Reprimo tissue-specific expression pattern is conserved between zebrafish and human.

Ricardo J Figueroa,Alejandro H Corvalán,Gonzalo Carrasco-Avino,Martin Lange,Juan C Opazo,Ignacio A Wichmann,Arndt F Siekmann,Gareth I Owen,Julio D Amigo,Marc Robinson-Rechavi

doi:10.1371/journal.pone.0178274

Abstract

Reprimo (RPRM), a member of the RPRM gene family, is a tumor-suppressor gene involved in the regulation of the p53-mediated cell cycle arrest at G2/M. RPRM has been associated with malignant tumor progression and proposed as a potential biomarker for early cancer detection. However, the expression and role of RPRM, as well as its family, are poorly understood and their physiology is as yet unstudied. In this scenario, a model system like the zebrafish could serve to dissect the role of the RPRM family members in vivo. Phylogenetic analysis reveals that RPRM and RPRML have been differentially retained by most species throughout vertebrate evolution, yet RPRM3 has been retained only in a small group of distantly related species, including zebrafish. Herein, we characterized the spatiotemporal expression of RPRM (present in zebrafish as an infraclass duplication rprma/rprmb), RPRML and RPRM3 in the zebrafish. By whole-mount in situ hybridization (WISH) and fluorescent in situ hybridization (FISH), we demonstrate that rprm (rprma/rprmb) and rprml show a similar spatiotemporal expression profile during zebrafish development. At early developmental stages rprmb is expressed in somites. After one day post-fertilization, rprm (rprma/rprmb) and rprml are expressed in the notochord, brain, blood vessels and digestive tube. On the other hand, rprm3 shows the most unique expression profile, being expressed only in the central nervous system (CNS). We assessed the expression patterns of RPRM gene transcripts in adult zebrafish and human RPRM protein product in tissue samples by RT-qPCR and immunohistochemistry (IHC) staining, respectively. Strikingly, tissue-specific expression patterns of the RPRM transcripts and protein are conserved between zebrafish and humans. We propose the zebrafish as a powerful tool to elucidate the both physiological and pathological roles of the RPRM gene family.

Highlights

The Reprimo (RPRM) gene family has only been identified in vertebrates and that is composed by three paralogs: RPRM, RPRML and RPRM3 [1]
Multiple sequence alignment of RPRM, RPRML and RPRM3 proteins between human, mouse and zebrafish showed a striking conservation of the C-terminus across species (Fig 2), while the N-terminus domain retained the most variability within the protein family
The high level of conservation (72.5% and 66% identity between human RPRM and zebrafish rprma, rprmb and rprm3 proteins, respectively; 55.7% identity between human and zebrafish RPRML proteins) observed in the RPRM protein sequences and domains suggests the functions of the proteins may be related; though further research is warranted to support this conclusion

Summary

Introduction

The Reprimo (RPRM) gene family has only been identified in vertebrates and that is composed by three paralogs: RPRM, RPRML and RPRM3 [1]. This group of genes diversified during the two rounds of whole genome duplication that occurred early in the evolution of vertebrates. RPRM is a potential class II (inactivation occurring by epigenetic mechanisms) tumor suppressor gene (TSG). Inactivation is driven principally by aberrant methylation of its promoter region [4, 7, 8] and correlates with genetic instability in human cancers, such as gastrointestinal tumors [9, 10]. No literature exists on the presence or role of Reprimo-like (RPRML) protein in either physiology or pathophysiology

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Conclusion