Subject(s)
Endotoxins , Genetic Engineering , Pest Control, Biological/methods , Animals , Bacillus thuringiensis/genetics , Bacillus thuringiensis/physiology , Bacillus thuringiensis Toxins , Bacterial Proteins/genetics , Bacterial Toxins/genetics , Cloning, Molecular , DNA Restriction Enzymes , DNA, Recombinant , Fungi/genetics , Fungi/physiology , Genes, Bacterial , Genes, Fungal , Genes, Viral , Hemolysin Proteins , Insect Viruses/genetics , Insect Viruses/physiology , Insecticides , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/physiology , Peptides/genetics , Peptides/physiology , Plants/genetics , Protein Precursors/genetics , Spores, Bacterial/physiologyABSTRACT
Bacteria with specific temperature sensitive lethal mutations in the gene for the beta' subunit of RNA polymerase synthesize both the beta and beta' subunits at a several fold higher rate at 42 degrees C than wild-type cells relative to total protein. Synthesis of the alpha and sigma subunits proceeds at essentially the wild-type rates under these conditions. In contrast, a mutant with a temperature sensitive lethal mutation in the beta subunit gene synthesizes beta and beta' at 42 degrees C at slightly lower rates than wild-type, while alpha and sigma synthesis is not significantly altered. In all of the mutants at 42 degrees C, newly synthesized alpha subunits are stable, while the beta, beta' and sigma subunits are rapidly degraded. The apparent uncoupling of betabeta' and alpha subunit synthesis seen in the beta' mutants at 42 degrees C might suggest that the synthesis of these subunits is at least in part controlled by different mechanisms.
Subject(s)
Escherichia coli/enzymology , Genes , Genes, Lethal , Hot Temperature , MutationABSTRACT
A temperature-sensitive mutant having a lethal mutation in the gene for the beta subunit of RNA polymerase (nucleosidetriphosphate:RNA nucleotidyltransferase, EC 2.7.7.6) exhibits an apparent 2- to 3-fold decrease in the rates of both beta and beta' subunit synthesis at the non-permissive temperature, relative to total protein. In contrast, a temperature-sensitive mutant with a lethal mutation in the gene encoding beta' has a 5- to 6-fold increase in the rates of beta and beta' synthesis at 42 degrees. These beta and beta' mutants also exhibit rapid degradation of both subunits at the high temperature.
Subject(s)
DNA-Directed RNA Polymerases/metabolism , Escherichia coli/enzymology , Mutation , DNA-Directed RNA Polymerases/biosynthesis , Electrophoresis, Polyacrylamide Gel , Escherichia coli/growth & development , Escherichia coli/metabolism , Genetics, Microbial , Isotope Labeling , Kinetics , Leucine/metabolism , Macromolecular Substances , Sodium Dodecyl Sulfate , Temperature , Time Factors , TritiumABSTRACT
A heat-inducible lysis-defective lambda prophage has been integrated directly into the E. coli chromosome at a site (bfe) very closely linked to the ribonucleic acid polymerase mutation rif(d), a dominant rifampin resistance allele. This unusual lysogen has facilitated the isolation of specialized transducing phages conferring rifampin resistance to sensitive cells, and carrying at least the beta subunit gene of RNA polymerase in intact form.
Subject(s)
Coliphages/isolation & purification , DNA-Directed RNA Polymerases/metabolism , Escherichia coli/enzymology , Genes , Alleles , Chromosomes, Bacterial , Culture Media , DNA, Bacterial/metabolism , DNA-Directed RNA Polymerases/antagonists & inhibitors , Drug Resistance, Microbial , Genes, Dominant , Lysogeny , Mutation , Phenotype , Rifampin/pharmacology , Temperature , Transduction, GeneticABSTRACT
lambdacI857s7drif(d)18 phage is a defective, specialized transducing phage carrying the RNA polymerase mutation rif(d), a dominant rifampicin-resistant allele. This phage DNA directs in vitro synthesis of the beta subunit of RNA polymerase in a DNA-dependent protein-synthesizing system. Prophage induction in cells that are lysogenic for this phage increases the rate of beta-subunit synthesis in vivo by at most 2-fold. No increase in subunit synthesis above the haploid level is observed in cells diploid for the beta gene.
