Decreased phosphorylation of a low molecular weight protein by cGMP-dependent protein kinase in variant HL-60 cells resistant to nitric oxide- and cGMP-induced differentiation.
We previously described the isolation of a variant subline of HL-60 cells that does not differentiate in response to nitric oxide (NO)-generating agents or to cGMP analogs. The variant cells have normal guanylate cyclase activity and normal NO-induced increases in the intracellular cGMP concentration. We now show that the variant cells have normal cGMP-dependent protein kinase (G-kinase) activity, both by an in vitro and in vivo assay, and using two-dimensional gel electrophoresis we have identified six G-kinase substrates in the parental cells. Of these six proteins, we found considerably less phosphorylation of one of the proteins in the variant cells than in parental cells, both in vitro and in intact cells, and by 35S-methionine/35S-cysteine incorporation we found much less of this protein in the variant cells than in parental cells. The protein is a shared substrate of cAMP-dependent protein kinase (A-kinase); since cAMP analogs still induce differentiation of the variant cells, it appears that the NO/cGMP/G-kinase and cAMP/A-kinase signal transduction pathways share some but not all of the same target proteins in inducing differentiation of HL-60 cells.
Scheele
J S
JS
Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla 92093-0652, USA.
Pilz
R B
RB
Clark
G
G
Gupta
N
N
Loo
D
D
Martis
P
P
Boss
G R
GR
D002454:Cell Differentiation / Q000187:drug effects*; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000187:drug effects / Q000378:metabolism; D006152:Cyclic GMP / Q000494:pharmacology*; D017869:Cyclic GMP-Dependent Protein Kinases / Q000187:drug effects / Q000378:metabolism*; D003545:Cysteine / Q000032:analysis; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D018922:HL-60 Cells / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D006801:Humans; D008715:Methionine / Q000032:analysis; D009569:Nitric Oxide / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000187:drug effects / Q000378:metabolism; D011506:Proteins / Q000032:analysis / Q000378:metabolism*; D013462:Sulfur Radioisotopes / Q000032:analysis
D011506:Proteins; D013462:Sulfur Radioisotopes; D009569:Nitric Oxide; D008715:Methionine; D017868:Cyclic AMP-Dependent Protein Kinases; D017869:Cyclic GMP-Dependent Protein Kinases; D011493:Protein Kinase C; D006152:Cyclic GMP; D003545:Cysteine