Cell growth stimulation by EGF: inhibition through antisense-oligodeoxynucleotides demonstrates important role of casein kinase II.
Casein kinase II (CKII) is a highly conserved ubiquitous serine/threonine kinase composed of two catalytically active subunits (alpha and/or alpha') and two presumably regulatory subunits (beta). CKII has numerous cellular functions including a possible role in mitogenic signaling. To address this question, growth-arrested primary human fibroblasts (IMR-90) were exposed prior growth stimulation by epidermal growth factor (EGF) to oligodeoxynucleotides complementary to the translation start region of mRNAs coding for CKII alpha and beta subunits. A significant inhibition of growth stimulation (up to 60%) was observed with both antisense-alpha and antisense-beta. The inhibition was reversible, became decreased with mutated antisense-oligodeoxynucleotides, and neutralized by simultaneous presence of respective sense-oligodeoxynucleotides. The expected down-regulation of CKII protein due to hybrid formation of antisense-oligodeoxynucleotides with target mRNAs was investigated by determination of the intracellular protein level of CKII beta-subunit by immunofluorescence and quantitative image analysis. The protein was revealed to be localized predominantly in the nucleus and to become significantly decreased due to antisense-beta treatment of cells. The maximum decrease coincided with the early phase (first several hours) of growth stimulation by EGF when antisense-beta incubation was started 6-2 h before growth stimulation, the period within which application of antisense-alpha and antisense-beta caused the maximum of inhibition of growth stimulation. Thus CKII obviously plays, with both subunit alpha and subunit beta, an important role in the early phase of mitogenic stimulation.
Pepperkok
R
R
Biochemical Cell Physiology, German Cancer Research Center, Heidelberg.
Lorenz
P
P
Jakobi
R
R
Ansorge
W
W
Pyerin
W
W
D001483:Base Sequence; D047388:Casein Kinases; D002455:Cell Division / Q000187:drug effects*; D002460:Cell Line; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D006801:Humans; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000494:pharmacology*; D016376:Oligonucleotides, Antisense / Q000138:chemical synthesis / Q000494:pharmacology*; D011494:Protein Kinases / Q000235:genetics* / Q000302:isolation & purification / Q000502:physiology; D012333:RNA, Messenger / Q000187:drug effects / Q000235:genetics
D046911:Macromolecular Substances; D009838:Oligodeoxyribonucleotides; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D004815:Epidermal Growth Factor; D011494:Protein Kinases; D047388:Casein Kinases