Corticosteroids are critical for development and for mediating stress responses across diverse vertebrate taxa. Study of frog metamorphosis has made significant breakthroughs in our understanding of corticosteroid signaling during development in non-mammalian vertebrate species. However, lack of adequate corticosterone (CORT) response genes in tadpoles make identification and quantification of CORT responses challenging. Here, we characterized a CORT-response gene
cyp21a2 Knockout Tadpoles Survive Metamorphosis Despite Low Corticosterone
Abstract Corticosteroids are so vital for organ maturation that reduced corticosteroid signaling during postembryonic 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 (WT) 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 WT 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 messenger RNA expression levels of crh, pomc, star, and cyp11b2 and an approximately 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 corticosterone 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.
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- Award ID(s):
- 2035732
- NSF-PAR ID:
- 10385255
- Date Published:
- Journal Name:
- Endocrinology
- Volume:
- 164
- Issue:
- 1
- ISSN:
- 1945-7170
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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frzb (frizzled related protein) previously identified inXenopus tropicalis tadpole tail skin by an RNA-seq study. We validated the RNA-seq results that CORT and not thyroid hormone inducesfrzb in the tails using quantitative PCR. Further, maximumfrzb expression was achieved by 100-250 nM CORT within 12-24 hours.frzb is not significantly induced in the liver and brain in response to 100 nM CORT. We also found no change infrzb expression across natural metamorphosis when endogenous CORT levels peak. Surprisingly,frzb is only induced by CORT inX. tropicalis tails and not inXenopus laevis tails. The exact downstream function of increasedfrzb expression in tails in response to CORT is not known, but the specificity of hormone response and its high mRNA expression levels in the tail renderfrzb a useful marker of exogenous CORT-response independent of thyroid hormone for exogenous hormone treatments andin-vivo endocrine disruption studies. -
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- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1210/endocr/bqac182
