Genetically Induced Mitotic Exchange in the Heterochromatin of DROSOPHILA MELANOGASTER

Multiple copies of the 18S and 28S ribosomal RNA cistrons are present in both the X and Y chromosomes of Drosophila melanogaster. Data are presented here that identify a locus, Rex, that causes exchange-like events between duplicated ribosomal complexes at the ends of an attached-XY chromosome. Rex: (1) is close to or in the basal heterochromatin of the X chromosome; (2) is semidominant and (its effect) is temperature sensitive; (3) acts maternally; and (4) affects behavior of paternally derived attached-XY chromosomes shortly after fertilization. Though, at this point, the existence of Rex is known only from its effects on behavior of a particular compound chromosome, it presents intriguing possibilities for understanding regulation of chromosome behavior and organization of the ribosomal cistrons.     

Uptake and metabolism of thyroid hormones by cultured monkey hepatocarcinoma cells. Effects of potassium cyanide and dinitrophenol

Cultured monkey hepatocarcinoma cell (NCLP-6E) were used to investigate the uptake and metabolism of thyroid hormones. Intracellular accumulation was shown by the failure to acutely release hormone from cells subsequently exposed to serum proteins, and by the metabolic trnasformation of the hormones to deiodinated products and their sulfates. When hepatocarcinoma cell monolayers were studied at hormone concentrations below 10(-10) M, neither KCN nor dinitrophenol inhibited uptake. Taken together with previous findings that uptake was neither saturable nor reduced at low temperature, these results indicate that this process was not active transport. Deiodination of both the phenolic and non-phenolic rings, however, was partially inhibited by KCN but not by dinitrophenol. Sulfation of 3,3'-diiodothyronine and 3'-monoiodothyronine was strongly inhibited by both KCN and dinitrophenol. Uptake of the hormones and their metabolites was also measured in suspended hepatocarcinoma cells and compared with the uptake by normal rat hepatocytes, human fibroblasts and human lymphocytes. In these experiments 1 micrometer triidothyronine and 0.47 mM dinitrophenol were used to inhibit deiodination and sulfation, respectively. Uptake was similar in all cell types. Accumulation was highest with 3,5,3'-triiodothyronine, intermediate with other compounds having iodines in both rings, lowest with compounds iodinated in only one ring, and absent with iodothronine sulfates. These findings help to explain the relative rates of metabolism of the iodothyronines and their release from the cells.     

Synthesis of glycolipids and phospholipids in hamster cells: dependence on cell density and the cell cycle.

Sakiyama et al. ('72) reported the isolation of a line of hamster cells (NIL 1c1) which contains only three glycolipids, hematoside, ceramide monohexoside and ceramide dihexoside. The incorporation of radiolabeled palmitate into hematoside during 24 hours was three fold higher in normal confluent, non growing cells than sparse, growing ones. Polyoma transformed cells did not exhibit this effect. We have continued studies with the untransformed cell line and have found that the higher incorporation of radiolabeled palmitate into hematoside by normal confluent cells is not due to a higher rate of turnover of hematoside at confluence but represents a true chemical increase. We have also found that this increase is not a gradual process during cell growth but instead occurs only when the cells become confluent and stop growing. The increase of hematoside at confluence is not due to a higher rate of synthesis of hematoside during G1, relative to the other phases of the cell cycle. We found the rate of synthesis of hematoside to be constant throughout the cell cycle. The rate of synthesis of phosphatidyl choline, phosphatidyl ethanolamine, phosphatidyl serine, phosphatidyl inositol and sphingomyelin was also studied as a function of the cell cycle. We found no large differences in the synthetic rate of any given phospholipid species throughout the cell cycle although the rate of synthesis of the glycerophospholipids was somewhat higher during late G1 and S. We did, however, find major differences in the rates of synthesis of the different phospholipid species.     

Regulation of the Escherichia coli methylgalactoside transport system by gene mglD.

Constitutive activity of the methylgalactoside transport system of Escherichia coli K-12 is shown to result from mutation of a genetic locus distinct from the two previously described regulatory loci for this permease. Employing an autoradiographic procedure whereby constitutive and inducible cells can be differentiated, it is demonstrated that this locus, termed mglD, is 20% cotransducible with ptsF by bacteriophage P1. Selection for constitutive mutants among an inducible population yielded cells who mutations mapped in mglD. Cotransduction of mglD with mglB, minus C, and minus A, three genes required for activity of the methylgalactoside transport system, is 95, 88, and 81%, respectively. The results of recombination studies employing three and four factors indicate that the order of genes in this region is ptsF, mglD, B, C, A.     

Isolation of a human plasmin-derived, functionally active, light (B) chain capable of forming with streptokinase an equimolar light (B) chain-streptokinase complex with plasminogen activator activity.

A functionally active human plasmin light (B) chain derivative, stabilized by the streptomyces plasmin inhibitor leupeptin, was isolated from a partially reduced and alkylated enzyme preparation by an affinity chromatography method with a L-lysine-substituted Sepharose column. This light (B) chain derivative was found to be relatively homogeneous by electrophoretic analysis in both an acrylamide gel/dodecyl sulfate system and on cellulose acetate. It possessed approximately 3% of the proteolytic activity (casein substrate) of the original enzyme, and it incorporated 0.09 mol of [3H]diisopropyl phosphorofluoridate per mol of protein. It contained 3.1 +/- 0.3 carboxymethylated cysteines per mol of protein and can be designated as a CmCys5-light (B) chain (CmCys)3. When this isolated light (B) chain derivative was mixed in equal molar amounts with streptokinase, the mixture developed both human and bovine plasminogen activator activities; the bovine activator activity was approximately 66% of the bovine activator activity of the equimolar human plasmin-streptokinase complex. Although this complex now incorporated 0.50 mol of [3H]diisopropyl phosphorofluoridate per mol of protein, its proteolytic activity, on a molar basis, was the same as the proteolytic activity of the isolated light (B) chain derivative. It was shown by electrophoretic analysis in both an acrylamide gel/epsilon-aminocaproic acid system and on cellulose acetate that the light (B) chain derivative and streptokinase forms an equimolar light (B) chain-streptokinase complex, indicating that the binding site for streptokinase is located on the light (B) chain of the enzyme. A functionally active equimolar light (B) chain-streptokinase complex was also isolated from a partially reduced and alkylated equimolar human plasmin-streptokinase complex by the affinity chromatography method. The plasminogen activator activities (human and bovine) of this light (B) chain-streptokinase complex were similar to those of the plasmin-streptokinase complex from which it was derived. Although this complex incorporated 0.70 mol of [3H]diisopropyl phosphorofluoridate per mol of protein, its proteolytic activity, on a molar basis, was only 14% of proteolytic activity of the plasmin-streptokinase complex.     

High-pressure liquid-solid chromatography of the ecdysones--insect molting hormones

A rapid, high-pressure liquid-solid Chromatographie technique was developed for the separation, detection and analysis of ecdy-steroids. The advantages in using this system for the identification of ecdysteroids in combination with NMR and mass spectrometry and its potential application in analyzing material from biological and synthetic, systems are presented.     

ISOELECTRIC POINTS FOR THE MYCELIUM OF FUNGI

1. Mycelium of Rhizopus nigricans when stained with certain acid and basic dyes and washed with buffer mixtures of 0.1 M phosphoric acid and sodium hydroxide responded much like an amphoteric colloid with an isoelectric point near pH 5.0. 2. When grown on potato dextrose agar the reaction of which was varied with phosphoric acid the extent of colony growth of Rhizopus nigricans plotted against the initial Sörensen value of the agar produced a double maximum curve with the minimum between the two maxima at initial pH 5.2. 3. When grown in potato dextrose broth the reaction of which was varied with phosphoric acid the dry matter produced by Rhizopus nigricans plotted against the Sörensen value of the broth produced a double maximum curve with the minimum between the two maxima at initial pH 5.2 or average pH 4.9. 4. Mycelium of Rhizopus nigricans placed in buffer mixtures of 0.01 M phosphoric acid and sodium hydroxide of pH 4.1 to 6.3, changed the reaction in most cases toward greater alkalinity. 5. Mycelium of Fusarium lycopersici stained with certain acid and basic dyes and washed with buffer mixtures of 0.1 M phosphoric acid and sodium hydroxide responded much like an amphoteric colloid with an isoelectric point near pH 5.5.