Analyses of steroid receptors are important for understanding molecular details of transcriptional control, as well as providing insight as to how an individual transacting factor contributes to cell identity and function. These studies have led to the identification of a superfamily of regulatory proteins that include receptors for thyroid hormone and the vertebrate morphogen retinoic acid. Although animals employ complex and often distinct ways to control their physiology and development, the discovery of receptor-related molecules in a wide range of species suggests that mechanisms underlying morphogenesis and homeostasis may be more ubiquitous than previously expected.

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The steroid and thyroid hormone receptor superfamily

Alternative processing of the RNA transcribed from the calcitonin gene appears to result in the production of a messenger RNA in neural tissue distinct from that in thyroidal 'C' cells The thyroid mRNA encodes a precursor to the hormone calcitonin whereas that in neural tissues generates a novel neuropeptide, referred to as calcitonin gene-related peptide (CGRP) The distribution of CGRP-producing cells and pathways in the brain and other tissues suggests functions for the peptide in nociception, ingestive behaviour and modulation of the autonomic and endocrine systems The approach described here permits the application of recombinant DNA technology to analyses of complex neurobiological systems in the absence of prior structural or biological information

Production of a novel neuropeptide encoded by the calcitonin gene via tissue-specific RNA processing

Identification of complementary DNAs encoding the human glucocorticoid receptor predicts two protein forms, of 777 (alpha) and 742 (beta) amino acids, which differ at their carboxy termini. The proteins contain a cysteine/lysine/arginine-rich region which may define the DNA-binding domain. Pure radiolabelled glucocorticoid receptor, synthesized in vitro, is immunoreactive and possesses intrinsic steroid-binding activity characteristic of the native glucocorticoid receptor.

/pdf/primary-structure-and-expression-of-a-functional-human-9s709u9sel.pdf

Primary structure and expression of a functional human glucocorticoid receptor cDNA

The glucocorticoid receptor (Gr, encoded by the gene Grl1) controls transcription of target genes both directly by interaction with DNA regulatory elements and indirectly by cross-talk with other transcription factors. In response to various stimuli, including stress, glucocorticoids coordinate metabolic, endocrine, immune and nervous system responses and ensure an adequate profile of transcription. In the brain, Gr has been proposed to modulate emotional behaviour, cognitive functions and addictive states. Previously, these aspects were not studied in the absence of functional Gr because inactivation of Grl1 in mice causes lethality at birth (F.T., C.K. and G.S., unpublished data). Therefore, we generated tissue-specific mutations of this gene using the Cre/loxP -recombination system. This allowed us to generate viable adult mice with loss of Gr function in selected tissues. Loss of Gr function in the nervous system impairs hypothalamus-pituitary-adrenal (HPA)-axis regulation, resulting in increased glucocorticoid (GC) levels that lead to symptoms reminiscent of those observed in Cushing syndrome. Conditional mutagenesis of Gr in the nervous system provides genetic evidence for the importance of Gr signalling in emotional behaviour because mutant animals show an impaired behavioural response to stress and display reduced anxiety.

Disruption of the glucocorticoid receptor gene in the nervous system results in reduced anxiety

The recent availability in culture of embryo-derived pluripotential cells which exhibit both a normal karyotype and a high differentiative ability has encouraged us to assess the potential of these cells to form functional germ cells following their incorporation into chimaeric mice We report here the results of blastocyst injection studies using three independently isolated XY embryo-derived cell lines (EK CP1 , EK CC1 1 and EKCC1 2) which produce a very high proportion (greater than 50%) of live-born animals that are overtly chimaeric Seven chimaeric male mice, derived from these three lines, have, so far, proved to be functional germ-line chimaeras

Formation of germ-line chimaeras from embryo-derived teratocarcinoma cell lines

A DNA fragment containing the promoter of the mouse metallothionein-I gene fused to the structural gene of rat growth hormone was microinjected into the pronuclei of fertilized mouse eggs. Of 21 mice that developed from these eggs, seven carried the fusion gene and six of these grew significantly larger than their littermates. Several of these transgenic mice had extraordinarily high levels of the fusion mRNA in their liver and growth hormone in their serum. This approach has implications for studying the biological effects of growth hormone, as a way to accelerate animal growth, as a model for gigantism, as a means of correcting genetic disease, and as a method of farming valuable gene products.

/pdf/dramatic-growth-of-mice-that-develop-from-eggs-microinjected-39d8dk7cie.pdf

Dramatic growth of mice that develop from eggs microinjected with metallothionein–growth hormone fusion genes

In order to define the molecular mechanisms by which glucocorticoids and thyroid hormone act to regulate growth hormone gene expression, the sites at which they exert their effects on growth hormone biosynthesis were examined in vivo and in a pituitary cell line. Glucocorticoids were shown to rapidly increase accumulation of growth hormone mRNA and nuclear RNA precursors. Glucocorticoids and thyroid hormone were shown to rapidly and independently increase growth hormone gene transcription. These events are shown to occur physiologically in animals and further establish the importance of growth hormone gene expression as a model for steroid regulation.

https://www.pnas.org/content/pnas/79/24/7659.full.pdf

Glucocorticoid and thyroid hormones transcriptionally regulate growth hormone gene expression

After adrenalectomy, the plasma levels of adrenocorticotropic hormone (corticotropin, ACTH)/endorphin peptides in rats rise dramatically in the first 4 hr while pituitary peptide levels fall sharply. Eight hours after adrenalectomy, plasma levels are near control values again but they then increase continuously over the next 8 days. Proopiomelanocortin (POMC) mRNA levels in the anterior pituitary (quantitated by hybridization with cloned POMC cDNA) increase 2-fold in the first 24 hours, reaching 15- to 20-fold the control level 18 days after adrenalectomy. When dexamethasone is administered to rats 8 days after adrenalectomy, the above events are reversed. Plasma ACTH falls to control levels within 2 hr whereas anterior pituitary POMC mRNA requires 5 days of treatment for return to control levels. The levels of POMC mRNA in the neurointermediate lobe and the hypothalamus are not altered by either treatment. Adrenalectomy increases transcription of the POMC gene in the anterior pituitary approximately 20-fold and halves transcription of the growth hormone gene within 1 hr of operation. Administration of dexamethasone immediately after adrenalectomy suppresses the increase in transcription of the POMC gene and increases the transcription of the growth hormone gene. Transcription of the POMC gene(s) in the neurointermediate lobe is not altered by either of these treatments.

https://www.pnas.org/content/pnas/80/22/6982.full.pdf

Neal C. Birnberg

Papers

Dramatic growth of mice that develop from eggs microinjected with metallothionein–growth hormone fusion genes

Glucocorticoid and thyroid hormones transcriptionally regulate growth hormone gene expression

Glucocorticoids regulate proopiomelanocortin gene expression in vivo at the levels of transcription and secretion

Dramatic growth of mice that develop from eggs microinjected with metallothionein-growth hormone fusion genes. 1982.

3 – Biosynthesis of ACTH and Related Peptides