Chloroplast and Cytoplasmic Ribosomes of Euglena: Selective Binding of Dihydrostreptomycin to Chloroplast Ribosomes

Dihydrostreptomycin binds preferentially to chloroplast ribosomes of wild-type Euglena gracilis Klebs var. bacillaris Pringsheim. The K(diss) for the wild-type chloroplast ribosome-dihydrostreptomycin complex is 2 x 10(-7) M, a value comparable with that found for the Escherichia coli ribosome-dihydrostreptomycin complex. Chloroplast ribosomes isolated from the streptomycin-resistant mutant Sm(1) (r)BNgL and cytoplasmic ribosomes from wild-type have a much lower affinity for the antibiotic. The K(diss) for the chloroplast ribosome-dihydrostreptomycin complex of Sm(1) (r) is 387 x 10(-7) M, and the value for the cytoplasmic ribosome-dihydrostreptomycin complex of the wild type is 1,400 x 10(-7) M. Streptomycin competes with dihydrostreptomycin for the chloroplast ribosome binding site, and preincubation of streptomycin with hydroxylamine prevents the binding of streptomycin to the chloroplast ribosome. These results indicate that the inhibition of chloroplast development and replication in Euglena by streptomycin and dihydrostreptomycin is related to the specific inhibition of protein synthesis on the chloroplast ribosomes of Euglena.     

Neurospora crassa Cytoplasmic Ribosomes: Isolation and Characterization of a Cold-Sensitive Mutant Defective in Ribosome Biosynthesis

Twenty-seven cold-sensitive mutants of Neurospora crassa were isolated by mutagenesis of wild-type conidia followed by filtration enrichment in complete medium at the nonpermissive temperature (10 C). Zone sedimentation analyses of cytoplasmic ribosomes isolated from the wild-type strain and from 14 of the mutant strains grown at 10 C indicate that one cold-sensitive mutant is defective in ribosome biosynthesis at that temperature: instead of the 2.3:1 mass ratio of 60S:37S ribosomal subunits characteristic of wild type, the mutant strain PJ30201 (called crib-1 for cytoplasmic ribosome biosynthesis) exhibits a mass ratio of approximately 7.2:1. Ribosomal subunits synthesized by strain PJ30201 at 25 C are present in wild-type proportions. The cold-sensitive and ribosomal phenotypes segregate together in tetrads isolated from crosses between strain PJ30201 and the wild type indicating that a single nuclear gene mutation is probably responsible for both mutant phenotypes. The crib-1 locus lies near the centromere in linkage group IV.     

Haem ligands of the ferricytochrome c of Ustilago sphaerogena

The mammalian-type cytochrome c of the basidiomycete Ustilago sphaerogena contains in a single polypeptide chain of 107 residues, two histidine residues located at positions 18 and 33, and one methionine residue situated at position 80 (Bitar et al., 1972). The reaction of Ustilago ferricytochrome c with bromoacetate at neutral pH resulted in the modification of histidine-33, but not of histidine-18 or of the invariant methionine residue. The activities of Ustilago cytochrome c with mitochondrial cytochrome c oxidase and with NADH-cytochrome c reductase were unaltered by the modification. The equilibrium constants for the formation of low-spin complexes of the ferrihaem octapeptide of horse cytochrome c (residues 14-21, including the haem bound covalently to cysteines 14 and 17) with imidazole, N(2)-acetylhistidine and monocarboxymethyl derivatives of N(2)-acetylhistidine were determined spectrophotometrically. Alkylation of the imidazole side-chain group of N(2)-acetylhistidine resulted in a marked decrease in its ability to form low-spin ferrihaem complexes. These results indicate that in Ustilago ferricytochrome c in solution histidine-33 is not involved in the central co-ordination complex. Since side-chain groups of residues other than histidine and methionine do not appear to be involved in the central complexes of other mammalian-type cytochromes c (Hettinger & Harbury, 1964, 1965; Myer & Harbury, 1965) it is likely that in Ustilago ferricytochrome c in solution at neutral pH, the side-chain groups of histidine-18 and methionine-80 are involved in the central co-ordination complex. The latter is stable over the pH range 2.6-8.4.     

Cultivation and Survival Studies of Neisseria gonorrhoeae in a Human Diploid Cell Strain

Neisseria gonorrhoeae was cultivated in a human diploid cell strain (WI-38). Eighty percent of the cultures contained viable gonococci for at least 4 m at 36°C, as evidenced by subculture to brain heart infusion broth. Monthly subcultures of bacteria could be made to fresh WI-38 cultures for at least 11 monthly passages with a 69% survival rate. The identity of gonococci was confirmed by morphology, gram staining, oxidase testing and fermentation reactions. Viability in brain heart infusion broth, minimum essential medium (Eagle), and WI-38 spent fluid was of much shorter duration. The organisms grown in WI-38 cultures appeared to orient largely in the vicinity of the WI-38 cells as well as within the cytoplasm of the cells.     

Recovery of the Ability to Synthesize DNA in Segments of Normal Size at Long Times After Ultraviolet Irradiation of Human Cells

DNA synthesized in human cells within the first hour after ultraviolet (UV) irradiation is made in segments of lower molecular weight than in nonirradiated cells. The size of these segments approximates the average distance between pyrimidine dimers in the parental DNA. This suggests that the dimers interrupt normal DNA synthesis and result in gaps in the newly synthesized DNA. However, DNA synthesized in human cells at long times after irradiation is made in segments equal or nearly equal to those synthesized by nonirradiated cells. The recovery of the ability to synthesize DNA in segments of normal size occurs in normal human cells, where the dimers are excised, and also in cells of the human mutants xeroderma pigmentosum (XP), where the dimers remain in the DNA. This observation implies that the pyrimidine dimer may not be the lesion that causes DNA to be synthesized in smaller than normal segments.     

The origin and structure of the secondary coat of the egg of Drosophila melanogaster

Summary The techniques of light and electron microscopy have been employed in a study of the protective coverings of the egg of Drosophila melanogaster. Data obtained during this investigation suggest the involvement of the follicle cells, in the production of one of these coverings and justify its classification as a secondary coat. The secondary coat of D. melanogaster is highly organized and has been divided into three Zones (I, II, IIII). The follicle cells enveloping the oocyte exhibit two phases of secretory activity each involving hypertrophy of the Golgi complex and rough endoplasmic reticulum, and the production of protein and polysaccharide components. The first phase concerns the elaboration of the material which gives rise to the homogeneous lamina referred to as Zone I. The second results in the release of an electron dense component which becomes organized into two laminae separated by struts or pillars; this construction is referred to as Zone II. At the completion of this secretory phase, the follicle cells assume a squamous morphology and a third Zone, composed of a homogeneous substance, appears between the follicle cells and Zone II.This investigation was supported by grant GM-08776 to one of us (EA) from the National Institutes of Health, United States Public Health Service.     

Value of Prospective Tissue Typing in Kidney Transplantation Between HL-A Identical Siblings

The predictability of leucocyte typing in kidney transplantation was assessed by an analysis of 37 kidney transplants from sibling donors. Recipients who were identical for the HL-A antigens with their donors gave highly predictable results. In comparison with those siblings who were incompatible or compatible but not identical their grafts functioned earlier, they required less immunosuppression, and had never had any rejections. They also appeared to have less postoperative morbidity. These results indicate that less immunosuppression than is current in many transplant centres could well be used with benefit in HL-A identical sibling transplants. This could reduce the risk of infection and possibly minimize the adverse effects of steroids on wound healing in these patients.     

LOCALIZATION OF A BASIC PROTEIN IN THE MYELIN OF VARIOUS SPECIES WITH THE AID OF FLUORESCENCE AND ELECTRON MICROSCOPY

In this study, alanine was shown to be the N-terminal amino acid of a basic protein of low molecular weight that was isolated from either human or guinea pig brain. Antibodies prepared against the guinea pig protein were labeled with either fluorescein or ferritin. Studies with the labeled antibodies showed that an immunohistochemically similar protein is found in the myelin sheaths of central and peripheral nervous tissues of chicken and frog and a variety of mammalian species. Loss of integrity of the myelin during processing was shown to enhance markedly the antigen-antibody reaction.