Stimulation by Cyclic Adenosine Monophosphate of Plasmid Deoxyribonucleic Acid Replication and Catabolite Repression of the Plasmid Deoxyribonucleic Acid-Protein Relaxation Complex

Colicinogenic factors ColE1 and ColE2 are bacterial plasmids that exist in Escherichia coli as supercoiled deoxyribonucleic acid (DNA) and as strand-specific, relaxation complexes of supercoiled DNA and protein. Newly replicated ColE1 DNA becomes complexed with protein after the replication event. This association of DNA and protein can take place under conditions in which DNA or protein synthesis is arrested. The addition of cyclic adenosine monophosphate (c-AMP) to normal cells growing in glucose medium results in a six- to tenfold stimulation in the rate of synthesis of the protein component(s) of the complex and a three- to fivefold stimulation in the rate of ColE1 DNA replication. Employing mutants deficient in catabolite gene activator protein or adenylate cyclase, it was shown that synthesis of both the plasmid-determined protein colicin E1 and the protein component(s) of the ColE1 relaxation complex is mediated through the c-AMP-catabolite gene activator protein system. Addition of c-AMP to ColE2-containing cells results in the stimulation of synthesis of ColE2 DNA and relaxation protein(s) as well as in the production of a protein component of the ColE2 relaxation complex that renders it sensitive to induced relaxation by heat treatment. In the case of ColE2, synthesis of the relaxation protein(s) is not dependent upon catabolite gene activator protein.     

The effect of the virus-serum incubation period upon vaccinia virus serum neutralization titers

Rabbit and human vaccinia virus immune sera were titrated by the plaque reduction serum neutralization (SN) method. Titers determined following a 2-h, 37 degree C virus-serum incubation period were not significantly different from those determined following an 18-h incubation period (2-h, 37 degree C incubation followed by a 16-h, 4 degree C period). However, control virus plaque counts decreased significantly after the longer incubation period. Numerous factors affecting SN titer estimation are reviewed. Standardization of SN test conditions may be useful in comparative potency evaluation of vaccinia-vectored recombinant DNA viral vaccines.     

The presence and binding characteristics of calmodulin in microsomal preparations enriched in sarcoplasmic reticulum from rabbit skeletal muscle

ATP-dependent oxalate facilitated calcium transport in sarcoplasmic reticulum (SR) preparations obtained from rabbit vastus lateralis muscle (fast skeletal muscle; F_sr) and soleus (slow skeletal muscle; S_sr) was determined. Addition of exogenous calmodulin did not stimulate calcium transport in either F_sr or S_sr preparations. F_sr and S_sr previously washed in 1 mM EGTA demonstrated a reduced capacity to transport Ca²⁺; the exogenous addition of calmodulin (0.24 μM) under these conditions, did not restore uptake activity but significantly decreased the steady-state level of Ca²⁺ uptake. Extracts of skeletal SR prepared by treatment with 0.2 mM EDTA and boiling produced significantly more stimulation of red cell Ca²⁺ATPase activity than extracts prepared by boiling alone. This stimulation of red cell Ca²⁺-ATPase was inhibited to a significant extent by $\text{48}\text{80}$, a known anti-calmodulin agent. Radioimmunoassay revealed that extracts prepared by boiling or EDTA-treatment followed by boiling contained considerable amounts of calmodulin. Washing with 1 mM EGTA, though, did not release any calmodulin from SR. These studies reveal that calmodulin is present in both F_sr and S_sr and can only be removed by harsh treatments. The role of calmodulin in skeletal muscle Ca²⁺-transport remains to be determined.     

Volume regulation in salt-acclimated toad (Bufo viridis): the role of urea and the urinary bladder

Body water (weight) was studied in the euryhaline toad Bufo viridis during high salt (500 mOsm NaCl) acclimation. Plasma osmolality was greatly increased upon salt acclimation mainly by urea, and was always hyperosmotic to the ambient solution. Water content was regulated quite efficiently in slowly acclimated undisturbed toads. Repeatedly catheterized toads behaved like osmometers when transferred to hyperosmotic solutions. Total urea loss was greatly reduced in salt acclimated toads, suggesting urine was not voided under these conditions. It is concluded that urea accumulation, inhibition of the urine voiding response and the urine in the bladder are the principal factors involved in volume regulation under conditions of salt acclimation.     

Evaluation of the Escherichia coli K12 inductest for detection of potential chemical carcinogens

46 chemicals of diverse classes and structures, including 30 known animal carcinogens, were evaluated for prophage-inducing ability using the Escherichia coli inductest with lysogenic strain GY5027 envA - uvrB- and indicator strain GY4015 ampR . The inductest detected 9 of 30 known carcinogens as genotoxic agents, including 3 polycyclic hydrocarbons, 2 aflatoxins, and 2 antitumor antimicrobials. Among the 21 carcinogens ineffective as prophage inducers were 3 aromatic amines (other than 2-aminoanthracene), 3 azo-aminoazo compounds, 2 methanesulfonates, and 2 nitro aromatics. In contrast, 18 and 17 of the 30 animal carcinogens were detected as genotoxic agents in the Salmonella/Ames test and E. coli WP2/ WP100 rec assay, respectively. The threshold sensitivity of the inductest was less than that of the Salmonella/Ames test for chemicals genotoxic in both tests. The ineffectiveness of the inductest as a routine test for detecting potential chemical carcinogens may be related to the nature of the DNA damage lesions formed by various genotoxic agents.     

Movement characteristics and power output of guinea-pig and hamster spermatozoa in relation to activation

Spermatozoa were collected from the cauda epididymidis of golden hamsters and guinea-pigs, and the acrosome reaction was induced in vitro. Movement characteristics of the spermatozoa were assessed with high-speed cinemicrography. Before the initiation of the acrosome reaction (preactivated spermatozoa), sperm movement in both species was characterized by progressive swimming by regular flagellar waves of moderate amplitude and relative high frequency. After the acrosome reaction (activated spermatozoa), sperm movement in both species was not progressive, and was characterized by whiplash-like flagellar undulations of significantly (P less than 0.05) higher amplitude and lower frequency. Calculation of the hydrodynamic power output by a new theory indicated that no significant change occurred after activation.     

Inhibition of 3,4-dihydroxy-l-phenylalanine decarboxylase in rat striatal synaptosomes by amino acids interacting with substrate transport

Dopamine synthesis from 3,4-dihydroxy-l-phenylalanine in rat striatal synaptosomes was inhibited by a number of amino acids with aromatic or large aliphatic side chains. Inhibition was not seen when aromatic amino acid decarboxylase activity was measured in disrupted synaptosomes. Similarly, inhibition of dopamine synthesis from tyrosine was seen in the presence of leucine. The inhibition most likely results from interactions of the amino acids with substrate transport across the synaptosome plasma membrane, rather than directly with the catalytic enzymes. The kinetic data obtained are used to infer information about the relevant transport process; they suggest the potential importance of amino acid efflux as a regulatory step.