Influenza viruses are T cell-independent B cell mitogens.

UV-inactivated influenza virus A strains of subtypes H1, H2, H3, and H6 were shown to be mitogenic for unprimed splenic lymphocytes from BALB/c mice. Representative viruses of these four subtypes all behaved as T cell-independent B cell mitogens. The magnitude of the proliferative response was determined by the subtype of the hemagglutinin molecule: H2 and H6 viruses were the most potent mitogens, and H3 viruses were moderately mitogenic, whereas H1 viruses induced only low, but significant, levels of proliferation. Mitogenesis was inhibited by antiviral sera and by monoclonal antibodies directed against hemagglutinin.     

Tissue accumulation of cadmium as a function of blood concentration

We have provided data relating Cd concentration in tissue to about a 40-fold range in blood Cd concentration. Osmotic pumps containing cadmium chloride were subcutaneously implanted in male New Zealand white rabbits. The pumps continuously delivered either 0.15 or 1.5 mg Cd/d. Blood and plasma levels of Cd were measured weekly throughout the study. After 28 d, the rabbits were killed and tissue concentrations of Cd determined by atomic absorption spectrophotometry. Less than 10% of the total Cd in the blood was carried in the plasma, the remainder was associated with the blood cells, where it was bound mainly to metallothionein. We found the blood and tissue levels of Cd were correlated for each tissue we investigated. There was a wide range in affinity of the tissues for Cd; liver and kidney had the highest Cd uptake, whereas brain affinity was about 500 times less than liver.     

Induction of heavy metal binding groups by dexamethasone and Zn in cultured Chinese hamster ovary cells

Cd binding capacity and pulse polarography were used to study the inducibility of sulfhydryl groups in cultured Chinese hamster ovary cells (wild type and a Cd-resistant mutant) in response to dexamethasone (dex) and Zn. Evidence is presented that both the wild type and the mutant responded to dex and Zn treatment by induction of sulfhydryl groups. In wild type for Zn and dex as well as in the mutant for dex, this induction seems to be in the form of sulfhydryls attached to particulate or membrane fractions in the cells. For Zn in the Cd-resistant mutant the induction was in the form of metallothionein.     

Multiple conformations of amino acid residues in ribonuclease A

The highly refined 1.26 A structure (R = 0.15) of phosphate-free bovine pancreatic ribonuclease A was modeled with 13 residues having discrete multiple conformations of side chains. These residues are widely distributed over the protein surface, but only one of them, Lys 61, is involved in crystal packing interactions. The discrete conformers have no unusual torsion angles, and their interactions with the solvent and with other atoms of the protein are similar to those residues modeled with a single conformation. For three of the residues--Val 43, Asp 83, and Arg 85--two correlated conformations are found. The observed multiple conformations on the protein surfaces will be of significance in analyzing structure-function relationships and in performing protein engineering.     

Comparison of nonphosphorylated and phosphorylated species of polyomavirus major capsid protein VP1 and identification of the major phosphorylation region.

The major virion protein of polyomavirus, VP1, consists of about six isoelectric species designated A through F. The minor species D, E, and F are phosphorylated and are thought to serve as viral receptors. We first wanted to distinguish whether all VP1 species are derived by post-translational modification from a common amino acid sequence or whether one or more of the species contain a region(s) of altered amino acid sequence resulting from alternate mRNA processing. We compared the VP1 species by detailed peptide mapping with several combinations of specific protease and radioisotopic labels. This approach enabled us to examine more than 80% of the predicted VP1 sequence, including the amino-and carboxy-termini. We found no evidence of sequence differences among any of the VP1 species. The specific incorporation of 32Pi was found to be the same for all of the phosphorylated species. Comparison of the phosphorylation sites of in vivo 32Pi-labeled D, E, and F by peptide mapping showed them to be identical. Each phosphorylated species contained a single major phosphopeptide and several minor phosphopeptides. The major phosphoamino acid, identified by acid hydrolysis, was phosphothreonine, with phosphoserine also present. By using chemical cleavage methods, we localized the major phosphorylation region to a central portion of the VP1 sequence. We discuss some features of this region and relate this information to functional implications of phosphorylation.     

The phorbol ester 12-0-tetradecanoyl-phorbol-13-acetate and stimulation of 3H-choline incorporation into endoplasmic reticulum membranes and other subcellular fractions of Krebs II ascites cells during in vitro incubation

Summary After transfer of Krebs II ascites cells from the mouse peritoneum to suspension culture addition of the phorbol ester 12-0-tetradecanoyl-phorbol-13-acetate (TPA) causes an early stimulation of ³H-choline incorporation into phosphatidylcholine (PC). Choline transport into the treated cells, however, was unaffected. Within 30 min of TPA treatment ³H-choline incorporation was almost 300% above the control level. During a 5 hr period of suspension culture the overall patterns of ³H-choline incorporation were similar in TPA-treated and control cultures though the rate was greatly accentuated by the presence of the phorbol ester. Incubation of cells with cycloheximide prior to incubation with TPA did not result in an inhibition of the TPA-directed ³H-choline incorporation. After 3 hr incubation with TPA there were large increases in radioactivity in all subcellular fractions. At 20 hr, however, the values were not far from those of the control. During the first 3 hr of incubation with TPA the incorporation of ³H-choline into light rough (LR) and smooth (S) membranes was stimulated to levels of 400% and 320% respectively above control values. At later times the profiles of radioactivity in membrane subfractions in TPA-treated and control cultures were similar. The results illustrate an early effect of TPA on PC biosynthesis in Krebs II ascites cells while at later times of incubation the stimulatory effect was virtually abolished.     

Purification and partial characterization of four proteins from human parotid saliva

Four proteins, which have been designated A, B, C and D, have been purified from human parotid saliva. These proteins are the major constituents of parotid saliva which migrate rapidly to the anode in polyacrylamide electrophoresis at pH9.5. Gel filtration and polyacrylamide electrophoresis were employed in the purification procedures. After purification all four preparations were tested for homogeneity by electrophoresis at pH2.8 and 9.5, by isoelectric focusing in the pH range 3-10, by immunodiffusion, and by sedimentation in the analytical ultracentrifuge. None of the proteins showed significant activity in assays for amylase, acid and alkaline phosphatase, protease, lysozyme, ribonuclease, peroxidase, beta-glucuronidase, beta-galactosidase, iron-binding activity and esterase. No cross-reactions were detected with antisera specific for lactoferrin and 15 serum proteins. All four proteins were rich in glutamic acid, proline and glycine and were lacking completely the sulphur-containing amino acids. Proteins A and C contained no threonine or tyrosine. Carbohydrate could be demonstrated only in protein A at a concentration of 4% of the total protein.