Distribution of mass in relaxed frog skeletal muscle and its redistribution upon activation.

Five orders of equatorial reflection were recorded from both relaxed and fully activated intact frog sartorius muscle using synchrotron x-ray radiation. Electron density maps of the myofilament lattice in axial projection were calculated from the integrated intensities by Fourier synthesis, using all possible phase combinations. These maps were evaluated systematically in terms of their compatibility with electron microscopically and biochemically derived properties of the lattice structure and with the minimum wavelength principle. For the relaxed state, one phase combination emerged as most consistent with these constraints: it shows a thick filament with a compact core surrounded by an annular shell of density. The distribution of mass suggests that the S-2 moiety of the myosin molecule is an integral part of the thick-filament backbone and the S-1 moiety makes up the shell and is tilted or slewed around the backbone. For the active state, there are two feasible maps, which differ according to whether or not the activation process is associated with phase inversion in two of the reflections. Both maps represent patterns of redistribution of mass upon activation in which the thick-filament backbone is practically unaffected and there is movement of density from the annular shell towards the thin filaments. In addition to this outward radial flux of density from the thick-filament periphery, the pattern of net mass transfer involves a pronounced azimuthal component in both cases. The total net mass transfer is equivalent to approximately 20% (no phase change) or approximately 40% (with phase change) of the S-1 mass. From the observed systematic increase in peak widths of the higher orders, the size of the crystalline domain in the myofilament lattice in the relaxed sartorius is estimated to be greater than 650 nm and the variations in myofilament lattice spacing among different myofibrils to be about +/- 3%. Furthermore, in the activated state, the equilibrium positions of the myofilaments are no longer well ordered, but are distributed statistically about the lattice points with a standard deviation of approximately 3 nm.     

Alterations in cholesterol and fatty acid biosynthesis in rat liver homogenates by aryloxy acids (Short Communication)

2,4-Dichlorophenoxyacetic acid and 2,4,5-trichlorophenoxyacetic acid inhibited the incorporation of [2-(14)C]mevalonate into cholesterol and non-saponifiable lipids. Both compounds inhibited the conversion of [1-(14)C]isopentenyl pyrophosphate into cholesterol and the synthesis of cholesterol and fatty acids from [2-(14)C]acetate. There was no inhibition of the conversion of [1-(14)C]mevalonate into CO(2). At low concentrations (0.5mm) of the compounds there was a stimulation of acetate incorporation into fatty acids.     

Purification and characterization of bovine aorta ribonucleases

1. Two ribonucleases (aorta ribonuclease I and aorta ribonuclease II) from bovine aorta were purified 4611-fold and 667-fold respectively. Ethanolic precipitation, acid extraction, isoionic precipitation at pH3.5 and Bio-Rex 70 column chromatography were the methods employed. 2. Aorta ribonuclease I exhibited no deoxyribonuclease or alkaline phosphatase activity. 3. Aorta ribonuclease I appeared to be homogeneous when subjected to discontinuous gel electrophoresis. 4. Aorta ribonuclease II exhibited the same properties as aorta ribonuclease previously isolated. 5. The activities of the aorta ribonucleases and pancreatic ribonuclease on homopolymers and dinucleoside phosphates were compared. 6. Aorta ribonuclease I exhibited optimum pH7.5 and, under the assay conditions used, optimum temperature 60 degrees .     

Viral Antibodies in Diabetes Mellitus

Sera from 123 patients with diabetes mellitus of recent onset, 155 patients with diabetes of more than two years'' duration, and 250 normal persons were collected over a period of two and a half years. All sera were tested for neutralizing antibody to Coxsackie virus types B1–6, and a sample was tested for complement-fixing antibody to a number of viral, rickettsial, and mycoplasmal antigens.In diabetics of recent onset no evidence was found of any excess of antibodies to mumps virus or some common respiratory viruses. Insulin-dependent diabetes within three months of onset were found to have higher antibody titres to Coxsackie B virus, particularly of type B4, than either normal subjects or patients with diabetes of longer duration.     

Translation and stability of proteins encoded by the plastid psbA and psbB genes are regulated by a nuclear gene during light-induced chloroplast development in barley

We have characterized a nuclear mutant of barley, viridis-115, lacking photosystem II (PSII) activity and compared it to wild-type seedlings during light-induced chloroplast development. Chloroplasts isolated from wild-type and viridis-115 seedlings illuminated for 1 h synthesized similar polypeptides and had similar protein composition. After 16 h of illumination, however, mutant plastids exhibited reduced ability to radiolabel D1, CP47, and several low Mr membrane polypeptides, and by 72 h, synthesis of these proteins was undetectable. Immunoblot analysis showed that plastids of dark-grown wild-type barley lacked several PSII proteins (D1, D2, CP47, and CP43) and that 16 h of illumination resulted in the accumulation of these polypeptides. In contrast, these polypeptides did not accumulate in illuminated viridis-115 seedlings, although mutant plastids accumulated two PSII proteins that participate in oxygen evolution, oxygen-evolving enhancers 1 and 3. Northern analysis showed that the levels of psbA and psbB mRNA in mutant plastids were equal to or greater than levels in wild-type plastids throughout the developmental period examined here. These results indicate that the nuclear mutation present in viridis-115 affects the translation and stability of the chloroplast-encoded D1 and CP47 polypeptides and that its influence is expressed after the onset of light-induced chloroplast development.     

Localization of sequences regulating ancestral and acquired sites of esterase6 activity in Drosophila melanogaster

We have broadly defined the DNA regions regulating esterase6 activity in several life stages and tissue types of D. melanogaster using P- element-mediated transformation of constructs that contain the esterase6 coding region and deletions or substitutions in 5' or 3' flanking DNA. Hemolymph is a conserved ancestral site of EST6 activity in Drosophila and the primary sequences regulating its activity lie between -171 and -25 bp relative to the translation initiation site: deletion of these sequences decrease activity approximately 20-fold. Hemolymph activity is also modulated by four other DNA regions, three of which lie 5' and one of which lies 3' of the coding region. Of these, two have positive and two have negative effects, each of approximately twofold. Esterase6 activity is present also in two male reproductive tract tissues; the ejaculatory bulb, which is another ancestral activity site, and the ejaculatory duct, which is a recently acquired site within the melanogaster species subgroup. Activities in these tissues are at least in part independently regulated: activity in the ejaculatory bulb is conferred by sequences between -273 and -172 bp (threefold decrease when deleted), while activity in the ejaculatory duct is conferred by more distal sequences between -844 and -614 bp (fourfold decrease when deleted). The reproductive tract activity is further modulated by two additional DNA regions, one in 5' DNA (-613 to -284 bp; threefold decrease when deleted) and the other in 3' DNA (+1860 to +2731 bp; threefold decrease when deleted) that probably overlaps the adjacent esteraseP gene. Collating these data with previous studies suggests that expression of EST6 in the ancestral sites is mainly regulated by conserved proximal sequences while more variable distal sequences regulate expression in the acquired ejaculatory duct site.      

Use of tumour marker immunoreactivity to identify primary site of metastatic cancer.

OBJECTIVES--To determine whether variations in the expression of tumour related antigens can predict the origin of tumours. DESIGN--Immunohistological study of tumour marker expression in primary adenocarcinomas and respective metastatic deposits. Antibodies to the following tumour markers were used: polymorphic epithelial mucin (NCRC-11 and SM3), carcinoembryonic antigen, carcinoembryonic antigen with non-specific antigen co-specificity, CA125, CA19.9, prostate specific antigens, and thyroglobulin. SETTING--Histopathology department of teaching hospital. SUBJECTS--100 pathology sections of metastatic adenocarcinoma and their related primary tumours. MAIN OUTCOME MEASURES--Concordance of reactivity between primary and metastatic tumours. Reactivity profiles of tumour sites. RESULTS--The correct primary site of origin was predicted in 70% (33/47) of tumours in men and 54% (27/43) tumours in women with antibodies SM3, 288, CA19.9, CA125, and PSA (men only). Specificities ranged from 68% for breast tumour to 98% for prostate tumour. CONCLUSION--Use of tumour markers in patients presenting with metastatic adenocarcinoma of unknown origin can help localise the probable primary sites and reduce the need for extensive and expensive imaging techniques.