engleski

Characterization of Nme6-like gene/protein from marine sponge Suberites domuncula

Nucleoside diphosphate kinases (NDPKs) are evolutionarily conserved enzymes involved in many biological processes such as metastasis, proliferation, development, differentiation, ciliary functions, vesicle transport and apoptosis in vertebrates. Biochemical mechanisms of these processes are still largely unknown. Sponges (Porifera) are simple metazoans without tissues, closest to the common ancestor of all animals. They changed little during evolution and probably provide the best insight into the metazoan ancestors’ genomic features. The purpos of this study was to address structural and functional properties of Group II Nme6 gene/protein homolog from the marine sponge Suberites domuncula, Nme6Sd, in order to elucidate its evolutionary history. Nme6Sd gene and promoter were sequenced and analysed with various bioinformatical tools. Nme6Sd and Nme6SdΔ31 were produced in E. coli strain BL21 and NDPK activity was measured using a coupled pyruvate kinase-lactate dehydrogenase assay. Subcellular localization in human tumour cells was examined by confocal scanning microscopy. Our results show that the sponge Nme6Sd compared to human Nme6 does not possess NDPK activity, does not localize in mitochondria at least in human cells although it has a mitochondrial signal sequence, lacks two recent introns that comprise miRNAs, and have different transcriptional binding sites in the promoter region. Therefore, we conclude that the structure and function of Nme6 gene/protein changed during metazoan evolution and that complex actions of Nme6 in different biological processes probably correlate with increasing complexity of the organism.

Porifera; NDPK; Nm23; Nme6; gene structure; subcellular localization

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