Yeast surviving factor Svf1 as a new interacting partner, regulator and in vitro substrate of protein kinase CK2

Abstract

Since Svf1 is phosphoprotein, we investigated whether it was a substrate for protein kinase CK2. According to the amino acid sequence Svf1 harbours 20 putative CK2 phosphorylation sites. Here, we have reported cloning, overexpression, purification and characterization of yeast Svf1 as a substrate for three forms of yeast CK2. Svf1 serves as a substrate for both the recombinant CK2alpha (Km 0.35 microM) and CK2alpha' (Km 0.18 microM) as well as CK2 holoenzyme (Km 1.1 microM). Different Km values argue that CK2beta(beta') subunit has an inhibitory effect on the activity of both CK2alpha and CK2alpha' towards surviving factor Svf1. Reconstitution of alpha'2betabeta' isoform of CK2 holoenzyme shows that beta/beta' subunits have regulatory effect depending on the kind of CK2 catalytic subunit. This effect was not observed in the case of alpha2betabeta' isoform, which may be due to interaction between Svf1 and regulatory CK2beta subunit (shown by co-immunoprecipitation experiments). Interactions between CK2 subunits and Svf1 protein may have influence on ATP as well as ATP-competitive inhibitors (TBBt and TBBz) binding. CK2 phosphorylates up to six serine residues in highly acidic peptide K199EVIPESDEEESSADEDDNEDEDEESGDSEEESGSEEESDSEEVEITYED248 of the Svf1 protein in vitro. Presented data may help to elucidate the role of protein kinase CK2 and Svf1 in the regulation of cell survival pathways.

Yeast Protein Asf1 Possesses Modulating Activity towards Protein Kinase CK2

“…As already described in our former studies [ 44 , 45 , 46 , 47 ], synthetic and natural inhibitors have different inhibitory potential depending on the protein substrate. We have investigated the inhibitory action of the Asf1 protein using already characterized substrates from yeast S. cerevisiae , such as recombinant Svf1, Fip1, and Elf1 [ 48 , 49 , 50 ]. The experiments using human catalytic subunits showed that the Asf1 protein exhibited an inhibitory effect towards the phosphorylation of different substrates.…”

Survival factor SvfA plays multiple roles in differentiation and is essential for completion of sexual development in Aspergillus nidulans

“…Activation of the expression of cpeA, a catalase-peroxidase gene, by StuA is also required for the formation of Hülle cells and cleistothecia during early sexual development 1,33 . It is also noted that Svf1 of S. cerevisiae is involved in cell survival by affecting the sphingolipid metabolism 40 and is a substrate of serine/threonine protein kinase CK2, which is essential for life in all eukaryotes by regulating the cell cycle, tumorigenesis, and apoptosis 41 . Interestingly, exogenous expression of the human anti-apoptotic gene Bcl-x L , which regulates apoptosis and the cell cycle 42 , could functionally complement the defect of Svf1 in S. cerevisiae 24 .…”