Acute Ethanol Increases IGF‐I‐Induced Phosphorylation of ERKs by Enhancing Recruitment of p52‐Shc to the Grb2/Shc Complex

Abstract

Ethanol plays a detrimental role in the development of the brain. Multiple studies have shown that ethanol inhibits insulin-like growth factor I receptor (IGF-IR) function. Because the IGF-IR contributes to brain development by supporting neural growth, survival, and differentiation, we sought to determine the molecular mechanism(s) involved in ethanol’s effects on this membrane-associated tyrosine kinase. Using multiple neuronal cell types, we performed Western blot, immunoprecipitation, and GST-pulldowns following acute (1 – 24 hours) or chronic (3 weeks) treatment with ethanol. Surprisingly, exposure of multiple neuronal cell types to acute (up to 24 hours) ethanol (50mM) enhanced IGF-I-induced phosphorylation of ERKs, without affecting IGF-IR tyrosine phosphorylation itself, or Akt phosphorylation. This acute increase in ERKs phosphorylation was followed by the expected inhibition of the IGF-IR signaling following 3-week ethanol exposure. We then expressed a GFP-tagged IGF-IR construct in PC12 cells and used them to perform fluorescence recovery after photobleaching (FRAP) analysis. Using these fluorescently-labeled cells, we determined that 50mM ethanol decreased the half-time of the IGF-IR-associated FRAP, which implied that cell membrane-associated signaling events could be affected. Indeed, co-immunoprecipitation and GST-pulldown studies demonstrated that the acute ethanol exposure increased the recruitment of p52-Shc to the Grb2-Shc complex, which is known to engage the Ras-Raf-ERKs pathway following IGF-1 stimulation. These experiments indicate that even a short and low-dose exposure to ethanol may dysregulate function of the receptor, which plays a critical role in brain development.

Effects of Intermittent versus Chronic-Moderate Ethanol Administration during Adolescence in the Adult Hippocampal Phosphoproteome

“…Thus, it seems that the pattern of alcohol administration has a larger effect on the hippocampal phosphoproteome than the dose of alcohol used, which could be related to the intermittent pattern of consumption−withdrawal that characterizes it, since this withdrawal alters the glutamatergic system, causing excitotoxicity, 59 although the mechanism by which hippocampal deficits occur after intermittent alcohol consumption is not fully known. Among the seven proteins identified, only GRB2-2 has been previously related to ethanol use 60 and our results showed an increase in the Low-I group compared to the M group. GRB2-2 is a multifunctional adaptor protein that links many different growth factor receptors and an important signal transducer for numerous receptor tyrosine kinases, 61 such as the PI3K/Akt pathway, 62 which, interestingly, is also affected by ethanol as shown in previous studies from our group.…”

Primary cilia safeguard cortical neurons in neonatal mouse forebrain from environmental stress-induced dendritic degeneration

“…In fact, it has been shown that EtOH positively and negatively affects Akt phosphorylation at low and high doses, respectively, in hepatic insulin signaling in rats (77). Differential effects on IGF-1R activation based on the type of EtOH treatment (i.e., chronic exposure versus acute low-dose exposure) have also been reported in cultured neurons (79). Cilia-dependent IGF-1R signaling is required for cell-cycle progression in cortical progenitor cells (53,80) and the differentiation of preadipocytes (52).…”