Protein phosphorylation in encystment-induced Colpoda cucullus: localization and identification of phosphoproteins

In Colpoda cucullus, the morphogenetic transformation was preceded by an enhancement of the in vivo protein phosphorylation level. Immunofluorescence microscopy using antiphosphoserine antibody showed that these phosphorylated proteins were localized in the macronucleus and other cytoplasmic regions. Biotinylated Phos-tag/ECL assays of isolated macronuclei showed that a 33-kDa protein (p33) was localized within them. The p33 obtained from isolated macronuclei was tentatively identified as ribosomal P0 protein by LC-MS/MS analysis. In addition, among the encystment-specific phosphoproteins obtained by phosphate-affinity chromatography, the 29-, 31-, and 33-kDa proteins (p29, p31, and p33) were tentatively identified as ribosomal P0 protein, whereas the 24-kDa phosphoprotein (p24) was tentatively identified as ribosomal S5 protein.