Detection and developmental regulation of the mRNA for the regulatory subunit of the cAMP-dependent protein kinase of D. discoideum by cell-free translation.
CAMP is an important effector of the development of Dictyostelium discoideum amoebae and could exert its effects on gene expression through the cytosolic cAMP‐dependent protein kinase (cAK). Antibodies, specific for the regulatory subunit (R) of the cAK, were used to investigate the developmental regulation of the corresponding mRNA (R‐mRNA) by in vitro translation and immunoprecipitation. Under such conditions, a single polypeptide of the same mol. wt. as R (42 kd) is detected, showing that the protein is not synthesized as a large precursor. The level of the R‐mRNA, which is low in vegetative cells, increases 10‐ to 20‐fold during the first hours of development. Its expression is stimulated by the treatment of AX3 cells with cAMP either added to a concentration of 1 mM or given as 0.1 microM pulses every 5 min, whereas such treatments have little or no effect in cells of strain AX2. The R‐mRNA remains highly expressed (0.01‐0.03% of translatable mRNA) throughout post‐aggregative development; it is not affected by mechanical disaggregation of the multicellular organism. The parallel developmental time courses of the translatable R‐mRNA and the R protein produced in vivo suggest that the expression of this polypeptide is regulated at the level of mRNA synthesis.