Search

We have isolated and sequenced all 23 members of the 22-kD alpha zein (z1C) gene family of maize. This is one of the largest plant gene families that has been sequenced from a single genetic background and includes the largest contiguous genomic DNA from maize with 346,292 bp to date. Twenty-two of the z1C members are found in a roughly tandem array on chromosome 4S forming a dense gene cluster 168,489-bp long. The twenty-third copy of the gene family is also located on chromosome 4S at a site approximately 20 cM closer to the centromere and appears to be the wild-type allele of the floury-2 (fl2) mutation. On the basis of an analysis of maize cDNA databases, only seven of these genes appear to be expressed including the fl2 allele. The expressed genes in the cluster are interspersed with nonexpressed genes. Interestingly, some of the expressed genes differ in their transcriptional regulation. Gene amplification appears to be in blocks of genes explaining the rapid and compact expansion of the cluster during the evolution of maize. more...

The action of several synthesis inhibitors of arachidonic acid metabolites on ovulation in the rat has been investigated, and the effectiveness of intraventricular microinjections vs intracerebral injections has been compared. Indomethacin, aspirin, and paracetamol, three prostaglandin (PG) synthesis inhibitors, were completely ineffective at the doses given in blocking ovulation when injected into the third ventricle on the morning of proestrus. Similar results were obtained with intraventricular injections of 15-hydroperoxyarachidonic acid (15-HPAA), which selectively inhibits the formation of prostacyclin from the endoperoxide intermediates, and with imidazole, an inhibitor of thromboxane biosynthesis. In contrast, intracerebral administration of either indomethacin or aspirin blocked ovulation in a significant proportion of rats when infused directly into the preoptic area or the anterior hypothalamic area of the brain. However, intracerebral injections of either paracetamol or 15-HPAA produced no ovulatory blockade. Although imidazole showed no apparent effect when injected in the anterior hypothalamus, infusion into the preoptic area blocked ovulation in a small proportion of animals. The results provide further evidence for the role of PGs in the hypothalamic control of ovulation and suggest that the PGs of the E and F series are most likely to be the PG type involved, and not thromboxanes or prostacyclin. more...

We report the purification of human corticotrophin-releasing factor-binding protein (hCRF-BP) using repeated affinity chromatography (with the aid of synthetic CRF immobilized on Sepharose 4B solid phase) followed by gel filtration. Presence of the binding protein was tracked throughout the procedure by its ability to inhibit binding of 125I-labelled hCRF to an hCRF antiserum; normal human plasma exhibits 80% inhibition in this system whereas sheep plasma, which does not contain an hCRF-BP, has no effect. Affinity cross-linking of 125I-labelled hCRF to the purified hCRF-BP was performed using disuccinimidyl suberate (1 mmol/l). SDS electrophoresis of the purified CRF cross-linked binding protein followed by radioautography resulted in one major band of Mr37 000 which corresponded to our original molecular weight estimate based on the elution position of the binding protein on Sephacryl S-200. A 107-fold purification of the hCRF-BP resulted in a preparation estimated to be 95% pure, with an overall yield of 5%.Journal of Endocrinology(1989) 122,23–31 more...

Regulation of the rapid compensatory growth seen in the remaining adrenal gland of rats following unilateral adrenalectomy is poorly understood. The role of adrenocorticotropic hormone (ACTH) is obscure1,2as immunoneutralization of circulating ACTH does not affect the observed compensatory growth or hyperplasia1. This finding, together with the fact that mechanical manipulation of one adrenal without extirpation is followed by growth only in the contralateral gland3, has led to the concept of neural regulation of compensatory adrenal growth via a loop from one adrenal through the hypothalamus and back to the contralateral gland which is independent of ACTH secretion3. We recently showed that peptides from the N terminal of ACTH precursor proopiocortin (POC), not containing the γ-melanocyte-stimulating hormone (γ-MSH) sequence, can stimulate adrenal mitogenesis and proposed that normal long-term adrenal growth and proliferation involves post-secretional proteolytic cleavage of pro-γ-MSH [or N-POC(1–74)] to generate the mitogenic factor N-POC(1–48/49) and a C-terminal fragment N-POC(50–74), or rat γ3-MSH4. We have now investigated this hypothesis further in rats by selectively quenching different regions of circulating POC peptides with specific antisera and observing the effect on the increases in weight, RNA and DNA normally seen in the remaining gland following unilateral adrenalectomy1–3. Our results, reported here, suggest that neurally mediated proteolytic cleavage of the circulating inactive mitogenic precursor pro-γ-MSH at the adrenal gland is the major mechanism of control of compensatory growth. more...

We have expressed human pre-procorticotrophin-releasing hormone (pre-proCRH) as a fusion protein to β-galactosidase in Escherichia coli. The chimeric fusion protein was found in insoluble bacterial inclusion bodies. The inclusion bodies were isolated, purified and solubilized, and used as imunogens in rabbits to raise antibodies against the neuropeptide moiety. The antibodies generated were characterized by immunoassays and immunocytochemical techniques. The immunoassay results showed that the recombinant pre-proCRH antibodies cross-reacted with the full-length CRH precursor and several cleavage products derived from it, i.e. CRH(1–41) and CRH(36–41). They did not cross-react with the CRH antagonist CRH(9–41). Extracts of stalk median eminence from various species were also studied. The antibodies cross-reacted with extracts from ovine, bovine, human and rat tissues, exhibiting parallel displacement curves to that of synthetic rat/human CRH(1–41) used as standard. They also cross-reacted with a skin extract of the frog, a species known to contain a CRH-related peptide, i.e. sauvagine, in this tissue. The immunocytochemical studies demonstrated that the antibodies generated against recombinant human preproCRH labelled neurones in the rat paraventricular nucleus of the hypothalamus. They exhibited the same pattern of staining as that obtained with an antibody generated against synthetic CRH(1–41). The results indicate that these antibodies can recognize CRH(1–41) or CRH-related molecules in the hypothalamus in situas well as in tissue extracts from several species. Hence, they will be useful tools in the study of the CRH biosynthetic pathway and its intracellular compartmentalization. more...

Samples of maternal blood, amniotic fluid and umbilical arterial and venous blood were collected from 11 women at 16–24 weeks of pregnancy. Corticotrophin-releasing hormone-41 (CRH-41) and ACTH were measured by immunoradiometric assay. The mean levels of ACTH were 11 pmol/l in maternal plasma, 12 pmol/l in fetal plasma and 9·7 pmol/l in amniotic fluid. The mean levels of CRH-41 were 1·6 pmol/l in maternal plasma and 0·7 pmol/l in fetal plasma. There was a positive correlation between maternal and fetal plasma CRH-41 and between maternal CRH-41 and ACTH. In fetal plasma there was a weak inverse correlation between CRH-41 and ACTH. This is the first demonstration of CRH-41 in the circulation of the mid-trimester human fetus, but on the basis of the present findings it is not possible to specify the exact source (fetal, placental or maternal).J. Endocr.(1987) 114,497–501 more...

Abstract. CRF-41 and vasopressin are the major part of the hypothalamic complex responsible for the release of ACTH. The potentiated response of the corticotroph to mixtures of the peptides and their co-localization in the same neurosecretory vesicles would indicate their obligatory co-secretion in many stress situations. The occurrence of CRF at peripheral sites especially in the placenta and in the plasma of women in their third trimester of pregnancy is suggestive of further roles for this peptide although care should be exercised in interpreting simple immunohistochemical staining in certain tissues. When ACTH is released from the pituitary several other peptides which form part of its precursor are co-released. The effect of these peptides on the adrenal gland and the way their activity is expressed has led to a post-secretional proteolytic control mechanism being proposed. more...