Structure and function of cancer-related developmentally regulated GTP-binding protein 1 (DRG1) is conserved between sponges and humans

Abstract

Cancer is a disease caused by errors within the multicellular system and it represents a major health issue in multicellular organisms. Although cancer research has advanced substantially, new approaches focusing on fundamental aspects of cancer origin and mechanisms of spreading are necessary. Comparative genomic studies have shown that most genes linked to human cancer emerged during the early evolution of Metazoa. Thus, basal animals without true tissues and organs, such as sponges (Porifera), might be an innovative model system for understanding the molecular mechanisms of proteins involved in cancer biology. One of these proteins is developmentally regulated GTP-binding protein 1 (DRG1), a GTPase stabilized by interaction with DRG family regulatory protein 1 (DFRP1). This study reveals a high evolutionary conservation of DRG1 gene/protein in metazoans. Our biochemical analysis and structural predictions show that both recombinant sponge and human DRG1 are predominantly monomers that form complexes with DFRP1 and bind non-specifically to RNA and DNA. We demonstrate the conservation of sponge and human DRG1 biological features, including intracellular localization and DRG1:DFRP1 binding, function of DRG1 in α-tubulin dynamics, and its role in cancer biology demonstrated by increased proliferation, migration and colonization in human cancer cells. These results suggest that the ancestor of all Metazoa already possessed DRG1 that is structurally and functionally similar to the human DRG1, even before the development of real tissues or tumors, indicating an important function of DRG1 in fundamental cellular pathways.

Transfection of Sponge Cells and Intracellular Localization of Cancer-Related MYC, RRAS2, and DRG1 Proteins

“…DRG1 is an evolutionary conserved GTPase that has been implicated in various tumors, but its role in cancer is not yet fully understood. Our study [ 73 ], as well as other studies, confirmed the localization of DRG1 in the cytosol of various cell lines (MCF-7 and HeLa human cancer cell lines, mouse 3T3 cells, and Drosophila melanogaster cells) [ 94 , 95 , 96 ].…”

“…The developmentally regulated GTP-binding protein (DRG) subfamily consists of two paralogs, DRG1 and DRG2. Both are involved in protein translation [ 91 ], microtubule regulation [ 92 ], and cell proliferation [ 73 , 93 ]. DRG1 is an evolutionary conserved GTPase that has been implicated in various tumors, but its role in cancer is not yet fully understood.…”

“…The expression of the human DRG1 protein requires the DFRP1 protein for its stabilization and localization in the cytosol of human cancer cells MCF-7 and HeLa [ 73 , 92 ]. Therefore, to determine the localization of the DRG1 protein, cDNAs for the sponge and human proteins were cloned into a vector containing the GFP marker, while the sponge and human DFRP1 were cloned into a vector containing the CHERRY reporter.…”

“…The resulting constructs for the analysis of MYC and RRAS2 intracellular localization are GFP- or CHERRY-tagged. Other plasmids (EsuDRG1-GFP, HsaDRG1-GFP, EsuDFRP1-CHERRY, HsaDFRP1-CHERRY, EsuDFRP1-MYC, and HsaDFRP1-MYC) were already published [ 73 ].…”

“…Previous research has shown that these proteins display a high similarity to homologs in “higher” metazoans, not only in primary but also in predicted secondary and tertiary structures, suggesting similar or identical biochemical and biological functions [ 59 , 63 , 64 , 65 , 66 , 67 , 68 , 69 ]. Although we have some insights into the biological function of sponge proteins, the evolutionary conservation of their function in sponges and humans is still poorly understood [ 70 , 71 , 72 , 73 ]. Due to the compartmentalization of the eukaryotic cell, the determination of the protein’s intracellular localization is the first step for the understanding of its activation state, interaction partners, correct chemical environment (e.g.…”