Protein kinase C-mediated down-regulation of cyclin D1 involves activation of the translational repressor 4E-BP1 via a phosphoinositide 3-kinase/Akt-independent, protein phosphatase 2A-dependent mechanism in intestinal epithelial cells.
We reported previously that protein kinase Calpha (PKCalpha), a negative regulator of cell growth in the intestinal epithelium, inhibits cyclin D1 translation by inducing hypophosphorylation/activation of the translational repressor 4E-BP1. The current study explores the molecular mechanisms underlying PKC/PKCalpha-induced activation of 4E-BP1 in IEC-18 nontransformed rat ileal crypt cells. PKC signaling is shown to promote dephosphorylation of Thr(45) and Ser(64) on 4E-BP1, residues directly involved in its association with eIF4E. Consistent with the known role of the phosphoinositide 3-kinase (PI3K)/Akt/mTOR pathway in regulation of 4E-BP1, PKC signaling transiently inhibited PI3K activity and Akt phosphorylation in IEC-18 cells. However, PKC/PKCalpha-induced activation of 4E-BP1 was not prevented by constitutively active mutants of PI3K or Akt, indicating that blockade of PI3K/Akt signaling is not the primary effector of 4E-BP1 activation. This idea is supported by the fact that PKC activation did not alter S6 kinase activity in these cells. Further analysis indicated that PKC-mediated 4E-BP1 hypophosphorylation is dependent on the activity of protein phosphatase 2A (PP2A). PKC signaling induced an approximately 2-fold increase in PP2A activity, and phosphatase inhibition blocked the effects of PKC agonists on 4E-BP1 phosphorylation and cyclin D1 expression. H(2)O(2) and ceramide, two naturally occurring PKCalpha agonists that promote growth arrest in intestinal cells, activate 4E-BP1 in PKC/PKCalpha-dependent manner, supporting the physiological significance of the findings. Together, our studies indicate that activation of PP2A is an important mechanism underlying PKC/PKCalpha-induced inhibition of cap-dependent translation and growth suppression in intestinal epithelial cells.
Guan
Lingjie
L
Department of Pharmacology and Therapeutics, Roswell Park Cancer Institute, Buffalo, NY 14263, USA.
Song
Kyung
K
Pysz
Marybeth A
MA
Curry
Kathryn J
KJ
Hizli
A Asli
AA
Danielpour
David
D
Black
Adrian R
AR
Black
Jennifer D
JD
D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D056741:Cyclin D; D016213:Cyclins / Q000378:metabolism*; D015536:Down-Regulation; D004847:Epithelial Cells / Q000378:metabolism; D039642:Eukaryotic Initiation Factors / Q000378:metabolism; D007413:Intestinal Mucosa / Q000378:metabolism*; D047908:Intracellular Signaling Peptides and Proteins; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D051571:Protein Kinase C-alpha / Q000378:metabolism*; D054648:Protein Phosphatase 2; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D051381:Rats; D015398:Signal Transduction
D002352:Carrier Proteins; D056741:Cyclin D; D016213:Cyclins; D039642:Eukaryotic Initiation Factors; D047908:Intracellular Signaling Peptides and Proteins; D019869:Phosphatidylinositol 3-Kinases; D010749:Phosphoprotein Phosphatases; D010750:Phosphoproteins; D051571:Protein Kinase C-alpha; D054648:Protein Phosphatase 2; D051057:Proto-Oncogene Proteins c-akt; C499526:Eif4ebp1 protein, rat