Role of nuclear proteins on [H]actinomycin D binding during lymphocyte mitogenesis

The uptake of [³H]actinomycin D ([³H]AD) by ConA-stimulated lymphocytes was followed during 96 h of incubation and correlated with the level of nuclear proteins in the nucleus, DNA synthesis and the degree of AD-induced inhibition of RNA and DNA synthesis. During the first 48 h there is a parallel increase of drug binding to cells and a rising level of non-histone proteins (NHP) in the nucleus. During the next 48 h, DNA synthesis occurs, drug uptake decreases and the nuclear level of NHP continues to rise. The level of histones remains constant during 96 h. The variations in cellular [³H]AD uptake during 96 h are not due to changes in cell membrane permeability, since similar variations in drug binding are observed in isolated cell nuclei. NHP, obtained as 0.25 M NaCl extracts of cell nuclei, increase binding of [³H]AD to nuclei isolated from non-stimulated lymphocytes, while histones have no such effect. NHP extracted with phenol, after washing the nuclei with salt and acid solutions, or extracted with 0.25 M NaCl from non-stimulated and stimulated lymphocytes and Chang liver cells are equally active to bind [³H]AD to nuclei of non-stimulated lymphocytes. NHP from Chang cells, purified by DNA-cellulose chromatography using calf thymus DNA, stimulated [³H]AD binding to lymphocyte nuclei, indicating that the drug-binding activity is due to proteins binding to DNA. NHP increase binding of [³H]AD to pure DNA in the absence of histones. The degree of [³H]AD binding to ConA-stimulated lymphocytes during 96 h correlated with the degree of inhibition of RNA and DNA synthesis by AD.     

Carotenoids of Phaffia rhodozyma, a red-pigmented fermenting yeast

The red-pigmented fermenting yeast Phaffia rhodozyma contains astaxanthin as the principal carotenoid pigment. Echinenone, 3-hydroxyechinenone and phoenicoxanthin were also isolated and identified; isocryptoxanthin and canthaxanthin were absent. Evidence is presented for a new carotenoid, 3-hydroxy-3′4′-didehydro-β,ψ-caroten-4-one. A possible biosynthetic scheme for the formation of astaxanthin in P. rhodozyma is suggested.     

On the detection of functional and structural enzyme mutants by coordinated affinity chromatography and isoelectric focusing

A method of studying structural and functional heterogeneity of enzymes has been developed and tested on chymotrypsin. The enzyme, prepared from single mouse pancreata, has been fractionated with respect to function and charge content by a combination of affinity chromatography and isoelectric focusing. By comparing chymotrypsin isolated from isogenic strains, chymotrypsinogen of strains A/Sn and NZB was found to be genetically heterogeneous, thus not revealed as different chymotrypsin forms of a single zymogen. Chymotrypsinogen originating from two loci was investigated, and structural and functional differences of the corresponding enzymes were determined. At both loci, structural allelomorphism was indicated. At one locus, the structural heterogeneity was also found to be reflected in functional heterogeneity of the corresponding enzymes. By mating the two strains and fractionating the enzyme of the cross, the differences were shown to be inherited.     

Flow microfluorometric system for screening gynecologic cytology specimens using propidium iodide-fluorescein isothiocyanate.

Seventy cervical cytology specimens have been screened by a xero resolution flow analyzer-sorter using propidium iodide and fluorescein isothiocyanate as fluorochromes for nucleus and cytoplasm, respectively. This system shows a 1% sensitivity for detection of abnormal cells using only crude visual data analysis. Screening of clinical specimens was performed on the instrument with a 5.8% false negative rate and a 11.8% false positive rate by comparison with routine visual cytologic evaluation of the same samples.     

Ecdysterone antagonism,mimicry, and synergism of juvenile hormone action on the Monarch butterfly reproductive tract

Previous research on Monarch butterflies has shown that juvenile hormone (JH) stimulates the development of the ovary and certain reproductive glands of both sexes. Ecdysterone injections into intact Monarchs demonstrate that low doses of this hormone inhibit ovarian development, and higher doses stimulate the male and female reproductive glands. In addition, experiments using neckligatured adults show that ecdysterone stimulates the reproductive glands of both sexes, in the apparent absence of JH, with the most pronounced effect being observed on the female colleterial gland. Other studies with neck-ligatured animals demonstrate that ecdysterone also synergizes with JH on the female gland and all three male glands. The feasibility of using Monarch reproductive glands for studies on the mode of action and interaction of JH and ecdysterone, and the possibility of a rôle of ecdysterone in the normal regulation of Monarch oögenesis, are discussed.     

CACHONINA ILLDEFINA SP. NOV. (DINOPHYCEAE): CHLOROPLAST TUBULES AND DEGENERATION OF THE PYRENOID1

A new species of the dinoflagellate genus Cachonina, C. illdefina sp. nov., was isolated from a red tide off El Capitan State Park, Santa Barbara County, California, in October 1973. The organism is light yellowgreen in color with deeply incised girdle and sulcal grooves. Electron microscopy of the organism, revealed a typical dinokaryotic nucleus. The chloroplasts of the organism are connected, and often contain microtubule-like elements, 25 nm diam. The pyrenoids are characterized as excluding chloroplast thylakoids and ribosomes, although containing an amorphous matrix and numerous tubular invaginations from the cytoplasm. The pyrenoids become detached from the chloroplasts and degenerate into small vesicles. C. illdefina is not bioluminescent.     

Amino-acyl tXNA as inhibitors or amino acid donors in peptide synthesis

Xenobiotic nucleic acids (XNAs) offer tremendous potential for synthetic biology, biotechnology, and molecular medicine but their ability to mimic nucleic acids still needs to be explored. Here, to study the ability of XNA oligonucleotides to mimic tRNA, we synthesized three L-Ala-tXNAs analogs. These molecules were used in a non-ribosomal peptide synthesis involving a bacterial Fem transferase. We compared the ability of this enzyme to use amino-acyl tXNAs containing 1′,5′-anhydrohexitol (HNA), 2′-fluoro ribose (2′F-RNA) and 2′-fluoro arabinose. L-Ala-tXNA containing HNA or 2′F-RNA were substrates of the Fem enzyme. The synthesis of peptidyl-XNA and the resolution of their structures in complex with the enzyme show the impact of the XNA on protein binding. For the first time we describe functional tXNA in an in vitro assay. These results invite to test tXNA also as substitute for tRNA in translation.