Evolution and Spatiotemporal Expression of ankha and ankhb in Zebrafish

TL;DR

This study explores the evolution and expression of two zebrafish genes, ankha and ankhb, which are related to a human gene linked to a rare bone disorder, Craniometaphyseal Dysplasia.

Contribution

The study identifies zebrafish as a potential model for studying CMD by analyzing the spatiotemporal expression and evolutionary relationship of ankha and ankhb.

Findings

Ankhb shows closer evolutionary relationship and stronger promoter activity compared to ankha.

Both ankha and ankhb are expressed in craniofacial regions and somites during zebrafish development.

In silico analysis suggests both paralogs may transport PPi similarly to the human ANK protein.

Abstract

Craniometaphyseal Dysplasia (CMD) is a rare skeletal disorder that can result from mutations in the ANKH gene. This gene encodes progressive anksylosis (ANK), which is responsible for transporting inorganic pyrophosphate (PPi) and ATP from the intracellular to the extracellular environment, where PPi inhibits bone mineralization. When ANK is dysfunctional, as in patients with CMD, the passage of PPi to the extracellular environment is reduced, leading to excess mineralization, particularly in bones of the skull. Zebrafish may serve as a promising model to study the mechanistic basis of CMD. Here, we provide a detailed analysis of the zebrafish Ankh paralogs, Ankha and Ankhb, in terms of their phylogenic relationship with ANK in other vertebrates as well as their spatiotemporal expression patterns during zebrafish development. We found that a closer evolutionary relationship exists between the zebrafish Ankhb protein and its human and other vertebrate counterparts, and stronger promoter activity was predicted for ankhb compared to ankha. Furthermore, we noted distinct temporal expression patterns, with ankha more prominently expressed in early development stages, and both paralogs also being expressed at larval growth stages. Whole-mount in situ hybridization was used to compare the spatial expression patterns of each paralog during bone development, and both showed strong expression in the craniofacial region as well as the notochord and somites. Given the substantial overlap in spatiotemporal expression but only subtle patterning differences, the exact roles of these genes remain speculative. In silico analyses predicted that Ankha and Ankhb have the same function in transporting PPi across the membrane. Nevertheless, this study lays the groundwork for functional analyses of each ankh paralog and highlights the potential of using zebrafish to find possible targeted therapies for CMD.

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