Differential functions of related family members Rybp and Yaf2 in cell survival

Abstract

Rybp and Yaf2 are the only members of their protein family and share 71% amino acid identity. Previously, these two proteins have been shown to interact with transcription factors including YY1 and members of the E2F, Myc, and Polycomb group families. Moreover, Rybp has been shown to interact with pro-apoptotic DED-containing proteins. Despite these known interactions, the differential cellular functions of Rybp and Yaf2 have not been fully elucidated.;Through two independent experimental methods, (i) studying the novel Rybp/Yaf2 interactions with the pro-apoptotic protein Hippi and (ii) Yaf2 morpholino knockdown in zebrafish, we have now demonstrated that Rybp and Yaf2 play different roles in cell survival. The Hippi protein was recovered from large-scale yeast two hybrid screens with Rybp as bait. Hippi is a pro-apoptotic protein that can complex with the Hip1 protein to activate Caspase 8-mediated apoptosis, possibly contributing to the loss of neurons seen in Huntington's disease. We demonstrated that Rybp could enhance both Hippi-mediated and Hip1/Hippi-mediated apoptosis. Furthermore, Rybp was required for Hippi to interact with Caspase 8, suggesting that Rybp may function as an adapter protein between Hip1/Hippi and Caspase 8. Importantly, Yaf2 can also interact with Hippi, but Yat2 inhibits Hippi-mediated apoptosis.;An anti-apoptotic role for Yat2 was also seen when expression of the zYaf2 ortholog was knocked down in the zebrafish. Embryos injected with either of two Yaf2 anti-sense morpholinos exhibited pre-somitic developmental arrest accompanied by significant apoptosis. The apoptosis was Caspase 8-mediated and was causal to developmental arrest because treating with a Caspase 8-specific inhibitor allowed the morphant embryos to survive early arrest. A hypomorphic phenotype emerged with lower doses of morpholino that included extensive degeneration of the anterior brain due to increased apoptosis.;Our findings suggest that Rybp and Yaf2 can encode opposing functions in the regulation of apoptosis, Rybp enhances apoptosis and Yaf2 inhibits apoptosis. However, the precise molecular roles of Rybp/Yaf2 as they relate to apoptosis/survival remain to be further elucidated. It is also important to determine how these roles in apoptosis/survival interrelate with Rybp/Yaf2's proposed activities in transcriptional regulation and posttranslational modification processes.