Selective nuclear translocation of protein kinase C alpha in Swiss 3T3 cells treated with IGF-I, PDGF and EGF.
To determine the subcellular distribution of PKC after GFs treatment we have employed a combined immunochemical and in situ confocal microscopy analysis. In quiescent Swiss 3T3 cells only a faint PKC positivity was observable in the nucleus while a strong reaction was seen in the cytoplasm. IGF-I and to a lesser extent PDGF and EGF induced, after 45 min of treatment, a nuclear translocation of PKC detected by a pan-anti-PKC antibody and nuclear fluorescence was distributed in the nuclear interior except for the nucleolar regions. Bombesin and FGF did not affect the sub-cellular distribution of the enzyme. To further the understanding of which PKC isoform was involved in the translocation process, we have tested nine isozyme-specific anti-PKC antibodies. Immunoblotting analysis revealed the presence in Swiss 3T3 fibroblasts of alpha, beta I, epsilon and zeta isoforms. In isolated nuclei from GF-exposed cells only the alpha isozyme was detected: immunostaining was very intense after IGF-I treatment and clearly observable after PDGF and EGF stimulation. This result was strongly supported by the in situ confocal microscopy which parallels the Western blot analysis. These data demonstrate that several, but not all, GFs acting through tyrosine kinase receptor induce the intranuclear translocation of PKC alpha and, because of the dramatic effect of IGF-I, strengthen the case for a link between the activation of nuclear inositol lipid cycle and PKC translocation induced by this GF.
Istituto di Anatomia Umana Normale, Università di Ferrara, Italy.
L M
LM
Neri
A M
AM
Billi
L
L
Manzoli
S
S
Rubbini
R S
RS
Gilmour
L
L
Cocco
A M
AM
Martelli
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002451:Cell Compartmentation; D002467:Cell Nucleus / Q000378:metabolism*; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005455:Fluorescent Antibody Technique; D006133:Growth Substances / Q000494:pharmacology*; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D007527:Isoenzymes / Q000378:metabolism*; D051379:Mice; D008853:Microscopy / Q000379:methods; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000302:isolation & purification; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D011493:Protein Kinase C / Q000302:isolation & purification / Q000378:metabolism*
D006133:Growth Substances; D007527:Isoenzymes; D009687:Nuclear Proteins; D010982:Platelet-Derived Growth Factor; D004815:Epidermal Growth Factor; D007334:Insulin-Like Growth Factor I; D011493:Protein Kinase C