The Reprimo Gene Family: A Novel Gene Lineage in Gastric Cancer with Tumor Suppressive Properties

Abstract

The reprimo (RPRM) gene family is a group of single exon genes present exclusively within the vertebrate lineage. Two out of three members of this family are present in humans: RPRM and RPRM-Like (RPRML). RPRM induces cell cycle arrest at G2/M in response to p53 expression. Loss-of-expression of RPRM is related to increased cell proliferation and growth in gastric cancer. This evidence suggests that RPRM has tumor suppressive properties. However, the molecular mechanisms and signaling partners by which RPRM exerts its functions remain unknown. Moreover, scarce studies have attempted to characterize RPRML, and its functionality is unclear. Herein, we highlight the role of the RPRM gene family in gastric carcinogenesis, as well as its potential applications in clinical settings. In addition, we summarize the current knowledge on the phylogeny and expression patterns of this family of genes in embryonic zebrafish and adult humans. Strikingly, in both species, RPRM is expressed primarily in the digestive tract, blood vessels and central nervous system, supporting the use of zebrafish for further functional characterization of RPRM. Finally, drawing on embryonic and adult expression patterns, we address the potential relevance of RPRM and RPRML in cancer. Active investigation or analytical research in the coming years should contribute to novel translational applications of this poorly understood gene family as potential biomarkers and development of novel cancer therapies.

Keywords: biomarker; development; evolution; gastric cancer; reprimo; tumor suppressive gene properties.

Figure 1

Domain structure and potential post-translational modification sites of human RPRM and RPRML proteins. Schematic representation shows the RPRM and RPRML putative N-glycosylation (green hexagons), serine-phosphorylation (blue circle) and sumoylation (purple triangle) sites. The N-terminal, transmembrane and C-terminal domains are represented by colored boxes (turquoise, yellow, and red, respectively). An * (asterisk) indicates positions which have a single, fully conserved residue. A: (colon) indicates conservation between groups of strongly similar properties—scoring >0.5 in the Gonnet PAM 250 matrix. A. (period) indicates conservation between groups of weakly similar properties—scoring 0.5 in the Gonnet PAM 250 matrix. Post-translational modification sites were predicted using *NetNGlyc (http://www.cbs.dtu.dk/services/NetNGlyc/), **NetPhos (http://www.cbs.dtu.dk/services/NetPhos/) and ***SumoPlot (http://www.abgent.com/sumoplot).

Figure 1

Domain structure and potential post-translational modification sites of human RPRM and RPRML proteins. Schematic representation shows the RPRM and RPRML putative N-glycosylation (green hexagons), serine-phosphorylation (blue circle) and sumoylation (purple triangle) sites. The N-terminal, transmembrane and C-terminal domains are represented by colored boxes (turquoise, yellow, and red, respectively). An * (asterisk) indicates positions which have a single, fully conserved residue. A: (colon) indicates conservation between groups of strongly similar properties—scoring >0.5 in the Gonnet PAM 250 matrix. A. (period) indicates conservation between groups of weakly similar properties—scoring 0.5 in the Gonnet PAM 250 matrix. Post-translational modification sites were predicted using *NetNGlyc (http://www.cbs.dtu.dk/services/NetNGlyc/), **NetPhos (http://www.cbs.dtu.dk/services/NetPhos/) and ***SumoPlot (http://www.abgent.com/sumoplot).

Figure 2

Evolution and diversification of RPRM genes in vertebrates. (A) During the diversification of vertebrates RPRM genes were differentially retained, as not all RPRM genes are present in all main groups of vertebrates. Thus, RPRM and RPRML genes were retained in all main groups of jawed vertebrates (gnathostomes), whereas RPRM3 was only retained in four distantly related groups: cartilaginous fish (e.g., sharks, skates, and rays), holostean fish (e.g., bowfish and gars), teleost fish (e.g., zebrafish) and coelacanths. In the group that includes lampreys and hagfish, two RPRM genes have been identified, however, phylogenetic and synteny analyses have failed in defining orthology. (B) Schematic representation of the evolution of the RPRM gene family in teleost fish. On the left, the RPRM gene family diversified as a product of the two rounds of whole-genome duplication occurred in the vertebrate ancestor, as well as, a teleost-specific whole genome duplication. On the left, the expression territories of the three RPRM gene lineages present in teleost fish.

Figure 3

RNA expression of RPRM and RPRML across different tissues. (A) Tissue-specific expression profile from 570 human donors available in the Genotype-Tissue Expression (GTEx) database [28]. Data is expressed as log10 of Transcripts Per Kilobase Million (TPM). (B) Expression levels of human tumor and matched normal tissue samples from Broad Institute TCGA Genome Data Analysis Center [30]. Data is expressed as log2 of RSEM (RNA-Seq by Expectation Maximization).

Figure 4

Schematic model of RPRM-mediated cell cycle and G2 arrest mechanisms. RPRM has been identified as a transcriptional target for: (1) p53 [4]; (2) histone deacetylase 7/FoxA1 (HDAC7/FoxA1) in an estrogen mediated mechanism [49]; and (3) for epigenetic silencing by hypermethylation of its promoter region [54]. A potential regulation by p73 it has also been proposed [3]. RPRM expression results in inhibited dephosphorylation of Cdc2, suppressing the activation of the Cdc2-Cyclin B1 complex. Thus, inducing cell cycle arrest at G2 suggesting a potential role for RPRM as a tumor suppressor gene [4]. The balance towards cell cycle arrest or proliferation can be shifted by multiple antagonistic effectors, amongst them RPRM. Straight lines with arrowheads indicate activation. Lines with no arrowhead indicate inhibition. Curved arrow on the bottom left indicates dephosphorylation of Cdc2/Cyclin B1 complex. Curved thick arrow on the bottom right indicates nuclear translocation of dephosphorylated Cdc2/Cyclin B1 at the G2/M checkpoint.

Figure 5

Unanswered questions in RPRM gene family. RPRM is as a p53-induced protein which induces cell cycle arrest at the G2/M checkpoint [4], through an unknown mechanism. Recently, RPRM genes have been shown to be expressed during brain, gut and blood vessel development [2]. Additional functions for RPRM include its role as a potential tumor suppressor gene (TSG), and a biomarker—through the assessment of the methylation status of its promoter region—for non-invasive detection of gastric cancer and other tumors [48,62]. Much like RPRM, RPRML is also expressed during embryonic development [2], but its role in physiological processes has never been investigated.