title: Structural insight into an evolutionarily ancient programmed cell death regulator - the crystal structure of marine sponge BHP2 bound to LB-Bak-2.
abstract: Sponges of the porifera family harbor some of the evolutionary most ancient orthologs of the B-cell lymphoma-2 (Bcl-2) family, a protein family critical to regulation of apoptosis. The genome of the sponge Geodia cydonium contains the putative pro-survival Bcl-2 homolog BHP2, which protects sponge tissue as well as mammalian Hek-293 and NIH-3T3 cells against diverse apoptotic stimuli. The Lake Baikal demosponge Lubomirskia baicalensis has been shown to encode both putative pro-survival Bcl-2 (LB-Bcl-2) and pro-apoptotic Bcl-2 members (LB-Bak-2), which have been implied in axis formation (branches) in L. baicalensis. However, the molecular mechanism of action of sponge-encoded orthologs of Bcl-2 remains to be clarified. Here, we report that the pro-survival Bcl-2 ortholog BHP2 from G. cydonium is able to bind the BH3 motif of a pro-apoptotic Bcl-2 protein, LB-Bak-2 of the sponge L. baicalensis. Furthermore, we determined the crystal structure of BHP2 bound to LB-Bak-2, which revealed that using a binding groove conserved across all pro-survival Bcl-2 proteins, BHP2 binds multi-motif Bax-like proteins through their BH3-binding regions. However, BHP2 discriminates against BH3-only bearing proteins by blocking access to a hydrophobic pocket that is critical for BH3 motif binding in pro-survival Bcl-2 proteins from higher organisms. This differential binding mode is reflected in a structure-based phylogenetic comparison of BHP2 with other Bcl-2 family members, which revealed that BHP2 does not cluster with either Bcl-2 members of higher organisms or pathogen-encoded homologs, and assumes a discrete position. Our findings suggest that the molecular machinery and mechanisms for executing Bcl-2-mediated apoptosis as observed in mammals are evolutionary ancient, with early regulation of apoptotic machineries closely resembling their modern counterparts in mammals rather than Caenorhabditis elegans or drosophila.
authors.affiliation: Department of Biochemistry & Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne,Victoria 3086, Australia.
authors.forename: Sofia
authors.initials: S
authors.lastname: Caria
authors.affiliation: Department of Chemistry & Physics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, Victoria 3086, Australia.
authors.forename: Mark G
authors.initials: MG
authors.lastname: Hinds
authors.affiliation: Department of Biochemistry & Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne,Victoria 3086, Australia.
authors.forename: Marc
authors.initials: M
authors.lastname: Kvansakul
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000235:genetics*; D051017:Apoptosis Regulatory Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D018360:Crystallography, X-Ray; D019143:Evolution, Molecular; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D041681:NIH 3T3 Cells; D011485:Protein Binding; D011487:Protein Conformation; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D051017:Apoptosis Regulatory Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; C437969:bhp2 protein, Geodia cydonium