Catalytically inactive SHIP2 inhibits proliferation by attenuating PDGF signaling in 3T3-L1 preadipocytes.
Inadequate proliferation and/or differentiation of preadipocytes may lead to adipose tissue dysfunction characterized by hypertrophied, insulin-resistant adipocytes. Platelet-derived growth factor (PDGF) may alter adipose tissue function by promoting proliferation of preadipocytes. Two principal signaling pathways that regulate proliferation are PI3K/PI(3,4,5)P3/Akt and Shc/Ras/ERK1/2. SH2 domain-containing inositol 5-phosphatase 2 (SHIP2) dephosphorylates PI(3,4,5)P3, and also binds to Shc. Our goal was to determine how SHIP2 affects these PDGF signaling routes. To assess the role of the 5-phosphatase domain, we expressed wild-type or catalytically inactive dominant-negative SHIP2 (P686A-D690A-R691A; PDR/AAA) in 3T3-L1 preadipocytes. Surprisingly, SHIP2 PDR/AAA inhibited proliferation more potently than wild-type SHIP2. After three days of proliferation, phospho-Akt, phospho-ERK1/2, and PDGF receptor (PDGFR) levels were reduced in PDR/AAA-expressing preadipocytes. SHIP2 PDR/AAA interference with PDGFR signaling was demonstrated using imatinib, an inhibitor of PDGFR tyrosine kinase. The anti-proliferative effect of imatinib observed in control preadipocytes was not significant in SHIP2 PDR/AAA-expressing preadipocytes, indicating a pre-existing impairment of PDGFR-dependent mitogenesis in these cells. The inhibition of PDGF-activated mitogenic pathways by SHIP2 PDR/AAA was consistent with a decrease in PDGFR phosphorylation caused by a drop in receptor levels in SHIP2 PDR/AAA-expressing cells. SHIP2 PDR/AAA promoted ubiquitination of the PDGFR and its degradation via the lysosomal pathway independently of the association between the E3 ubiquitin ligase c-Cbl and PDGFR. Overall, our findings indicate that SHIP2 PDR/AAA reduces preadipocyte proliferation by attenuating PDGFR signaling.
Artemenko
Yulia
Y
Chronic Disease Program, Ottawa Health Research Institute and Departments of Medicine and Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, Ontario, Canada.
Gagnon
Annemarie
A
Sorisky
Alexander
A
D041721:3T3-L1 Cells; D017667:Adipocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D019943:Amino Acid Substitution; D000818:Animals; D001483:Base Sequence; D001549:Benzamides; D002454:Cell Differentiation; D049109:Cell Proliferation / Q000187:drug effects; D017931:DNA Primers / Q000235:genetics; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D000068877:Imatinib Mesylate; D000072181:Inositol Polyphosphate 5-Phosphatases; D051379:Mice; D016297:Mutagenesis, Site-Directed; D000072183:Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D010879:Piperazines / Q000494:pharmacology; D010982:Platelet-Derived Growth Factor / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D050721:Proto-Oncogene Proteins c-cbl / Q000378:metabolism; D011743:Pyrimidines / Q000494:pharmacology; D017479:Receptors, Platelet-Derived Growth Factor / Q000037:antagonists & inhibitors; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013234:Stem Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D054875:Ubiquitination
D001549:Benzamides; D017931:DNA Primers; D010879:Piperazines; D010982:Platelet-Derived Growth Factor; D047428:Protein Kinase Inhibitors; D011743:Pyrimidines; D011994:Recombinant Proteins; D000068877:Imatinib Mesylate; D050721:Proto-Oncogene Proteins c-cbl; D017479:Receptors, Platelet-Derived Growth Factor; D051057:Proto-Oncogene Proteins c-akt; D048049:Extracellular Signal-Regulated MAP Kinases; D010744:Phosphoric Monoester Hydrolases; D000072181:Inositol Polyphosphate 5-Phosphatases; C000606435:Inppl1 protein, mouse; D000072183:Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases; C493849:Cbl protein, mouse