The effects of decreased growth temperature on the cystine content of cystinotic fibroblasts

Cystinotic fibroblasts transferred from 37 degrees C to 28 degrees C accumulated additional cystine over the period from 4 to 7 days of incubation at 28 degrees C, after which the additional cystine was lost; warming (to 37 degrees C) of cells with elevated cystine stores led to rapid cystine loss. These results, taken together with previously published data showing cystine release from cystinotic fibroblasts incubated at above-normal temperature, are interpreted as indicating the presence in the cystinotic fibroblast lysosome membrane of a cystine-porter whose efficacy is increased by an increase in membrane fluidity. This porter may be the residual activity of the cystine porter that is known to be deficient in cystinosis, or it may be a second as yet unrecognized porter. It is further proposed that this porter is responsible for the presumed efflux of cystine from cystinotic lysosomes.     

Effects of male partners upon proceptivity in ovariectomized estradiol-treated marmosets (Callithrix jacchus)

Effects of male partners upon the expression of female proceptivity were examined in two experiments using 16 ovariectomized marmosets. Experiment 1 showed that the female's proceptive tongue-flicking display (PTF) is triggered specifically by eye contact with the male. Stimulation of PTF by administration of estradiol-17 beta (E2) to ovariectomized females depends in part upon the male's responsiveness to female displays, such as "staring" and "freezing," which may serve to attract his attention and to establish eye contact. Experiment 2 provided evidence that females' proceptivity decreases if their male partners are lesioned in the preoptic area/anterior hypothalamus. Such males are sexually hypoactive and less responsive to the females' visual displays. However, E2 still activates PTF if females succeed in initiating eye contact with males. Results indicate that variability in effects of E2 upon proceptivity in marmosets may be influenced by subtle aspects of facial communication between the sexes as well as by individual differences in hormonal sensitivity. Copulatory activity in males is not essential for E2 to exert its stimulatory action upon proceptivity in female marmosets.     

A general principle for the allocation of limited resources

Summary A marginal fitness theorem is derived for the allocation of a limited resource among alternative activities that have effects on the fitness of an individual. The marginal advantage theorem states that at the evolutionarily stable strategy (ESS), the marginal gains from increasing each of the allocations (expressed as partial derivatives of the fitness advantage of a rare mutant) are equal. The theorem is true for all proportional allocations (a + b + c + ...=j), regardless of the number of allocations, the nature of the response curves describing the direct effects of the allocations [f(a), etc.], or the way the effects of different allocations combine into fitness. The theorem is extended to size-number compromises and packaging strategies. The marginal advantage theorem is used to derive general theorems about the marginal effects of allocations [f (a), etc.] at the ESS and matching rules concerned with the total fitness to cost ratios of allocations at the ESS. The marginal advantage theorem is applicable to diverse allocation strategies, and provides a method for obtaining ESS allocations for any number of allocations and their components.     

Regulation fo protein kinase C activity by various lipids

Protein kinase C has recently attracted considerable attention because of its importance in the control of cell division, cell differentiation, and signal transduction across the cell membrane. The activity of this enzyme is altered by several lipids such as diacylglycerol, free fatty acids, lipoxins, gangliosides, and sulfatides. These lipids may interact with protein kinase C either directly or through calcium ions and produce their regulatory effect (activation or inhibition) on the activities of the enzymes phosphorylated by this kinase. These processes widen our perspective of the regulation of intercellular and intracelluular communication.Abbreviations used (PK-C) Protein kinase C - (cAMP-PK) cAMP dependent protein kinase - (DAG) diacylglycerol - (PtdSer) phosphatidylserine - (InsP ₃) inositol 1,4,5-trisphosphate - (PtdIns 4,5-P₂) inositol 4,5 bisphosphate - (FFA) free fatty acid - (MBP) myelin basic protein - (ATP) adenosine triphosphate - (GTP) guanine triphosphate - (TPA) 12-tetradecanoylphorbol-13-acetate - (EGF) epidermal growth factor - (PDGF) platelet derived growth factor - (NeuNAc) and N-acetylneuraminic acid     

Leader protein of foot-and-mouth disease virus is required for cleavage of the p220 component of the cap-binding protein complex.

Suppression of host protein synthesis in cells infected by poliovirus and certain other picornaviruses involves inactivation of the cap-binding protein complex. Inactivation of this complex has been correlated with the proteolytic cleavage of p220, a component of the cap-binding protein complex. Since picornaviral RNA is not capped, it continues to be translated as the cap-binding protein complex is inactivated. The cleavage of p220 can be induced to occur in vitro, catalyzed by extracts from infected cells or by reticulocyte lysates translating viral RNA. Expression of polioviral protease 2A is sufficient to induce p220 cleavage, and the presence in 2A of an 18-amino-acid sequence representing a putative cysteine protease active site correlates with the ability of different picornaviruses to induce p220 cleavage. Foot-and-mouth disease virus (FMDV) infection induces complete cleavage of p220, yet the FMDV genome codes for a 2A protein of only 16 amino acids, which does not include the putative cysteine protease active site. Using cDNA plasmids encoding various regions of the FMDV genome, we have determined that the leader protein is required to initiate p220 cleavage. This is the first report of a function for the leader protein, other than that of autocatalytic cleavage from the FMDV polyprotein.     

Antibodies to pollen exines

A polyclonal antiserum and monoclonal antibodies have been prepared to purified pollen exines of Calocedrus decurrens Florin. The location of the antigen is in the exine, as shown by light-and electron-microscopic immunocytochemistry. The greatest reduction in antibody binding follows treatment of the exine with chemicals known to alter sporopollenin. These results provide evidence that sporopollenin is antigenic. Exines of ten species of gymnosperms and angiosperms also bound the polyclonal antiserum, indicating similarity of sporopollenin structure.     

Molecular analysis of plasmid DNA repair within ultraviolet-irradiated Escherichia coli. I. T4 endonuclease V-initiated excision repair

The process by which DNA-interactive proteins locate specific sequences or target sites on cellular DNA within Escherichia coli is a poorly understood phenomenon. In this study, we present the first direct in vivo analysis of the interaction of a DNA repair enzyme, T4 endonuclease V, and its substrate, pyrimidine dimer-containing plasmid DNA, within UV-irradiated E. coli. A pyrimidine dimer represents a small target site within large domains of DNA. There are two possible paradigms by which endonuclease V could locate these small target sites: a processive mechanism in which the enzyme "scans" DNA for dimer sites or a distributive process in which dimers are located by random three-dimensional diffusion. In order to discriminate between these two possibilities in E. coli, an in vivo DNA repair assay was developed to study the kinetics of plasmid DNA repair and the dimer frequency (i.e. the number of dimer sites on a given plasmid molecule) in plasmid DNA as a function of time during repair. Our results demonstrate that the overall process of plasmid DNA repair initiated by T4 endonuclease V (expressed from a recombinant plasmid within repair-deficient E. coli) occurs by a processive mechanism. Furthermore, by reducing the temperature of the repair incubation, the endonuclease V-catalyzed incision step has been effectively decoupled from the subsequent steps including repair patch synthesis, ligation, and supercoiling. By this manipulation, it was determined that the overall processive mechanism is composed of two phases: a rapid processive endonuclease V-catalyzed incision reaction, followed by a slower processive mechanism, the ultimate product of which is the dimer-free supercoiled plasmid molecule.     

Identification of N-glycolylneuraminic acid-containing gangliosides of cat and sheep erythrocytes. 252Cf fission fragment ionization mass spectrometry in the analysis of glycosphingolipids

A number of gangliosides were isolated from cat and sheep erythrocytes for use in analyzing the specificity of a panel of human anti-heterophile monoclonal antibodies. The structures of these compounds were determined by a combination of different procedures, including sugar analysis, glycosidase treatment, periodate oxidation, TLC immunostaining, methylation analysis, and mass spectrometry. These methods identified the cat erythrocytes gangliosides (C1 and C2) as N-glycolylneuraminic acid (NeuGc)-containing hematosides; C1 was shown to be NeuGc alpha 2----8NeuGc alpha 2----3Gal beta I----4Glc-Cer [NeuGc)2GD3) and C2 to be NeuAc alpha 2----8NeuGc alpha 2----3Gal beta 1----4Glc-Cer [NeuAc-NeuGc-)GD3). The two sheep gangliosides (S1 and S2) were found to be novel glycolipids based on the paragloboside sequence; S1 was identified as NeuGc alpha 2----8NeuGc alpha 2----3Gal beta 1----4GlcNAc beta 1----3Gal beta 1----4Glc-Cer [NeuGc)2-disialylparagloboside) and S2 as NeuAc alpha 2----8NeuGc alpha 2----3Gal beta 1----4GlcNAc beta 1----3Gal beta 1----4Glc-Cer [NeuAc-NeuGc-)-disialylparagloboside). Structural analysis of these compounds was aided by the use of 252Cf fission fragment ionization time-of-flight mass spectrometry. This method provided easily interpretable spectra on methylated derivatives which were particularly useful in determining the sialic acid composition of the gangliosides and the sequence of their disialosyl side chains.     

A single base mutation in the gene for type III collagen (COL3A1) converts glycine 847 to glutamic acid in a family with Ehlers-Danlos syndrome type IV. An unaffected family member is mosaic for the mutation

Summary Ehlers-Danlos syndrome type IV, an inherited connective tissue disease, is usually caused by mutations in the gene for type III collagen. Here, we describe a glycine to glutamic acid substitution in a patient with this syndrome. Previous studies had shown that fibroblasts from the patient, his mother and brother secreted a reduced amount of type III collagen and also produced an overmodified form of the protein that was preferentially retained intracellularly. Peptide mapping experiments indicated that the mutation was located within cyanogen bromide peptide 9. This was supported by chemical cleavage analysis and sequencing of cDNA encoding this region. Allele-specific oligonucleotide hybridisation of genomic DNA confirmed that a G to A mutation converted Gly 847 to Glu. The mutation was present in two other affected family members and also in a third, who was clinically unaffected. Further analysis of this unaffected individual revealed reduced mutant:normal ratios in DNA obtained from both blood and hair samples, showing that she was mosaic for the mutation.