Role of epidermal growth factor and ErbB2 receptors in 3T3-L1 adipogenesis.
OBJECTIVE
Epidermal growth factor (EGF) stimulates proliferation in 3T3-L1 preadipocytes, but EGF action in differentiation is less clear. EGF promotes differentiation at concentrations <1 nM but inhibits differentiation at higher concentrations, suggesting a dual role in adipogenesis. We hypothesized that differences in EGF receptor activation and downstream signaling mediate distinct biological effects of EGF at low vs. high abundance.


METHODS
We compared the effects of low (0.1 nM) vs. high (10 nM) EGF on the activation of EGF receptors, proximal signaling molecules Src and Shc, and the downstream mitogen-activated protein kinase (MAPK) pathways extracellular regulated kinase (ERK) and p38 in proliferating and differentiated 3T3-L1 cells.


RESULTS
Both low and high EGF activated ERK and p38 in preadipocytes. Src inhibitors PP1 and PP2 blocked ERK and p38 activation by low but not high EGF, and only high EGF increased Shc phosphorylation. Selective inhibition of the EGF receptor (EGFR) with AG1478 blocked ERK and p38 activation at both concentrations; however, selective inhibition of the ErbB2 receptor (EB2R) with AG825 or small interfering RNA (siRNA) blocked low but not high EGF activation of ERK and p38. Coimmunoprecipitation of EGFR with EB2R and Src was observed with low EGF in preadipocytes but at both concentrations in adipocytes. EB2R inhibition during differentiation decreased p38 activity and peroxisome proliferator-activated receptor gamma (PPARgamma) abundance.


CONCLUSIONS
Our results show that EGFR homodimers mediate action of EGF at high abundance, but at low abundance, EGF promotes differentiation through EGFR/EB2R heterodimer activation of Src and p38. These results may partially explain the observations that high EGF concentrations inhibit, whereas low concentrations support, preadipocyte differentiation.
Harrington
Molly
M
Department of Pediatrics, Rhode Island Hospital and Brown Medical School, Providence, Rhode Island 02903, USA.
Pond-Tor
Sunthorn
S
Boney
Charlotte M
CM
D041721:3T3-L1 Cells; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D050156:Adipogenesis / Q000187:drug effects*; D000818:Animals; D000081247:CSK Tyrosine-Protein Kinase; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004815:Epidermal Growth Factor / Q000494:pharmacology* / Q000502:physiology; D066246:ErbB Receptors / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D051379:Mice; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000378:metabolism / Q000502:physiology*; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism; D019061:src-Family Kinases
D048868:Adaptor Proteins, Signal Transducing; D000081247:CSK Tyrosine-Protein Kinase; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D048049:Extracellular Signal-Regulated MAP Kinases; D011505:Protein-Tyrosine Kinases; D018719:Erb-b2 Receptor Tyrosine Kinases; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D048051:p38 Mitogen-Activated Protein Kinases; D019061:src-Family Kinases; C528034:Shc1 protein, mouse