	License type: CC BY 4.0

Affiliation: Hiroshima University, Nagoya University

Abstract

Spatially and temporally controlled gene expression, including transcription, several mRNA processing steps, and the export of mature mRNA to the cytoplasm, is essential for developmental processes. It is well known that RNA helicases of the DExD/H-box protein family are involved in these gene expression processes, including transcription, pre-mRNA splicing, and rRNA biogenesis. Although one DExD/H-box protein, Prp5, a homologue of vertebrate Ddx46, has been shown to play important roles in pre-mRNA splicing in yeast, the in vivo function of Ddx46 remains to be fully elucidated in metazoans. In this study, we isolated zebrafish morendo (mor), a mutant that shows developmental defects in the digestive organs and brain, and found that it encodes Ddx46. The Ddx46 transcript is maternally supplied, and as development proceeds in zebrafish larvae, its ubiquitous expression gradually becomes restricted to those organs. The results of whole-mount in situ hybridization showed that the expression of various molecular markers in these organs is considerably reduced in the Ddx46 mutant. Furthermore, splicing status analysis with RT-PCR revealed unspliced forms of mRNAs in the digestive organ and brain tissues of the Ddx46 mutant, suggesting that Ddx46 may be required for pre-mRNA splicing during zebrafish development. Therefore, our results suggest a model in which zebrafish Ddx46 is required for the development of the digestive organs and brain, possibly through the control of pre-mRNA splicing.

Highlights

Precursor mRNA splicing is essential for gene expression in metazoan cells, and the splicing reaction proceeds via a coordinated series of RNA-RNA, RNA-protein, and proteinprotein interactions, which lead to exon ligation and the release of the intron lariat [1]–[4]
Histochemical and immunohistochemical analyses exhibited that the exocrine pancreas and liver in the morha4 mutant were smaller than those in wild-type (WT) larvae (Figure 1I–L, Figure S1), whereas the size of the endocrine pancreas was normal in this mutant (Figure 1I and J)
Hydrolysis of Prp5 is necessary for the stable association of U2 small nuclear ribonucleoprotein particles (snRNPs) with pre-mRNA and pre-spliceosome formation in S. cerevisiae and S. pombe [6], [7]

Summary

Introduction

Precursor mRNA (pre-mRNA) splicing is essential for gene expression in metazoan cells, and the splicing reaction proceeds via a coordinated series of RNA-RNA, RNA-protein, and proteinprotein interactions, which lead to exon ligation and the release of the intron lariat [1]–[4]. The DExD/H-box helicases share nine conserved motifs; motifs Q, I, II, and VI are required for NTP/ATP binding and catalyze its hydrolysis [5]–[8]. These proteins have been shown to play important roles in all aspects of RNA metabolism, including the modulation of RNA structures and association/dissociation of RNA-protein complexes, such as pre-mRNA splicing, rRNA biogenesis, transcription, RNA stability and turnover, RNA export, and translation [5]–[8]. Prp (a homologue of vertebrate Ddx46) is necessary, along with ATP hydrolysis, for stable association of U2 snRNP with pre-mRNA and prespliceosome formation in S. cerevisiae and Schizosaccharomyces pombe [9]–[11]. The in vivo function of Ddx in metazoans remains to be elucidated,

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