Hereditary respiration deficiency in Saccharomycodes ludwigii

Saccharomycodes ludwigii, supposed to be petite-negative, gave rise to respiration-deficient mutants when acriflavine and ultraviolet irradiation, respectively, were applied to this yeast, strain IFO 1194. The frequency of such mutants was very low as compared with that in Saccharomyces cerevisiae and other petite-positive yeasts. Cytochrome composition was characterized by spectrophotometry at the temperature of liquid nitrogen. The respiratory mutants examined contained cytochrome c unaltered in quality and quantity. Cytochrome b was often present only in small amounts though never absent, while cytochrome a+a₃ was either present or absent. The respiratory mutants could form zygotes after conjugation with a wild-type culture of opposite mating type ( vs. a). The hybridization and segregation analysis of spore tetrads showed the inheritance of respiratory mutant character to be either Mendelian or non-Mendelian and similar to that of pet (nuclear) and rho^- (cytoplasmic) mutants, respectively, in Saccharomyces cerevisiae.     

Genetic analysis of nurse dams selected for six-week body weight or postweaning gain in mice.

A modified crossfostering technique was developed to compare the performance of nurse dams in selected and control populations of mice. The H6 and M16 populations were selected for increased 6-week body weight and 3- to 6-week postweaning gain, respectively, while the C2 and ICR populations were the respective controls. Crossfostering was performed using H6, M16 and their reciprocal F1 crosses as nurse dams in the selected crossfostering group and C2 ICR and their reciprocals in the control group. Measurements recorded for nurse dams included mean body weight of 8 young within a nursed litter at birth (MWB) and 12 days of age (MW12). The latter was used as a measure of postnatal maternal performance. Other traits recorded for nurse dams were number born (NB), body weight at parturition (DWP) and 12 days postpartum (DW12), and weight gain (DWG), feed intake (FED) and efficiency (EFF = DWG/FED) for the first 12 days of lactation. The correlated response in MW12 was negative (P less than .01) for M16 and essentially zero for H6. Both lines exhibited positive (P less than .01) correlated responses in DWP and DW12 and no change in EFF. Only the H6 line increases significantly in DWG and FED as a result of selection. NB increased in M16 and H6, but was significant for the latter population only. Population differences in selection response [(M16-ICR)-(H6-C2)] were significant for FED only, primarily due to average direct genetic effects. Direct comparisons of M16 and H6 indicated that M16 was larger in DWP and DW12 but smaller in DWG and EFF. Average direct genetic effects favored M16 for NB, DWP, and DW12, whereas average maternal genetic effects favored H6 for NB, DWP, DW12 and FED. Percent direct heterosis, in F1 crosses of selected populations was significant for MW12 (13.7%) ,FED (10.8%) and NB (11.4%). Direct heterosis in F1 crosses of the controls was significant for MW12 (9.4%), NB (6.6%), DWP (3.5%), DW12 (3.3%) and FED (4.4%). The effects of MW12, DWG and metabolic body size (MBS) accounted for 47% of the variation in FED, pooled within populations. Of these variables, MW12 accounted for the highest proportion (32%) of variation in total feed intake.     

Studies on the circadian rhythm of phosphoenolpyruvate carboxykinase. III. Circadian rhythm in the kidney.

Phosphoenolypyruvate carboxykinase [EC 4.1.1.21] activity in rat kidney shows a circadian rhythm with the highest activity between 0200 h and 0800 h and the lowest activity between 1400 h and 2000 h. The rhythm was observed in both sexes and throughout the year. Actinomycin D and cycloheximide effectively blocked the circadian increase in enzyme activity. These findings suggest that the circadian increase in phosphoenolypyruvate carboxykinase activity is due to net synthesis of enzyme protein through newly synthesized mRNA. In experiments with kidney cortex slices, gluconeogenesis from the radioactive precursor, [14C]malic acid, was considerably higher at 0200 h than at 1400 h, varying in parallel with the change in the enzyme activity.     

Evidence for direct binding of Clostridium botulinum type E derivative toxin and its fragments to gangliosides and free fatty acids

Clostridium botulinum type E derivative toxin and its heavy chain bound to gangliosides GT1b, GD1a and GQ1b and saturated and unsaturated free fatty acids with chain lengths of 14-20 carbons. The L-H-1 fragment lacking the carboxyl-terminal portion of the heavy chain bound to free fatty acids but not to gangliosides. These observations led us to a new hypothesis on the mechanism of binding between botulinum toxin and gangliosides; the carboxyl-terminal portion (H-2 fragment) of the heavy chain binds to an oligosaccharide residue of gangliosides and then the amino-terminal portion (H-1 fragment) interacts with the hydrophobic portion of gangliosides consisting of fatty acids.     

Immunoelectronmicroscopic localization of extracellular matrix components produced by bovine corneal endothelial cells in vitro

Bovine corneal endothelial cells deposit an extracellular matrix in short-term cultures, which contains various morphologically distinct structures when analysed by electron microscopy after negative staining. Amongst these were long-spacing fibers with a 150 nm periodicity, which appeared also to be assembled into more complex hexagonal lattices. Another structure was fine filaments, 10-40 nm in diameter, which occasionally exhibited 67 nm periodic cross-striation. Non-striated 10-20 nm filaments sometimes formed radially oriented bundles arranged in networks and fuzzy granular material was associated with the filaments in the bundles. Often, these bundles extended into solitary filaments, 10-20 nm in diameter, with a smooth surface. In addition, amorphous patches were seen, which contained dense aggregates of fibrillar and granular material. In longer-term cultures, some of the structures coalesced to form large fibrillar bundles. By using specific antibodies to various extracellular matrix components and immunolabeling with gold some of these structures could be identified as to their protein composition. Whereas fibronectin antibodies labeled a variety of structures--fine filaments with granular materials, radially oriented bundles, patchy amorphous aggregates and small granular material scattered throughout the background--type III collagen antibody predominantly labeled filaments with periodic banding (10-40 nm in diameter). A small amount of type III specific labeling was also observed over the networks of radially oriented fibrils and fine filaments associated with granular material. Type IV collagen and laminin antibodies localized in areas of the patchy amorphous aggregates. Type VI collagen antibodies, on the other hand, labeled fine filaments and the gold particles showed a pattern of 100 nm periodicity. Many of the fine 10-20 nm filaments exhibited a tubular appearance on cross-section, but they were not reactive with any of the antibodies used. Also negative were the long-spacing fibers and assemblies--including hexagonal lattices--containing this structural element.     

A new approach to high sensitivity differential hybridization

We describe a new approach to differential hybridization, designed to identify cDNA clones representing rare mRNA species. Duplicate filters carrying a library of cDNA from phorbolmyristate acetate (PMA)-induced EL-4 cells in λgt11 were hybridized with high concentrations of unlabeled, cloned, single-stranded cDNA from induced and control EL-4 cells, respectively. Plaques binding single-stranded cDNA were revealed by a second round of hybridization with ³⁵S-labeled DNA complementary to the vector moiety of the single-stranded cDNA. Plaques corresponding to PMA-induced mRNAs occurring at a level of about 1 part in 15000 were isolated. We believe the method is at least ten times more sensitive than conventional differential hybridization.