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Characterization of Developmentally Regulated cAMP/Ca 2+ ‐Independent Protein Kinases from Dictyostelium discoideum
Author(s) -
Pelech Steven,
Paddon Harry,
Kwong Linda,
Weeks Gerald
Publication year - 1989
Publication title -
development, growth and differentiation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.864
H-Index - 66
eISSN - 1440-169X
pISSN - 0012-1592
DOI - 10.1111/j.1440-169x.1989.00351.x
Subject(s) - dictyostelium discoideum , protein kinase a , kinase , phosphorylation , biochemistry , casein kinase 2 , casein kinase 1 , biology , casein , histone , mitogen activated protein kinase kinase , microbiology and biotechnology , dna , gene
Cell‐free extracts of the slime mold Dictyostelium discoideum were assayed for phosphorylating activity towards endogenous proteins and towards histone H1, casein and myelin basic protein (MBP). During development, protein kinase activity towards all of these substrates steadily increased and peaked between the aggregation and the pseudoplasmodial stages. Particulate‐associated kinase activity was solubilized with 1% CHAPS, and separated into 300–400 kDa and ∼ 100 kDa components on Sephacryl S‐300. The 300–400 kDa peak exhibited the most pronounced developmental increase in MBP phosphorylating activity. It was further fractionated on DEAE‐Sephacel and heparin‐Sepharose, and in each case, it coeluted with the histone H1 phosphorylating activity. The activity of this kinase was unaffected by cAMP and calmodulin, but it was reduced to 50% by ∼ 350 mM NaCl, 5 mM NaF and 40 μg polylysine/ml. The ∼ 100 kDa peak exhibited the most pronounced increase in casein kinase activity during development. Most of the casein phosphorylating activity did not bind to DEAE‐Sephacel; it was distinct from casein kinase 2, which was not developmentally regulated. In parallel with these elevated kinase activities during development, there was increased in vitro phosphorylation of a number of Dictyostelium proteins, including two major phosphoproteins of 140 and 94 kDa.