4E-binding protein phosphorylation and eukaryotic initiation factor-4E release are required for airway smooth muscle hypertrophy.
The molecular mechanisms of airway smooth muscle hypertrophy, a feature of severe asthma, are poorly understood. We previously established a conditionally immortalized human bronchial smooth muscle cell line with a temperature-sensitive SV40 large T antigen. Temperature shift and loss of large T cause G1-phase cell cycle arrest that is accompanied by increased airway smooth muscle cell size. In the present study, we hypothesized that phosphorylation of eukaryotic initiation factor-4E (eIF4E)-binding protein (4E-BP), which subsequently releases eIF4E and initiates cap-dependent mRNA translation, was required for airway smooth muscle hypertrophy. Treatment of cells with chemical inhibitors of PI 3-kinase and mammalian target of rapamycin blocked protein synthesis and cell growth while decreasing the phosphorylation of 4E-BP and increasing the binding of 4E-BP to eIF4E, consistent with the notion that 4E-BP1 phosphorylation and eIF4E function are required for hypertrophy. To test this directly, we infected cells with a retrovirus encoding a phosphorylation site mutant of 4E-BP1 (AA-4E-BP-1) that dominantly inhibits eIF4E. Upon temperature shift, cells infected with AA-4E-BP-1, but not empty vector, failed to undergo hypertrophic growth. We conclude that phosphorylation of 4E-BP, eIF4E release, and cap-dependent protein synthesis are required for hypertrophy of human airway smooth muscle cells.
Zhou
Limei
L
Department of Pediatrics and Communicable Diseases, University of Michigan, 1150 W. Medical Center Dr., Room 3570, MSRBII, Box 0688, Ann Arbor, MI 48109-0688, USA.
Goldsmith
Adam M
AM
Bentley
J Kelley
JK
Jia
Yue
Y
Rodriguez
Michael L
ML
Abe
Mark K
MK
Fingar
Diane C
DC
Hershenson
Marc B
MB
D048868:Adaptor Proteins, Signal Transducing; D001980:Bronchi / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology*; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D018797:Cell Cycle Proteins; D049108:Cell Enlargement / Q000187:drug effects; D002460:Cell Line; D002867:Chromones / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D039561:Eukaryotic Initiation Factor-4E / Q000378:metabolism*; D006801:Humans; D006984:Hypertrophy; D007093:Imidazoles / Q000494:pharmacology; D009025:Morpholines / Q000494:pharmacology; D009130:Muscle, Smooth / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology*; D009154:Mutation; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000187:drug effects; D011725:Pyridines / Q000494:pharmacology; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases; D013696:Temperature; D014162:Transfection; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002867:Chromones; C087000:EIF4EBP1 protein, human; D004791:Enzyme Inhibitors; D039561:Eukaryotic Initiation Factor-4E; D007093:Imidazoles; D009025:Morpholines; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010750:Phosphoproteins; D011725:Pyridines; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D011494:Protein Kinases; C546842:MTOR protein, human; D058570:TOR Serine-Threonine Kinases; D048051:p38 Mitogen-Activated Protein Kinases; C090942:4-(4-fluorophenyl)-2-(4-hydroxyphenyl)-5-(4-pyridyl)imidazole; D020123:Sirolimus