Induction of megakaryocytic differentiation and modulation of protein kinase gene expression by site-selective cAMP analogs in K-562 human leukemic cells.
Two classes (site 1- and site 2-selective) of cAMP analogs, which either alone or in combination demonstrate a preference for binding to type II rather than type I cAMP-dependent protein kinase isozyme, potently inhibit growth in a spectrum of human cancer cell lines in culture. Treatment of K-562 human leukemic cells for 3 days with 30 and 10 microM 8-chloroadenosine 3',5'-cyclic monophosphate (8-Cl-cAMP) (site 1-selective) resulted in 60% and 20% growth inhibition, respectively (with over 90% viability). N6-Benzyl-cAMP (site 2-selective) (30 microM) treatment resulted in 20% growth inhibition by day 3. When 8-Cl-cAMP (10 microM) and N6-benzyl-cAMP (30 microM) were both added, growth was almost completely arrested. The growth inhibition was accompanied by megakaryocytic differentiation in K-562 cells. The untreated control cells expressed little or no detectable levels of glycoprotein IIb-IIIa surface antigen complex. 8-Cl-cAMP (30 microM) treatment for 3 days substantially increased the antigen expression, while N6-benzyl-cAMP caused little or no change in the antigen expression. When cells were treated with 8-Cl-cAMP in combination with N6-benzyl-cAMP, antigen expression was synergistically enhanced, and cells demonstrated megakaryocyte morphology. By Northern blotting, we examined the mRNA levels of the type I and type II protein kinase regulatory subunits (RI alpha and RII beta), the catalytic subunit, and c-myc during 8-Cl-cAMP treatment. The steady-state level of RII beta cAMP receptor mRNA sharply increased within 1 hr of treatment and remained elevated for 3 days, while that of the RI alpha receptor markedly decreased to below control level within 6 hr and remained low during treatment. However, 8-Cl-cAMP did not affect the mRNA level of the catalytic subunit. 8-Cl-cAMP treatment also brought about a rapid decrease in c-myc mRNA. Thus, differential regulation of cAMP receptor genes is an early event in cAMP-induced differentiation and growth control of K-562 leukemia cells.
Tortora
G
G
Cellular Biochemistry Section, National Cancer Institute, Bethesda, MD 20892.
Clair
T
T
Katsaros
D
D
Ally
S
S
Colamonici
O
O
Neckers
L M
LM
Tagliaferri
P
P
Jahnsen
T
T
Robins
R K
RK
Cho-Chung
Y S
YS
D000954:Antigens, Surface / Q000032:analysis; D015152:Blotting, Northern; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D000242:Cyclic AMP / Q000031:analogs & derivatives* / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D008533:Megakaryocytes / Q000166:cytology*; D011494:Protein Kinases / Q000235:genetics*; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger / Q000235:genetics
D000954:Antigens, Surface; D011518:Proto-Oncogene Proteins; D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger; D000242:Cyclic AMP; D011494:Protein Kinases