Dysfunction of the endoplasmic reticulum (ER) has been reported in a variety of human pathologies, including cancer. However, the contribution of the ER to the early stages of normal cell transformation is largely unknown. Using primary human melanocytes and biopsies of human naevi (moles), we show that the extent of ER stress induced by cellular oncogenes may define the mechanism of activation of premature senescence. Specifically, we found that oncogenic forms of HRAS (HRAS(G12V)) but not its downstream target BRAF (BRAF(V600E)), engaged a rapid cell-cycle arrest that was associated with massive vacuolization and expansion of the ER. However, neither p53, p16(INK4a) nor classical senescence markers--such as foci of heterochromatin or DNA damage--were able to account for the specific response of melanocytes to HRAS(G12V). Instead, HRAS(G12V)-driven senescence was mediated by the ER-associated unfolded protein response (UPR). The impact of HRAS on the UPR was selective, as it was poorly induced by activated NRAS (more frequently mutated in melanoma than HRAS). These results argue against premature senescence as a converging mechanism of response to activating oncogenes and support a direct role of the ER as a gatekeeper of tumour control.
Phosphorylation of the cell adhesion protein CEACAM1 increases insulin sensitivity and decreases insulindependent mitogenesis in vivo. Here we show that CEACAM1 is a substrate of the EGFR and that upon being phosphorylated, CEACAM1 reduces EGFR-mediated growth of transfected Cos-7 and MCF-7 cells in response to EGF. Using transgenic mice overexpressing a phosphorylation-defective CEACAM1 mutant in liver (L-SACC1), we show that the effect of CEACAM1 on EGF-dependent cell proliferation is mediated by its ability to bind to and sequester Shc, thus uncoupling EGFR signaling from the ras/MAPK pathway. In L-SACC1 mice, we also show that impaired CEACAM1 phosphorylation leads to ligand-independent increase of EGFR-mediated cell proliferation. This appears to be secondary to visceral obesity and the metabolic syndrome, with increased levels of output of free fatty acids and heparin-binding EGF-like growth factor from the adipose tissue of the mice. Thus, L-SACC1 mice provide a model for the mechanistic link between increased cell proliferation in states of impaired metabolism and visceral obesity.
Phosphorylation of the cell adhesion protein CEACAM1 increases insulin sensitivity and decreases insulindependent mitogenesis in vivo. Here we show that CEACAM1 is a substrate of the EGFR and that upon being phosphorylated, CEACAM1 reduces EGFR-mediated growth of transfected Cos-7 and MCF-7 cells in response to EGF. Using transgenic mice overexpressing a phosphorylation-defective CEACAM1 mutant in liver (L-SACC1), we show that the effect of CEACAM1 on EGF-dependent cell proliferation is mediated by its ability to bind to and sequester Shc, thus uncoupling EGFR signaling from the ras/MAPK pathway. In L-SACC1 mice, we also show that impaired CEACAM1 phosphorylation leads to ligand-independent increase of EGFR-mediated cell proliferation. This appears to be secondary to visceral obesity and the metabolic syndrome, with increased levels of output of free fatty acids and heparin-binding EGF-like growth factor from the adipose tissue of the mice. Thus, L-SACC1 mice provide a model for the mechanistic link between increased cell proliferation in states of impaired metabolism and visceral obesity.
The New Zealand obese (NZO) mouse strain shares with
the related New Zealand black (NZB) strain a number of
immunophenotypic traits. Among these is a high proportion
of B-1 B lymphocytes, a subset associated with autoantibody
production. Approximately 50% of NZO/HlLt
males develop a chronic insulin-resistant type 2 diabetes
syndrome associated with 2 unusual features: the presence
of B lymphocyte–enriched peri-insular infiltrates and
the development of anti-insulin receptor autoantibodies
(AIRAs). To establish the potential pathogenic contributions
ofBlymphocytes and AIRAs in this model, a disrupted immunoglobulin heavy chain gene (Igh-6) congenic on the
NZB/BlJ background was backcrossed 4 generations into
the NZO/HlLt background and was then intercrossed to
produce mice that initially segregated for wild-type versus
the mutant Igh-6 allele and thus permitted comparison
of syndrome development. A new flow cytometric assay
(AIRA binding to transfected Chinese hamster ovary
cells stably expressing mouse insulin receptor) showed IgM
and IgG subclass AIRAs in serum from Igh-6 intact males,
but not in Igh6null male serum. However, the absence of
B lymphocytes and antibodies distinguishing mutant from
wild-type males failed to significantly affect diabetes-free
survival. The Igh6nullmales gained weight less rapidly than
wild-type males, probably accounting for a retardation, but
not prevention, of hyperglycemia. Thus, AIRA and the Blymphocyte
component of the peri-insulitis in chronic diabetics
were not essential either to development of insulin
resistance or to eventual pancreatic beta cell failure and
loss. A new substrain, designated NZL, was generated by
inbreeding Igh-6 wild-type segregants. Currently at the F10
generation, NZL mice exhibit the same juvenile-onset obesity
as NZO/HlLt males, but develop type 2 diabetes at a
higher frequency (> 80%). Also, unlike NZO/HlLt mice that
are difficult to breed, the NZL/Lt strain breeds well and thus
offers clear advantages to obesity/diabetes researchers.
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