Expression of insulin receptors and of 60-kDa receptor substrate in rat mature and immature enterocytes.
The mechanism(s) by which rat immature enterocytes exhibit increased responsiveness to insulin before weaning is unknown. Therefore, we have analyzed the distribution, ontogeny, and molecular properties of insulin receptors (IR) and of related substrates in immature and mature enterocytes. IR were studied by radioligand binding assays, cross-linking labeling, immunohistochemistry, and in vitro phosphorylated substrates by immunoprecipitation. Regardless of age, 125I-insulin binding to IR was five times higher in crypt cells than in villus cells and two times higher in the ileum than in the jejunum. Binding capacity to villus cells from sucklings (day 14) exceeded three times that of older animals (day 30 and day 60). Scatchard analysis of equilibrium binding data confirmed an age-related decrease in low- and high-affinity receptor classes without change in affinity constants. In concordance, both alpha- and beta-IR subunits were more abundant in immature than in mature membranes. In vitro, insulin elicited the phosphorylation of three membrane proteins (96, 60 and 42 kDa), whose signals were virtually inhibited by preincubating membranes with antireceptor monoclonal antibodies. By immunoprecipitation, the 60-kDa signal was rapidly detected as a tyrosine-phosphorylated protein, expressed in mature and immature membranes, and identified as a receptor substrate phosphorylated in vitro by the IR tyrosine kinase. In conclusion, 1) increased responsiveness of rat immature enterocytes to insulin could be related to high membrane concentrations of IR and 2) normal rat enterocytes express a 60-kDa phosphotyrosine protein identified as a direct substrate of the IR tyrosine kinase.
Buts
J P
JP
Laboratory of Pediatric Gastroenterology and Nutrition, Faculty of Medicine, Catholic University of Louvain, Brussels, Belgium.
De Keyser
N
N
Marandi
S
S
Maernoudt
A S
AS
Sokal
E M
EM
Rahier
J
J
Hermans
D
D
D000375:Aging / Q000378:metabolism*; D000818:Animals; D018507:Gene Expression Regulation, Developmental; D007082:Ileum; D007328:Insulin / Q000378:metabolism; D055504:Insulin Receptor Substrate Proteins; D007413:Intestinal Mucosa / Q000166:cytology / Q000254:growth & development / Q000378:metabolism*; D007583:Jejunum; D008871:Microvilli / Q000378:metabolism / Q000648:ultrastructure; D010750:Phosphoproteins / Q000096:biosynthesis*; D010766:Phosphorylation; D051381:Rats; D017208:Rats, Wistar; D011972:Receptor, Insulin / Q000096:biosynthesis*
D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; C526296:Irs1 protein, rat; D010750:Phosphoproteins; D011972:Receptor, Insulin