Streptomycin retards the phenotypic maturation of chick myogenic cells

Summary As part of an effort to optimize conditions required for the complete maturation of muscle cells in vitro, we have investigated the effects of the antibiotics penicillin, streptomycin, and Fungizone (amphotericin B) on the development of cultured chick embryo skeletal muscle. It is shown that even low dosages of streptomycin, but not penicillin or Fungizone, retard protein synthesis and accumulation in these cultures. Myosin accumulation was also reduced and the appearance of striations in fused cells was delayed in myotubes formed in medium containing streptomycin. Additional data suggest that this overall retardation of myogenesis is due to the influence of streptomycin on maturing myotubes rather than early proliferation and cell fusion. These results are discussed with regard to recent efforts to promote the full maturation of muscle cells grown in culture. This research was supported by National Institutes of Health Grant NS 155882 and a Task Force on Drug Development Research Contract from The Muscular Dystrophy Association.     

Mutant hemoglobin stability depends upon location and nature of single point mutation

The temperature dependence of the rates of heme release from the beta subunits of methemoglobin A and 5 beta mutant methemoglobins has been determined. The rates were largest for two hemoglobins with mutations distal to heme, previously known to be unstable. The other 3 mutants also released heme faster than A. These hemoglobins, with single point mutations at the alpha 1/beta 2 interface, were previously thought to be stable. The low reported yields of the 5 mutant proteins covaries with the relative rates of heme release from the met species.     

Measurement of diameters of estuarine bacteria and particulates in natural water samples by use of a submicron particle analyzer

Desulfovibrio vulgaris strain Madison outcompetedMethanobacterium strain ivanov for hydrogen when sulfate was in excess because of higher cell yield and growth rate and a greater affinity for hydrogen as a consequence of a lower Km and higher Vmax for in vivo hydrogenase activity.Desulfovibrio vulgaris displayed a growth yield of 1.1 g/mol H₂, a Km for tritium exchange of 4 M, and a specific in vivo hydrogenase activity of 2.17 DPM³H₂O×10³/g cell protein/h; whereasMethanobacterium strain ivanov had a yield of 0.6 g/mol H₂, a Km for tritium exchange of 14 M, and a specific in vivo hydrogenase activity of 0.38 DPM³H₂O×10³/g cell protein/h. Under these physiological conditions, the Gibbs free-energy change associated with methanogenesis and sulfidogenesis from H₂ was calculated to be-47.4 kJ/mol and-62.9 kJ/mol, respectively. When sulfidogenesis was limited by sulfate concentration, the methanogen was able to successfully compete with the sulfidogen for hydrogen. Competition between methanogens and sulfidogens for hydrogen is explained in terms of thermodynamic, kinetic, and other important considerations not discussed in the previous literature.     

Structure and enzymic activity of ribonuclease-A esterified at glutamic acid-49 and aspartic acid-53

The dimethyl ester of bovine pancreatic ribonuclease-A (dimethyl RNAase-A), the initial product of esterification of RNAase-A in anhydrous methanolic HCl, was isolated in a homogeneous form. The two carboxy functions esterified in this derivative are those of glutamic acid-49 and aspartic acid-53. There were no changes in the u.v.-absorption spectral characteristics, the accessibility of the methionine residues, the resistance of the protein to proteolysis by trypsin and the antigenic behaviour of RNAase-A as a result of the esterification of these two carboxy groups. Dimethyl RNAase-A exhibited only 65% of the specific activity of RNAase-A, but still had the same K_m value for both RNA and 2′:3′-cyclic CMP. However, the V_max. was decreased by about 35%. On careful hydrolysis of the methyl ester groups at pH9.5, dimethyl RNAase-A was converted back into RNAase-A. Limited proteolysis of dimethyl RNAase-A by subtilisin resulted in the formation of an active RNAase-S-type derivative, namely dimethyl RNAase-S, which was chromatographically distinct from dimethyl RNAase-A and had very nearly the same enzymic activity as dimethyl RNAase-A. Fractionation of dimethyl RNAase-S by trichloroacetic acid yielded dimethyl RNAase-S-protein and dimethyl RNAase-S-peptide, both of which were inactive by themselves but regenerated dimethyl RNAase-S when mixed together. Dimethyl RNAase-A-peptide was identical with RNAase-S-peptide. RNAase-S-protein could be generated from dimethyl RNAase-S-protein by careful hydrolysis of the methyl ester groups at pH9.5. The interaction of dimethyl RNAase-S-protein with RNAase-S-peptide appears to be about 4-fold weaker than that between the RNAase-S-protein and RNAase-S-peptide. Conceivably, the binding of the S-peptide `tail' of dimethyl RNAase-A with the remainder of the molecule is similarly weaker than that in RNAase-A, and this brings about subtle changes in the geometrical orientation of the active-site amino acid residues of these modified methyl ester derivatives. It is suggested that these changes could be responsible for the generation of the catalytically less-efficient RNAase-A and RNAase-S molecules (dimethyl RNAase-A and dimethyl RNAase-S respectively).     

Effect of ultraviolet radiation on the microsporidian Octosporea muscaedomesticae with reference to protectants provided by the host Phormia regina

The effect of ultraviolet light on the microsporidian Octosporea muscaedomesticae in relation to infection in the adult black blowfly, Phormia regina, was investigated. A 30-Watt germicidal lamp, 253.7-nm wavelength, was used as source of uv light in five investigations. In addition, sunlight served as a uv source in two studies. Viable naked dried spores exposed to the uv lamp at a distance of 10 cm were killed after 15 min. Viable naked spores in an aqueous suspension were killed after 30 min of exposure to the uv lamp and after 3 hr of exposure to bright sunlight, respectively. Daily 30-min uv lamp exposures on living hosts harboring all life phases of the parasite did not interfere with the ensuing infection in the blowfly's midgut and the pathogen's developmental cycle. Spores harvested from uv-treated infected hosts were found to be as infective as spores retrieved from hosts not treated with uv. Spores contained in dried fecal droplets and exposed up to 3 hr to the uv lamp, or 12 hr to bright sunlight, respectively, remained infective. Addition of uric acid to a preparation of naked spores prior to 15- and 30-min uv irradiations yielded 100% infection in both host groups. A uv-protective function is ascribed to components provided by the host's tissues and feces.     

MORPHOGENESIS OF CAPITATE GLANDULAR HAIRS OF CANNABIS SATIVA (CANNABACEAE)

The glandular secretory system in Cannabis sativa L. (marihuana) consists of three types of capitate glandular hairs (termed bulbous, capitate-sessile, and capitate-stalked) distinguishable by their morphology, development, and physiology. These gland types occur together in greatest abundance and developmental complexity on the abaxial surface of bracts which ensheath the developing ovary. Bulbous and capitate-sessile glands are initiated on very young bract primordia and attain maturity during early stages of bract growth. Capitate-stalked glands are initiated later in bract growth and undergo development and maturation on medium, to full sized bracts. Glands are epidermal in origin and derived, with one exception, from a single epidermal initial. The capitate-stalked gland is the exception and is of special interest because it possesses a multicellular stalk secondarily derived from surrounding epidermal and subepidermal cells. Glands differentiate early in development into an upper secretory portion and a subtending auxiliary portion. The secretory portion, depending on gland type, may range from a few cells to a large, flattened multicellular disc of secretory cells. The secretory portion produces a membrane-bound resinous product which caps the secretory cells. Capitate-stalked glands are considered to be of particular evolutionary significance because they may represent a gland type secondarily derived from existing capitate-sessile glands.     

The identification of membrane glycocomponents in polyacrylamide gels: A rapid method using I-labeled lectins

A rapid method is described for the identification of lectin binding membrane glycocomponents in polyacrylamide gels. The method requires only small quantities of membrane material and is applicable to a wide variety of lectins. Solubilized membrane components are electrophoretically separated according to molecular weight in a SDS-polyacrylamide gel. The gel is then incubated in a solution containing the ¹²⁵I-lectin. After elution of unbound ¹²⁵I-lectin, the gel is dried down and those bands which have bound the ¹²⁵I-lectin are identified by radioautography. The amount of bound ¹²⁵I-lectin can be quantified by either densitometric scanning of the radioautogram or by liquid scintillation counting of the dried gel.