Cardiolipin biosynthesis and remodeling enzymes are altered during development of heart failure

Cardiolipin (CL) is responsible for modulation of activities of various enzymes involved in oxidative phosphorylation. Although energy production decreases in heart failure (HF), regulation of cardiolipin during HF development is unknown. Enzymes involved in cardiac cardiolipin synthesis and remodeling were studied in spontaneously hypertensive HF (SHHF) rats, explanted hearts from human HF patients, and nonfailing Sprague Dawley (SD) rats. The biosynthetic enzymes cytidinediphosphatediacylglycerol synthetase (CDS), phosphatidylglycerolphosphate synthase (PGPS) and cardiolipin synthase (CLS) were investigated. Mitochondrial CDS activity and CDS-1 mRNA increased in HF whereas CDS-2 mRNA in SHHF and humans, not in SD rats, decreased. PGPS activity, but not mRNA, increased in SHHF. CLS activity and mRNA decreased in SHHF, but mRNA was not significantly altered in humans. Cardiolipin remodeling enzymes, monolysocardiolipin acyltransferase (MLCL AT) and tafazzin, showed variable changes during HF. MLCL AT activity increased in SHHF. Tafazzin mRNA decreased in SHHF and human HF, but not in SD rats. The gene expression of acyl-CoA: lysocardiolipin acyltransferase-1, an endoplasmic reticulum MLCL AT, remained unaltered in SHHF rats. The results provide mechanisms whereby both cardiolipin biosynthesis and remodeling are altered during HF. Increases in CDS-1, PGPS, and MLCL AT suggest compensatory mechanisms during the development of HF. Human and SD data imply that similar trends may occur in human HF, but not during nonpathological aging, consistent with previous cardiolipin studies.     

Structure of seasonal migrations of the littoral hermit crab Clibanarius vittatus (Bosc)

A seasonal migration in the hermit crab Clibanarius vittatus (Bosc) is described and its structure examined for size-related or sexual differences that might lead to differential mortalities. The largest crabs, which were nearly all males, began to leave the shore in early summer, but the smaller females stayed until late autumn. The smaller crabs also returned to the shore at least one month in advance of the large crabs in the spring. No size-related or sexual differences were found in the abilities of the crabs to regulate their volumes when moved from 34.5%. to 10.5%. salinity which is the observed salinity range in their usual habitat.     

Eosin B

The two known isomeric dibromodinitrofiuoresceins have been prepared in a fairly pure state, and their absorption spectra determined.

Commercial samples of eosin B are not 4, 5-dibromo—2, 7-dinitrofluorescein, as stated in dye indices. They are mixtures which contain other bromonitro derivatives of fluorescein as well as di-bromodinitro derivatives.

The color acid method provides a substantially reliable means of determining actual dye content with commercial samples of the dye, but the reduction method may prove decidedly misleading.     

Alcohol dehydrogenase isozymes in the mouse: Genetic regulation, allelic variation among inbred strains and sex differences of liver and kidney A2 isozyme activity

Genetic analysis of a proposed cis-acting temporal locus ( Adh-3t ), which regulates alcohol dehydrogenase C₂ (ADH-C₂) acitivity in mouse epididymis extracts, among F₁ (ddN × BALB/c) × ddN male backcross progeny provided evidence for genetic distinctness between the structural ( Adh-3 ) and temporal ( Adh-3t ) loci on chromosome 3. Genetic analysis also confirmed the close, linkage of Adh-1 (encoding liver and kidney ADH-A₂) and Adh-3 (encoding stomach ADH-C₂) to within 0.3 centimorgans on the mouse genome. Evidence is presented for a proposed closely linked cis-acting temporal locus (designated Adh-1t ) for the A₂ isozyme (encoded by Adh-1 ) controlling the activity of this enzyme in mouse kidney extracts, but having no apparent affect on liver and intestine ADH-A₂ activities. An extensive survey of the distribution of Adh-1, Adh-3 and Adh-3t alleles among 65 strains of mice is reported — with the exception of two Japanese strains (ddN and KF), linkage disequilibrium between Adh-3 and Adh-3t was observed. Sex differences in mouse liver and kidney ADH-A₂ activities were observed, with male/female ratios of approximately 0.6 and 3 respectively for these tissue extracts.     

Photoaffinity labeling of GDP-fucose:nLcOse4Cer alpha 1----3-fucosyltransferase from human small cell lung carcinoma NCI-H69 cells with the GDP-fucose analog GDP-hexanolaminyl-4-azidosalicylic acid

An iodinatable photoactive analog of GDP-fucose, GDP-hexanolaminyl-4-azidosalicylic acid, has been prepared and applied to studies of the previously described alpha 1----3-fucosyltransferase from NCI-H69 cells (Holmes, E. H., Ostrander, G. K., and Hakomori, S. (1985) J. Biol. Chem. 260, 7619-7627). The NCI-H69 cell alpha 1----3-fucosyltransferase was obtained from a 0.2% Triton X-100-solubilized enzyme fraction after affinity purification on a GDP-hexanolamine-Sepharose column and gel filtration through a fast protein liquid chromatography Superose 12 column. Increasing concentrations of the photoaffinity reagent were found to result in loss of up to 35% of the original enzyme activity at under 100 microM final concentrations. The inactivation was photolysis dependent and could be prevented by the addition of GDP-fucose prior to photolysis. The photoprobe behaved as a competitive inhibitor with respect to GDP-fucose with a Ki of 23 microM, identical to that of GDP. Photoincorporation of 125I-labeled GDP-hexanolaminyl-4-azidosalicylic acid into the enzyme fraction labeled a slow migrating protein band in a native polyacrylamide gel which corresponded to enzyme activity. Inclusion of GDP-fucose prevented photolabeling of this band. Sodium dodecyl sulfate gel electrophoresis of the photolabeled, GDP-fucose-protected band yielded a 125I-labeled protein band that migrated at Mr 45,000, most probably corresponding to an alpha 1----3-fucosyltransferase protein subunit. These studies suggest photoaffinity labeling using nucleotide affinity ligands linked to photoactivatable, heterobifunctional cross-linking reagents may be generally applicable to photoaffinity labeling glycosyltransferase enzyme proteins.     

Mammalian Fbh1 is important to restore normal mitotic progression following decatenation stress

We have addressed the role of the F-box helicase 1 (Fbh1) protein during genome maintenance in mammalian cells. For this, we generated two mouse embryonic stem cell lines deficient for Fbh1: one with a homozygous deletion of the N-terminal F-box domain (Fbh1^f/f), and the other with a homozygous disruption (Fbh1^?/?). Consistent with previous reports of Fbh1-deficiency in vertebrate cells, we found that Fbh1^?/? cells show a moderate increase in Rad51 localization to DNA damage, but no clear defect in chromosome break repair. In contrast, we found that Fbh1^f/f cells show a decrease in Rad51 localization to DNA damage and increased cytoplasmic localization of Rad51. However, these Fbh1^f/f cells show no clear defects in chromosome break repair. Since some Rad51 partners and F-box-associated proteins (Skp1-Cul1) have been implicated in progression through mitosis, we considered whether Fbh1 might play a role in this process. To test this hypothesis, we disrupted mitosis using catalytic topoisomerase II inhibitors (bisdioxopiperazines), which inhibit chromosome decatenation. We found that both Fbh1^f/f and Fbh1^?/? cells show hypersensitivity to topoisomerase II catalytic inhibitors, even though the degree of decatenation stress was not affected. Furthermore, following topoisomerase II catalytic inhibition, both Fbh1-deficient cell lines show substantial defects in anaphase separation of chromosomes. These results indicate that Fbh1 is important for restoration of normal mitotic progression following decatenation stress.