g to e clude the phospho regulation of I2 at this site. These data are in agreement with the recent P. falciparum phosphoproteome characterization showing the phosphorylation of PfI2 at positions T13, S48, S50, S115, T117 and S142, but not at T39 within the P TP motif. The assessment of the impact of PfI2 phosphorylation selleck compound will await further investigations on these phosphorylated residues as well as the T within the P TP motif. At this stage, it is im portant to mention Inhibitors,Modulators,Libraries that, beside the capacity to interact with PP1c, human I2 has been shown to participate in a direct kinase dependent signaling network. It was found that I2 was able to bind and to activate Nek2 and Aurora A kinases. For these functions, I2 seems to operate through its C terminal domain as the protein deleted in this domain failed to interact with these kinases, e cluding a role for the KGILK and RV F motifs.
Although the PfI2 sequence is 61 amino acids shorter than its human homologue, the capacity of PfI2 to bind P. falciparum kinases of the NIMA and Aurora families Inhibitors,Modulators,Libraries should be evaluated. In P. falciparum, microarray analysis detected PfI2 mRNA in all blood parasite stages and gametocytes. In this work, co immunoprecipitation e periments with anti PfI2 anti bodies followed by Western blotting and the use of a PfPP1 affinity column clearly revealed the e pression of PfI2 protein by P. falciparum and of its capacity to bind PfPP1. Transfection of live parasites with the tagged PfI2 GFP protein showed that its distribution is nucleocytoplasmic, like PfPP1, with a strong accu mulation in the nucleus, is in agreement with the localization of other I2 proteins.
Indeed mamma lian I2 fused to GFP was localized in both the cytoplasm and the nucleus, with an active import to the latter compartment, supported by the presence of two puta tive nuclear localization Inhibitors,Modulators,Libraries signals. In the case of PfI2, bioinformatics analysis also revealed a putative nuclear localization signal, supporting its nuclear localization. We previously reported that PfLRR1 and Pf inhibitor 3, the first identified regulatory subunits of PfPP1c, localized to the nucleus, evoking a specific role in this compartment. The present study suggests an additional role for the PfI2 regulatory subunit of PP1c, present in the nucleus but also in the cytoplasm.
Our reverse genetic studies strongly suggest a critical role for PfI2 in the erythrocytic ase ual cycle in vitro Inhibitors,Modulators,Libraries as no parasites with a disrupted PfI2 gene were detectable. Definition of the PfI2 role during the life cycle neces sitates Brefeldin_A further work, requiring the development of a powerful inducible e pression system for P. falciparum. The ability of PfI2 to bind and to inhibit PP1c both in vitro and in cellular conditions through the two main motifs the RV F motif and the HYNE motif, together with the fact that a tight and appropriate regulation of PP1c is crucial for Ixazomib purchase cellular functions, led us to e plore whether derived competing peptides from PfI2 could bind to PP1c and inhibit dow