Yeast Gcn4p stabilization is initiated by the dissociation of the nuclear Pho85p/Pcl5p complex.

Protein stability of the c-jun-like yeast bZIP transcriptional activator Gcn4p is exclusively controlled in the yeast nucleus. Phosphorylation by the nuclear Pho85p cyclin-dependent protein kinase, a functional homolog of mammalian Cdk5, initiates the Gcn4p degradation pathway in complex with the cyclin Pcl5p. We show that the initial step in Gcn4p stabilization is the dissociation of the Pho85p/Pcl5p complex. Pcl7p, another nuclear and constantly present cyclin, is required for Gcn4p stabilization and is able to associate to Pho85p independently of the activity of the Gcn4p degradation pathway. In addition, the nuclear cyclin-dependent Pho85p kinase inhibitor Pho81p is required for Gcn4p stabilization. Pho81p only interacts with Pcl5p when Gcn4p is rapidly degraded but constitutively interacts with Pcl7p. Our data suggest that Pcl7p and Pho81p are antagonists of the Pho85p/Pcl5p complex formation in a yet unknown way, which are specifically required for Gcn4p stabilization. We suggest that dissociation of the Pho85p/Pcl5p complex as initial step in Gcn4p stabilization is a prerequisite for a shift of equilibrium to an increased amount of the Pho85p/Pcl7p complexes and subsequently results in decreased Gcn4p phosphorylation and therefore increased stability of the transcription factor.

A polyphenol rich plant extract, CYSTUS052, exerts anti influenza virus activity in cell culture without toxic side effects or the tendency to induce viral resistance.

Infections with influenza A viruses still pose a major threat to humans and several animal species. The occurrence of highly pathogenic avian influenza viruses of the H5N1 subtype capable to infect and kill humans highlights the urgent need for new and efficient countermeasures against this viral disease. Here we demonstrate that a polyphenol rich extract (CYSTUS052) from the Mediterranean plant Cistus incanus exerts a potent anti-influenza virus activity in A549 or MDCK cell cultures infected with prototype avian and human influenza strains of different subtypes. CYSTUS052 treatment resulted in a reduction of progeny virus titers of up to two logs. At the effective dose of 50 microg/ml the extract did not exhibit apparent harming effects on cell viability, metabolism or proliferation, which is consistent with the fact that these plant extracts are already used in traditional medicine in southern Europe for centuries without any reported complications. Viruses did not develop resistance to CYSTUS052 when compared to amantadine that resulted in the generation of resistant variants after only a few passages. On a molecular basis the protective effect of CYSTUS052 appears to be mainly due to binding of the polymeric polyphenol components of the extract to the virus surface, thereby inhibiting binding of the hemagglutinin to cellular receptors. Thus, a local application of CYSTUS052 at the viral entry routes may be a promising approach that may help to protect from influenza virus infections.