The major type-1 protein phosphatase catalytic subunits are the same gene products in rabbit skeletal muscle and rabbit liver

The catalytic subunit of protein phosphatase-1 (PP1) isolated from rabbit liver had the same electrophoretic mobility as, and yielded peptide maps identical to those of the 33 kDa form of rabbit skeletal muscle PP1. The predicted amino-acid sequences of PP1 obtained from three rabbit liver cDNA clones were identical to that of PP1 alpha from rabbit skeletal muscle. These findings suggest that the distinctive substrate specificities and regulatory properties of hepatic and skeletal muscle type-1 protein phosphatases are not conferred by the catalytic subunits themselves, but by regulatory subunits that are complexed to the catalytic subunits in vivo.     

The receptors for prolactin and growth hormone are localized in the same region of human chromosome 5

Prolactin receptor (PRLR) and growth hormone receptor (GHR) are encoded by members of a gene family containing regions of identical sequences. To determine their chromosomal locations, cDNA probes for these genes were used. Analysis of hybridization to several somatic cell hybrids, together with hybridization in situ to metaphase chromosomes, resulted in the assignment of the loci for both receptors to human chromosome 5 in the region p13----p14. Thus, these proteins may be encoded by a cluster of related genes that evolved from a common ancestral gene, in a manner consonant with that of their ligands.     

Deficient glycosylation of arylsulfatase A in pseudo arylsulfatase-A deficiency

Summary Deficient arylsulfatase-A activity is diagnostic of a neurodegenerative human lysosomal storage disease, metachromatic leukodystrophy. Paradoxically, similar enzyme deficiency also occurs in normal individuals, who are known as being pseudo arylsulfatase-A deficient. We showed previously that this phenotype is associated with a structural gene mutation that produces an exceptionally labile enzyme. We now report on the nature and consequence of this mutation. When the mutant arylsulfatase-A is deglycosylated by endoglycosidase H, only one smaller molecular species was generated, instead of the two from the normal enzyme. This is consistent with the loss of one of the two N-linked oligosaccharide side chains known to be present on the wild-type enzyme. Quantitative analysis of mannose and leucine incorporation showed that the mutant enzyme incorporated two- to tenfold less mannose than the normal enzyme on a molar basis. This deficient glycosylation was specific to arylsulfatase-A. Another lysosomal enzyme not affected in this mutation, beta-hexosaminidase, was glycosylated normally in the mutant cells. The remaining single oligosaccharide side chain released from the mutant arylsulfatase-A by pronase digestion was normally processed to complex and high-mannose forms. However, the high-mannose side chains contained 30% fewer phosphorylated residues than those of the normal enzyme. Nevertheless, this reduced level of phosphorylation did not prevent targeting of the mutant enzyme to the lysosomes, a process normally mediated through phosphorylated mannose residues. In conclusion, pseudo arylsulfatase-A deficiency is a unique human mutation associated with reduced glycosylation and phosphorylation of a lysosomal enzyme with the loss of one of the two carbohydrate side chains. The mutation results in greatly reduced enzyme stability, thus indicating a role for oligosaccharides in maintaining enzyme stability within the degradative environment of the lysosomes. However, the residual catalytic activity or subcellular targeting of the mutant enzyme was not affected. These properties probably account for the benign clinical presentation of pseudo arylsulfatase-A deficiency.Abbreviations PD Pseudo arylsulfatase-A Deficiency - ARA Arylsulfatase-A     

The human PDGF receptor α-subunit gene maps to chromosome 4 in close proximity to c-kit

Summary The gene encoding the -subunit of the human platelet-derived growth factor receptor (PDGFRA) maps to band q11–q12 of chromosome 4 by in situ hybridization, which was confirmed by Southern analysis of a Chinese hamster × human cell hybrid that retains only human chromosome 4.     

Diurnal Variations of Beta-Endorphin at Rest and After Moderate Intensity Exercise

To determine the effect of time of day on circulating beta-endorphin concentrations 14 men exercised at 75% of their maximal capacity at 0600, 1200, 1800 and 2400 hr. Each trial was separated by 3-5 days and preceded by a normal sleep cycle except for the 0600 hr trials which was preceded by 6 hr sleep. Resting physiological data indicated normal diurnal variations in heart rate, core temperature and oxygen uptake, being lowest during the 0600 hr trials and highest during the 1800 hr trials. Resting plasma beta-endorphin concentrations averaged 11.9 +/- 8.4 pmol/l during the 0600 hr trials, significantly greater than the 2400 hr trials (6.4 +/- 3.6 pmol/l; P less than 0.05). No other significant differences existed at rest. Post exercise beta-endorphin concentrations were elevated and found to be inversely related to time of day with the 0600 hr trials having the highest mean (25.7 +/- 14.7) and the 2400 hr trials the lowest (14.7 +/- 8.3). These data suggest that the plasma beta-endorphin concentrations at rest and after exercise are affected by the time of day. The results also suggest that the changes in beta-endorphin associated with exercise are not major contributors to cardiorespiratory control or changes in psychological effect associated with exercise.     

Comparison of antiprogestin stimulation of uterine prostaglandin synthesis in vitro

Progesterone has an inhibitory effect on prostaglandin synthesis in urine tissue and this effect is reversible with progesterone receptor antagonists. Although antiprogesterone steroids such as RU486 (Mifepristone) are effective at inducing abortion in women they have an improved efficacy when used with exogenous synthetic prostaglandin. In the guinea-pig such antagonists sensitize the uterus but do not result in increased myometrial activity and therefore may not induce endogenous PG synthesis. In this study the effects of antiprogestins on a preparation of rat uterus perifused with progesterone were studied. ZK98 734 caused a rapid and sustained increase in 6-oxoPGF synthesis which rose within the first 90 minutes. This rapid response suggested that some mechanism other than the induction of fresh protein synthesis was involved. A similar increase was not seen with pregnant guinea-pig myometrium/decidua perifused in a similar manner, suggesting that some other mechanism was responsible for the relatively low PG production in pregnancy. However increases in 6-oxoPGF in response to antiprogestins were recorded when pregnant guinea-pig decidua/myometrium was incubated for 4 hours. In these experiments 1 microM ZK98 734 and 1 microM ZK98 299 (Onapristone) gave a 2.7 fold increase in PG production whereas RU486 gave a 1.6 fold increase. Both 1 microM ZK98 734 and 1 microM ZK98 299 also gave a significant increase in PGE production but no increase in PGF was observed. These findings suggest that some antiprogestins might have a better effect on the stimulation of endogenous PG synthesis or on the rate of catabolism of prostanoids.     

Antibodies which recognize the C-terminus of the inhibitory guanine-nucleotide-binding protein (Gi) demonstrate that opioid peptides and foetal-calf serum stimulate the high-affinity GTPase activity of two separate pertussis-toxin substrates.

We investigated the mechanisms of receptor-mediated stimulation of high-affinity GTPase activity in response to opioid peptides and to foetal-calf serum in membranes of the neuroblastoma X glioma hybrid cell line NG108-15. Increases in GTPase activity in response to both of these ligands was abolished by prior exposure of the cells to pertussis toxin. Pertussis toxin in the presence of [32P]NAD+ catalysed incorporation of radioactivity into a broad band of approx. 40 kDa in membranes prepared from untreated, but not from pertussis-toxin-pretreated, cells. Additivity studies indicated that the responses to opioid peptides and to foetal-calf serum were mediated by separate guanine-nucleotide-binding proteins (G-proteins). Whereas opioid peptides produced an inhibition of adenylate cyclase in membranes of untreated cells, foetal-calf serum did not. Affinity-purified antibodies which recognize the C-terminus of the inhibitory G-protein identified a 40 kDa polypeptide in membranes of NG108-15 cells. These antibodies attenuated opioid-stimulated high-affinity GTPase activity, but did not markedly affect the response to foetal-calf serum. We conclude that receptors for the opioid peptides function via the inhibitory G-protein (Gi), whereas foetal-calf serum activates a second pertussis-toxin-sensitive G-protein, which has a C-terminal sequence significantly different from that of Gi.