GRKO mice express an aberrant dexamethasone-binding glucocorticoid receptor, but are profoundly glucocorticoid resistant

The introduction of a targeted insertion mutation into exon 2 of the gel-it coding for thr glucocorticoid receptor (GR) enabled production of glucocor ticoid receptor knock-out (GRKO) mice. GRKO mice on a C57BL/6/129sv mixed g enetic background show a variable phenotype. with 90% of -/- mice dying at birth with respiratory insufficiency but 10% of mutant mice surviving to ma turity. To investigate the possibility of residual GR expression in survivi ng GRKO mice wt: have measured binding of the synthetic glucocorticoid dexa methasone in tissue extracts from adrenalectomized mice. High affinity bind ing of dexamethasone in protein extracts of liver. kidney. lung and brain f rom adult GRKO mice is found at levels 30-60% those in wild-type mice. with heterozygotes (+/-) having intermediate levels. PCR and ribonuclease prote ction analysis showed comparable levels of GR mRNA on the 3' side of the ge ne-targeted insertional mutation in exon 2 of the GR gene, with almost no G R mRNA detected from exons 1 and 2 on the 5' side of the gene-targeted inse rtional mutation, Western blot analysis using a C-terminal specific GR anti body detects a 39 kDa GR fragment in extracts from adult GRKO mice. Despite the evidence for expression of a ligand-binding domain fragment of the glu cocorticoid receptor these mice are profoundly glucocorticoid resistant. wi th elevated levels of plasma ACTH and corticosterone. Thymocytes from adult and fetal GRKO mice are resistant to dexamethasone-induced apoptosis and c ultured fetal hepatocytes from GRKO mice are completely refractory to gluco corticoid induction of the gluconeogenic enzyme glucose-6-phosphatase. Thus although the surviving adult homozygous GRKO mice express a dexamethasone- binding GR fragment, their classic target tissues remain profoundly glucoco rticoid insensitive. (C) 2001 Elsevier Science Ireland Ltd. All rights rese rved.