Interleukin 2-induced tyrosine phosphorylation. Interleukin 2 receptor beta is tyrosine phosphorylated

Interaction of interleukin 2 (IL2) with its high affinity membrane receptor complex (IL2R) is sufficient to induce proliferation of T lymphocytes. However, the biochemical mechanisms by which IL2 induces this process remain unresolved. The IL2R complex consists of at least two distinct polypeptides that bind IL2, a 75-kDa intermediate affinity subunit (IL2R beta) and a 55-kDa low affinity subunit (IL2R alpha). As indicated by Western blotting with anti-phosphotyrosine-specific antibodies and confirmed by phosphoamino acid analysis, we now demonstrate that interaction of the T cell growth factor interleukin 2 (IL2) with its high affinity receptor on IL2-sensitive human peripheral blood lymphoblasts induces tyrosine phosphorylation of proteins of 92, 80, 78, 70-75, and 57 kDa. IL2 induced tyrosine phosphorylation in YT 2C2 cells which express only the 75-kDa intermediate affinity IL2 binding molecule (IL2R beta) but not in cells which either express only the 55-kDa low affinity IL2 receptor molecule (IL2R alpha) or no IL2-binding sites. Therefore, IL2R beta, in the absence of IL2R alpha, appears sufficient to transduce the transmembrane signal leading to tyrosine phosphorylation. Two different antibodies reactive with phosphotyrosine specifically immunoprecipitated IL2R beta cross-linked to radiolabeled IL2. These findings suggest that IL2R beta is a substrate for the tyrosine kinase which is activated by IL2 binding to its receptor. Thus, like several other growth factor receptors, activation of the IL2R results in an increase in tyrosine phosphorylation with the receptor itself serving as one substrate.     

Ultracytochemistry of cholera-toxin binding sites in ciliary processes

Summary Cholera toxin reduces the rate of formation of aqueous humor in concentrations (10^–11 M) that do not disturb the morphology of the aqueoushumor forming epithelial cells of the ciliary processes of the rabbit eye. The search for an endogenous mediator of aqueous-humor formation comparable to cholera toxin in its mode of operation prompted us to map the distribution of cell surface receptors for cholera toxin in the ciliary processes of the eyes of rabbits. Cytochemical studies were carried out with the use of conjugates of cholera toxin to fluorescein isothiocyanate (CT-FITC) and to horseradish peroxidase (CT-HRP), and of the B subunit of cholera toxin to horseradish peroxidase (B-HRP). Multiple fluorescent CT-FITC binding sites were observed on the outer nonpigmented epithelial layer near the crests of the processes. Processes incubated with CT-HRP in vitro showed surface staining of 30–40% of the nonpigmented epithelial cells. A prominent reaction product was observed along the basal and lateral plasma membranes of these cells. In vivo studies carried out after arterial infusion of B-HRP showed a reproducible dense reaction product between the apical surfaces of the pigmented epithelium (PE) and of the nonpigmented epithelium (NPE) facing each other. Aggregations of reaction product were observed with the electron microscope in the extracellular space between the apices of PE and NPE. The apical plasma membrane of the endothelium of the blood vessels near the crests of the ciliary processes was stained after either in vivo or in vitro exposure to peroxidase conjugates. These findings indicate that the cell-surface receptors which mediate the action of cholera toxin on aqueous humor formation are very likely localized in the apical plasma membranes of the epithelium of the ciliary processes.Supported in part by USPHS grant # EY-00237, the Connecticut Lions Eye Research Foundation, Inc., and Research to Prevent Blindness, Inc.     

Isolation and characterisation of the Xenopus laevis albumin genes: loss of 74K albumin gene sequences by library amplification.

The blood of the frog X.laevis contains 2 albumins of 68,000 and 74,000 daltons which are encoded in the liver by two related mRNAs. When an amplified X.laevis DNA library was screened with cloned albumin cDNA only 68,000 dalton albumin gene sequences were isolated. Hybridisation of the albumin cDNA to Southern-blots of Eco R1 digested X.laevis DNA showed that the sequences present in the recombinants did not account for all the fragments which hybridised on the Southern-blots. This indicated that 74K albumin gene sequences exist but that they are not present in the amplified DNA library. A X.laevis genomic library was therefore constructed and screened for albumin genes without amplification. Both 68K and 74K albumin gene sequences were isolated. Recombinants containing 74K albumin gene sequences grew extremely poorly and this probably explains why the 74K albumin sequences were ot isolated from the amplified library. Characterisation of the cloned DNA indicates that there is one sequence coding for the 68K albumin but two different sequences coding for the 75K albumin.     

Hair morphology and the relationships of species inSolanum sect.Basarthrum

The usefulness of features of leaf hairs in distinguishing subgenera and sections is well documented in bothRhododendron andSolanum. In this analysis of the taxa ofSolanum sect.Basarthrum (23 species), and of a sample of closely related taxa from sect.Petota (22 species), it is shown that such features serve to delineate subsectional groups such as series and some species as well. SectionBasarthrum has an unexpected diversity of hair types. Although this group has been characterized by 2-celled bayonet hairs, more than one half of the taxa in the section bear multicellular finger hairs, and 3 species also possess branched hairs. Thus, major rearrangements of the species previously assigned to sect.Basarthrum are indicated or supported by pubescence features. The taxa studied from seriesEtuberosa andJuglandifolia (both of sect.Petota) show hair types that a) are relatively primitive for the section, and b) show linkage between sects.Petota andBasarthrum.     

CHROMOSOME NUMBERS IN SOUTH AMERICAN HAPLOPAPPUS CASS. (COMPOSITAE)

Chromosome counts are reported for 33 species from all four sections of the genus Haplopappus in South America. These include first reports for 28 species and two putative hybrids. All chromosome numbers reported herein are 2n = 5_II, with the exception of H. prunelloides with 2_n = 6_II. Unlike the North American species, the morphological diversity of South American taxa is not concomitant with chromosomal variation.     

Microbiology of Wetwood: Importance of Pectin Degradation and Clostridium Species in Living Trees

Wetwood samples from standing trees of eastern cottonwood (Populus deltoides), black poplar (Populus nigra), and American elm (Ulmus americana) contained high numbers of aerobic and anaerobic pectin-degrading bacteria (10⁴ to 10⁶ cells per g of wood). High activity of polygalacturonate lyase (≤0.5 U/ml) was also detected in the fetid liquid that spurted from wetwood zones in the lower trunk when the trees were bored. A prevalent pectin-degrading obligately anaerobic bacterium isolated from these wetwoods was identified as Clostridium butyricum. Pectin decomposition by C. butyricum strain 4P1 was associated with an inducible polygalacturonate lyase and pectin methylesterase, the same types of pectinolytic activity expressed in the wetwood of these trees. The pH optimum of the extracellular polygalacturonate lyase was alkaline (near pH 8.5). In vitro tests with sapwood samples from a conifer (Douglas fir, Pseudotsuga menziesii) showed that tori in membranes of bordered pits are degraded by pure cultures of strain 4P1, polygalacturonate lyase enzyme preparations of strain 4P1, and mixed methanogenic cultures from the tree samples of wetwood. These results provide evidence that pectin in xylem tissue is actively degraded by C. butyricum strain 4P1 via polygalacturonate lyase activity. The importance of pectin degradation by bacteria, including Clostridium species, appears paramount in the formation and maintenance of the wetwood syndrome in certain living trees.     

Structural studies of urinary oliogosaccharides from patients with mannosidosis

Eight neutral oligosaccharide fractions were obtained from the pooled urine of two patients with mannosidosis by Bio-Gel P2 and Bio-Gel P4 column chromatography. The structures of seventeen oligosaccharides were determined by monosaccharide composition analysis, methylation studies, acetolysis, Smith degradation, and ¹³C NMR analysis. Three of the proposed structures, Manα1-3Manβ1-4GlcNAc, Manα1-2Manα1-3Manβ1-4GlcNAc, and Manα1-2Manα1-2Manα1-3Manβ1-4GlcNAc are identical to those first published by Norden et al. (N. E. Norden, A. Lundblad, S. Svennson, P. A. Ockerman, and S. Autio, 1973. J. Biol. Chem.248, 6210–6215; N. E. Norden, A. Lundblad, S. Svennson, and S. Autio, 1974. Biochemistry13, 871–874). Thirteen of them, Manα1-3Manα1-6(Manα1-3)-Manβ1-4GlcNAc, Manα1-3Manα1-6(Manα1-2Manα1-3)Manβ1-4GlcNAc, and 11 isomers of (Manα1-2)_0–4[Manα1-6(Manα1-3)Manα1-6(Manα1-3)Manβ1-4GlcNAc], are the same as those first published by Yamashita et al. (K. Yamashita, Y. Tachibana, K. Mihara, S. Okada, H. Yabuuchi, and A. Kobata, 1980, J. Biol. Chem.255, 5126–5133); a tetrasac-charide, Manα1-6(Manα1-3)Manβ1-4GlcNAc, is newly reported and several other structural possibilities are proposed.     

Biofeedback treatment of dysmenorrhea

Nine dysmenorrheic women were run in EMG and thermal biofeedback procedures with concurrent autogenic relaxation practice. Significant reductions in subjective estimates of symptomology associated with dysmenorrhea were noted in all subjects. EMG levels correlated positively with the reductions in symptoms. Thermal levels did not correlate with EMG. In fact no consistent patterns in thermal measures were noted. However, thermal biofeedback cannot be ruled out as an effective treatment for dysmenorrhea since reductions in symptoms occurred during thermal biofeedback training. Another significant aspect of the present study is the effectiveness of long treatment procedures. A six month period was employed and significant reductions in symptoms were noted following two months of biofeedback treatment. Finally, the importance of beginning biofeedback treatment prior to onset of menstrual symptoms is indicated.     

Injury and recovery of Bacillus megaterium from mild chlorhexidine treatment

Treatment of Bacillus megaterium cell suspensions with 12 /μmol/1 chlorhexidine diacetate for 5 min led to an approximate 50%, reduction in viability when plated onto tryptone soya agar (TSA). Fifty percent of the surviving fraction were unable to form colonies on TSA containing 5.5% w/v KCI. Such loss of KCl tolerance is indicative of membrane damage, and was recovered within 30 min of incubation in tryptone soya broth (TSB). Multiplication of the damaged organisms did not recommence in this medium until after 60 min. Inclusion of inhibitors of respiration, and of protein, RNA and DNA synthesis in the TSB recovery medium did not significantly affect either the rate or extent of the recovery of KCl tolerance by damaged organisms.     

Reversed-phase high-performance liquid chromatography analysis of 6-thioguanine applicable to pharmacologic studies in humans

6-Thioguanine (6TG) and its metabolites were analyzed in human plasma with a reversed-phase high-performance liquid chromatographic method. 6TG and related compounds were extracted from plasma with an equal volume of 2 N perchloric acid at a 50–100% recovery efficiency. The neutralized extracts were chromatographed on a μBondapak C₁₈ column by two separate isocratic conditions. 6TG, 6-thiouric acid, 6-thioxanthine, 6-thioguanosine, and 6-methylthiouric acid were analyzed with 0.01 M sodium acetate, pH 3.5–10% methanol as the mobile phase and 340 nm for detection. 6-Methylthioguanine and three unknown metabolites were separated with acetate—25% methanol and 310 nm detection. One of the unknowns was identified as 6-methylthioguanosine. External standard calibration was used for quantitation. The 6TG detection limit was 0.8 nmol/ml in plasma.