LAUSR

Corticosteroids are vital for organ maturation such that reduced corticosteroid signaling during post-embryonic development causes death in terrestrial vertebrates. Indeed, death occurs at metamorphosis in frogs lacking proopiomelanocortin (pomc) or the glucocorticoid receptor (GR; nr3c1). Some residual corticosteroids exist in pomc mutants to activate the wild-type GR and mineralocorticoid receptor (MR), and the elevated corticosteroids in GR mutants may activate MR. Thus, we expected a more severe developmental phenotype in tadpoles with inactivation of 21-hydroxylase, which should eliminate all interrenal corticosteroid biosynthesis. Using CRISPR/Cas9 in Xenopus tropicalis we produced an 11-base pair deletion in cyp21a2, the gene encoding 21-hydroxylase. Growth and development were delayed in cyp21a2 mutant tadpoles, but unlike the other frog models, they survived metamorphosis. Consistent with an absence of 21-hydroxylase, mutant tadpoles had a 95% reduction of aldosterone in tail tissue, but they retained some corticosterone (∼40% of wild-type siblings), an amount, however, too low for survival in pomc mutants. Decreased corticosteroid signaling was evidenced by reduced expression of corticosteroid-response gene, klf9 and by impaired negative feedback in the hypothalamus-pituitary-interrenal axis with higher mRNA expression levels of crh, pomc, star, and cyp11b2 and a ∼30-fold increase in tail content of progesterone. In-vitro tail tip culture showed that progesterone can transactivate the frog GR. The inadequate activation of GR by CORT in cyp21a2 mutants was likely compensated for by sufficient corticosteroid signaling from other GR ligands to allow survival through the developmental transition from aquatic to terrestrial life.