Regulation of Calmodulin- and Dopamine-Stimulated Adenylate Cyclase Activities by Light in Bovine Retina

Abstract: Neural retina from most species contains 3,4-dihydroxyphenylethylamine (dopamine) receptors coupled to stimulation of adenylate cyclase activity. It has been demonstrated that release of dopamine from its neurons and subsequent occupation of dopamine receptors is increased by light. In this study, we have shown that adenylate cyclase activity in bovine retina is highly responsive to the endogenous Ca²⁺-binding protein, cal-modulin, and that calmodulin can increase dopamine-sen-sitive adenylate cyclase activity in bovine retina. We further demonstrate that both dopamine- and calmodulin-stimulated adenylate cyclase activities can be regulated by alterations in light. Bovine retinas were dissected from the eye under a low-intensity red safety light, defined as dark conditions, and incubated for 20 min in an oxygenated Krebs Henseleit buffer under either dark or light conditions. The retinas were then homogenized and adenylate cyclase activity measured in a paniculate fraction washed to deplete it of endogenous Ca²⁺ and calmodulin. Activation of adenylate cyclase activity by calmodulin, dopamine, and the nonhydrolyzable GTP analog, gua-nosine-5′-(β,γ-imido)triphosphate (GppNHp), was significantly (60%) greater in paniculate fractions from retinas that had been incubated under dark conditions as compared to those incubated under light conditions. Basal, Mn²⁺-, and GTP-stimulated adenylate cyclase activities were not altered by changes in lighting conditions. Calmodulin could increase the maximum stimulation of adenylate cyclase by dopamine in retinas incubated under either dark or light conditions, but the degree of its effect was greater in retinas incubated under light conditions. Activation of adenylate cyclase by calmodulin, dopamine, and GppNHp in paniculate fractions from retinas incubated under light conditions was indistinguishable from the activation obtained when retinas were incubated in the dark in the presence of exogenous dopamine. These results suggest that an increased release of dopamine occurs in light. The decreased response of adenylate cyclase to exogenous dopamine can then be explained by a subsequent down-regulation of dopamine receptor activity. The down-regulation of dopamine receptor activity can also regulate activation of adenylate cyclase by GppNHp and calmodulin. The results suggest that dopamine, calmodulin, and GppNHp are modulators of a common component of adenylate cyclase activity, and this component is regulated by light.     

Structure of tomato bushy stunt virus: V. Coat protein sequence determination and its structural implications

We report the chemically determined sequence of most of the polypeptide chain of the coat protein of tomato bushy stunt virus. Peptide locations have been determined by comparison with the high-resolution electron density map from X-ray crystallographic analysis as well as by conventional chemical overlaps. Three small gaps remain in the 387-residue sequence. Positively charged side-chains are concentrated in the N-terminal part of the polypeptide (the R domain) as well as on inward-facing surfaces of the S domain. There is homology of S-domain sequences with structurally corresponding residues in southern bean mosaic virus.     

The effect of metabolic acidosis on the synthesis and turnover of rat renal phosphate-dependent glutaminase.

Regulation of the mitochondrial phosphate-dependent glutaminase activity is an essential component in the control of renal ammoniagenesis. Alterations in acid-base balance significantly affect the amount of the glutaminase that is present in rat kidney, but not in brain or small intestine. The relative rates of glutaminase synthesis were determined by comparing the amount of [35S]methionine incorporated into specific immunoprecipitates with that incorporated into total protein. In a normal animal, the rate of glutaminase synthesis constitutes 0.04% of the total protein synthesis. After 7 days of metabolic acidosis, the renal glutaminase activity is increased to a value that is 5-fold greater than normal. During onset of acidosis, the relative rate of synthesis increases more rapidly than the appearance of increased glutaminase activity. The increased rate of synthesis reaches a plateau within 5 days at a value that is 5.3-fold greater than normal. Recovery from chronic acidosis causes a rapid decrease in the relative rate of glutaminase synthesis, but a gradual decrease in glutaminase activity. The former returns to normal within 2 days, whereas the latter requires 11 days. The apparent half-time for glutaminase degradation was found to be 5.1 days and 4.7 days for normal and acidotic rats respectively. These results indicate that the increase in renal glutaminase activity associated with metabolic acidosis is due primarily to an increase in its rate of synthesis. From the decrease in activity that occurs upon recovery from acidosis, the true half-life for the glutaminase was estimated to be 3 days.     

Flies as a source of enteric pathogens in a rural village in Thailand.

The village of Ban Pong in northeastern Thailand was studied from January through December 1981 to determine the importance of flies as a source of enteric pathogens. The number of flies (predominantly Musca domestica) increased in kitchens and animal pens in the hot dry spring, when the incidence of diarrhea was highest in the village. Enterotoxigenic Escherichia coli, Shigella spp., non-O1 Vibrio cholerae, and Vibrio fluvalis were isolated from fly pools in yards (69%), animal pens (38%), bathrooms (35%), and kitchens (8%). Enterotoxigenic E. coli was isolated from one fly pool in May and from another in June, when the incidence of such infections was highest in the village. Flies often carry and presumably disseminate enteric pathogens in rural Thailand.     

Structure of tomato busy stunt virus IV. The virus particle at 2.9 A resolution

The structure of tomato bushy stunt virus has been determined crystallographically to 2.9 A resolution. Details are presented of both the molecular structure and the methods by which it has been solved. The icosahedrally symmetric viral shell is composed of 180 protein subunits (Mr 43,000), with three similar but distinct modes of subunit bonding. This capacity for alternative packing is due to localized flexibility in the folded polypeptide (hinges between domains) and to multiple conformations for surface side-chains. The polypeptide backbone has an essentially invariant fold within a compact domain. A mechanism for correct positioning of the different modes of subunit interaction is evident from the structure of the TBSV particle. Thirty-five residues of the polypeptide chain fold in an ordered way on 60 of the 180 subunits, forming an internal framework. Interaction of folded domains with this framework permits accuracy of long-range geometry (correct curvature and closure) to be determined by unambiguous switching between alternative local contact angles. RNA packs tightly into the particle interior. Protein-RNA interactions occur through parts of the subunit that are flexibly linked to the well-ordered domains of the shell. This variable interaction imposes minimum restrictions on the folding of the RNA chain.     

Immunoreactive met-enkephalin in the canine adrenal; response to acute hypovolemic stress

Immunoreactive met-enkephalin was measured in the adrenal, kidney, liver, and intestine of the dog using radioimmunoassay. Adrenal tissue concentrations were 20 to 200-fold higher than the other tissues studied. In response to acute hypovolemic stress, the concentration of met-enkephalin in the adrenal vein of the dog increased 6-fold over basal peripheral arterial levels. These results suggest that the canine adrenal gland is a rich source of this opioid peptide and that the adrenal releases met-enkephalin in response to acute stress.     

Divalent cation sites in tomato bushy stunt virus. Difference maps at 2-9 A resolution

Difference electron density maps, using as few as four 1/2 degrees oscillation photographs, have been computed for tomato bushy stunt virus crystals soaked in EDTA. GdCl3 and silicotungstate. The maps define a double divalent cation site, responsible for regulating expansion of the virus particle, as well as sites for binding tungstate anions.