Modulation of insulin-like growth factor actions in L6A1 myoblasts by insulin-like growth factor binding protein (IGFBP)-4 and IGFBP-5: a dual role for IGFBP-5.
We have previously shown that the insulin-like growth factors (IGFs) stimulate both proliferation and differentiation of skeletal muscle cells in culture, and that these actions in L6A1 muscle cells may be modulated by three secreted IGF binding proteins (IGFBPs), IGFBP-4, -5, and -6. Since we found that the temporal expression pattern of IGFBP-4 and IGFBP-5 differed dramatically during the transition from proliferating myoblasts to differentiated myotubes, we undertook the current study to examine the effects of purified IGFBP-4 and IGFBP-5 on IGF-stimulated actions in L6A1 muscle cells. As has been shown for other cell types, we found that IGFBP-4 had only inhibitory actions, inhibiting IGF-I and IGF-II-stimulated proliferation and differentiation. In contrast, IGFBP-5 exhibited both inhibitory and stimulatory actions. When added in the presence of 30 ng/ml IGF-I, IGFBP-5 (250 ng/ml) inhibited all markers of the early proliferative response: the tyrosine phosphorylation of the cytoplasmic signaling molecules IRS-1 and Shc, the activation of the MAP kinases, ERK1 and 2, the elevation of c-fos mRNA, the early inhibition of the elevation in myogenin mRNA, and the increase in cell number. In contrast, IGFBP-5 stimulated all aspects of the myogenic response to IGF-I: the later rise in myogenin mRNA, the elevation of creatine kinase activity, and the fusion of myoblasts into myotubes. This dual response to IGFBP-5 was greatest when it was added at a molar ratio of IGFBP-5 to IGF-I of 2:1. In contrast, when IGFBP-5 was added in the presence of IGF-II, it inhibited both proliferation and differentiation. Neither IGFBP had any effect when added in the presence of R3 IGF-I, an analog with substantially reduced affinity for IGFBPs. Our results suggest that the role of IGFBP-4 is mainly to sequester excess IGFs, and thus inhibit all actions. IGFBP-5, however, is capable of eliciting a dual response, possibly due to its unique ability to associate with the cell membrane.
Ewton
D Z
DZ
Department of Biology, Syracuse University, New York 13244, USA. dzewton@mailbox.syr.edu
Coolican
S A
SA
Mohan
S
S
Chernausek
S D
SD
Florini
J R
JR
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D055504:Insulin Receptor Substrate Proteins; D018974:Insulin-Like Growth Factor Binding Protein 4 / Q000494:pharmacology* / Q000502:physiology; D018975:Insulin-Like Growth Factor Binding Protein 5 / Q000494:pharmacology* / Q000502:physiology; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology / Q000502:physiology*; D007335:Insulin-Like Growth Factor II / Q000494:pharmacology; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D018485:Muscle Fibers, Skeletal / Q000166:cytology* / Q000201:enzymology; D018008:Myogenin / Q000235:genetics; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011506:Proteins / Q000378:metabolism; D016760:Proto-Oncogene Proteins c-fos / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D055765:Shc Signaling Adaptor Proteins; D015398:Signal Transduction / Q000187:drug effects*; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D014443:Tyrosine / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D004791:Enzyme Inhibitors; D005419:Flavonoids; C526292:IRS1 protein, human; D055504:Insulin Receptor Substrate Proteins; D018974:Insulin-Like Growth Factor Binding Protein 4; D018975:Insulin-Like Growth Factor Binding Protein 5; C503799:MYOG protein, human; D018008:Myogenin; D010750:Phosphoproteins; D011506:Proteins; D016760:Proto-Oncogene Proteins c-fos; D012333:RNA, Messenger; C528024:SHC1 protein, human; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D014443:Tyrosine; D007334:Insulin-Like Growth Factor I; D007335:Insulin-Like Growth Factor II; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one