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Vol. 10, Issue 12, 4355-4367, December 1999
¶ and
*Dipartimento di Istologia ed Embriologia Medica, Università
"La Sapienza," 00161 Rome, Italy; Myogenic cell differentiation is induced by
Arg8-vasopressin, whereas high cAMP levels and protein
kinase A (PKA) activity inhibit myogenesis. We investigated the role of
type 4 phosphodiesterase (PDE4) during L6-C5 myoblast differentiation.
Selective PDE4 inhibition resulted in suppression of differentiation
induced by vasopressin. PDE4 inhibition prevented vasopressin-induced
nuclear translocation of the muscle-specific transcription factor
myogenin without affecting its overall expression level. The effects of
PDE4 inhibition could be attributed to an increase of cAMP levels and
PKA activity. RNase protection, reverse transcriptase PCR,
immunoprecipitation, Western blot, and enzyme activity assays
demonstrated that the PDE4D3 isoform is the major PDE4 expressed in
L6-C5 myoblasts and myotubes, accounting for 75% of total
cAMP-hydrolyzing activity. Vasopressin cell stimulation caused a
biphasic increase of PDE4 activity, which peaked at 2 and 15 min and
remained elevated for 48 h. In the continuous presence of
vasopressin, cAMP levels and PKA activity were lowered. PDE4D3
overexpression increased spontaneous and vasopressin-dependent
differentiation of L6-C5 cells. These results show that PDE4D3 plays a
key role in the control of cAMP levels and differentiation of L6-C5
cells. Through the modulation of PDE4 activity, vasopressin inhibits
the cAMP signal transduction pathway, which regulates myogenesis
possibly by controlling the subcellular localization of myogenin.
Dipartimento di
Sanità Pubblica e Biologia Cellulare, Università "Tor
Vergata," 00133 Rome, Italy; Unité 352, Institut National de la Santé et de la Recherche Médicale,
Biochimie et Pharmacologie, Institut National des Sciences
Appliquées de Lyon, 69621 Villeurbanne, France; and
§Division of Reproductive Biology, Department of
Gynecology and Obstetrics, Stanford University School of Medicine,
Stanford, California 94305
Joint senior authors.
¶
Corresponding author. E-mail address:
adamo{at}axrma.uniroma1.it.
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