TGF-β1: immunosuppressant and viability factor for T lymphocytes

Transforming growth factor-beta (TGF-β) is a multifunctional cytokine with multiple roles in the immune system. To date, it has been difficult to develop a comprehensive picture of the effect of TGF-β on T lymphocytes, because TGF-β not only acts directly on T lymphocytes, but also acts indirectly by regulating the function of antigen-presenting cells. In early studies, it was mostly the inhibitory function of TGF-β that was demonstrated; recently, however TGF-β was recognized as an antiapoptotic survival factor for T lymphocytes. The outcome of the TGF-β effect on T lymphocytes was shown to strongly depend on their stage of differentiation and on the cytokine milieu. TGF-β cannot be classified as a classical Th1 or Th2 cytokine. However, recently the existence of the TGF-β-producing Th3 subset was described which might play an important regulatory role during an immune response. A better understanding of the molecular mechanism of how TGF-β inhibits or stimulates T lymphocytes will help to predict the complex functions of this cytokine.     

Essential oils of seven Stachys taxa from Croatia

The essential oils of Stachys alpina L., Stachys officinalis (L.) Trevis., Stachys palustris L., Stachys recta L. subsp. recta, S. recta L. subsp. subcrenata (Vis.) Briq., Stachys salviifolia Ten., and Stachys sylvatica L. were obtained by hydrodistillation and analysed by gas chromatography (GC) and gas chromatography–mass spectrometry (GC–MS). Sesquiterpene hydrocarbons were the main group of constituents of all taxa, except S. alpina, which was rich in oxygenated sesquiterpenes. S. alpina and S. palustris had a significant aldehyde fraction and a high amount of alcohols. Some differences in the essential oil composition of two subspecies of S. recta (S. recta subsp. recta and S. recta subsp. subcrenata), growing under almost identical conditions, have been found.     

Effects of ATP and pyrophosphate on properties of glucocorticoid-receptor complexes from rat thymus cells

Cytosols from rat thymus cells incubated with glucocorticoid contain nonactivated and activated receptors and mero-receptor complexes, in relative amounts that depend on the incubation conditions. These forms can be separated by a rapid minicolumn chromatographic technique based on their differential affinities for DNA, DEAE, and hydroxylapatite. We have used this method to examine the effects of ATP, pyrophosphate (PPi), and related compounds on cytosolic complexes. In addition to ATP, already known to promote activation at 0 degrees C, PPi, ADP, and other triphosphates at millimolar concentrations promoted activation of nonactivated complexes. AMP and Pi had little effect. ATP and PPi at millimolar concentrations also reduced binding of activated complexes to DNA. Characterization of the ATP- and PPi-activated complexes by gel filtration and ion exchange chromatography revealed two DNA-binding forms. One was essentially identical (Stokes radius of approximately 5.4 nm, elution from DEAE at approximately 50 mM KCl) to the normal activated complex obtained directly from cells incubated at 37 degrees C. The other had a Stokes radius of approximately 3.1 nm and had no affinity for DEAE. Analysis by minicolumns and gel filtration showed that ATP and PPi prevented formation of mero-receptor complexes, a process which occurs relatively rapidly in untreated thymus cytosols. These compounds did not alter properties of preformed mero-receptor. The accumulation of 3.1-nm complexes in thymus cytosols in which formation of mero-receptor is prevented suggests that this form is an intermediate, normally short-lived, in the conversion of 5.4 nm complexes to mero-receptor.     

Phosphorylated sites within the functional domains of the approximately 100-kDa steroid-binding subunit of glucocorticoid receptors

The steroid-binding subunit of the glucocorticoid receptor is known to be a approximately 100-kDa phosphoprotein composed of an immunogenic, DNA-binding, and steroid-binding domain. When isolated from WEHI-7 cells, this protein contains between two and three phosphoryl groups per steroid-binding site (Mendel WEHI-7 cells, this protein contains between two and three phosphoryl groups per steroid-binding site (Mendel et al., 1987). To identify the domains that contain these phosphorylated sites, we have analyzed the phosphate content of selected proteolytic fragments of the approximately 100-kDa steroid-binding protein from nonactivated and activated receptors. The approximately 100-kDa steroid-binding protein from WEHI-7 cells grown in the presence of [32P]orthophosphate was covalently labeled with [3H]dexamethasone 21-mesylate, purified with the BuGR2 monoclonal antibody, digested with chymotrypsin or trypsin, and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Chymotrypsin digestion of this protein yields a approximately 45-kDa fragment containing both the steroid-binding and DNA-binding domains, which contained both 32P and 3H. Trypsin digestion of the protein yields a approximately 29-kDa fragment encompassing the steroid-binding domain but not the DNA-binding domain of the approximately 100-kDa protein, which also contained both 32P and 3H. The 32P/3H ratio of each fragment provides a measure of phosphate content per steroid-binding site and indicated that each fragment has approximately 30% of the phosphate content of the intact protein. This is sufficient to account for one of the three receptor phosphoryl groups.(ABSTRACT TRUNCATED AT 250 WORDS)     

The rabbit jejunal 'imino carrier' and the ileal 'imino acid carrier' describe the same epithelial function.

For rabbit jejunal brush border vesicles an 'imino carrier' has been defined as a sodium-dependent, alanine-resistant transporter of cyclic imino acids, while for intact rabbit jejunal and ileal epithelia an 'imino acid carrier' has been defined as a sodium-dependent transporter of both aliphatic and cyclic imino acids. This study on intact rabbit intestine examines whether these two terms describe the same epithelial function. The KPro1/2 and the KProi against JMeAIBmc are identical and so are KMeAIB1/2 and KMeAIBi against JPromc. Likewise, KLeui is the same against the transport of both proline and MeAIB. It is, therefore, concluded that the terms 'imino carrier' and 'imino acid carrier' describe the same epithelial function: A sodium-dependent, relatively high afinity, saturable transporter of both aliphatic and cyclic imino acids. Estimates of the apparent affinity and inhibitory constants for MeAIB, proline and leucine confirm that the jejuno-ileal variation of amino acid transport along the rabbit small intestine is a variation of maximal transport capacity.     

Transformation of Dendrobium orchid using particle bombardment of protocorms

Transformed dendrobium orchids (Dendrobium x Jaquelyn Thomas hybrids) were recovered from protocorms bombarded by particles coated with the plasmid pGA482GG/cpPRV4, which contains the plant expressible Nos-NPT II and papaya ringspot virus (PRV) coat protein (CP) genes. Approximately 280 protocorms from four crosses were bombarded and potentially transformed tissues were identified by growth and green color on half-strength Murashige and Skoog medium supplemented with 2% sucrose and 50–100 mg 1^–1 kanamycin sulfate. Kanamycin concentrations that prevented growth of nontransformed tissues could not be used for long-term selection because such levels suppressed the regeneration of potentially transformed tissues. PCR and restriction analysis 21 months after treatment found 13 of 13 plants from two crosses, which appeared kanamycin-tolerant, to contain the Nos-NPT II gene, while only one of these plants carried the vector-linked PRV CP-gene. These results support use of particle bombardment for transformation of this important ornamental monocot.     

Oxygen activation by isolated chloroplasts from Euglena gracilis: Isolation and properties of a fluorescent compound that catalyzes monovalent oxygen reduction

Three yellow fluorescent compounds isolated from Euglena gracilis strain Z show chromatographic and spectroscopic similarities with unconjugated pteridines but are not identical with simple 6-substituted pterins of the neopterin or lumazine type. Two of the compounds contain phosphate, apparently in different types of bonding; these two compounds can be converted into a third compound with the same fluorescent and spectroscopic properties by chemical dephosphorylation. One (water-soluble) phosphatecontaining fluorescent compound stimulates a ferredoxin-dependent monovalent oxygen reduction by isolated Euglena chloroplasts in the dark with NADPH + H⁺ as electron donor.     

Stabilization of thymic glucocorticoid-receptor complexes by the calcium-activated protease inhibitor, calpastatin

We previously described a heat-stable factor from WEHI-7 mouse thymoma, rat liver, spleen, and human chronic lymphocytic leukemia cells that prevents degradation of glucocorticoid-receptor complexes (GRC) in cytosols from rat thymus and acute non-lymphocytic leukemia cells. We now show that the factor has many properties in common with calpastatin, a naturally occurring inhibitor of a family of neutral calcium-activated proteases called calpains. Liver GRC-stabilizing activity and calpastatin activity, in addition to surviving boiling, co-chromatography on columns of DEAE-cellulose ion exchange or agarose A-0.5M gel filtration matrices, and have identical isoelectric points of 5.1. This factor should be especially useful for studying GRC function in the presence of calcium.     

The organization of the ribosomal RNA genes of Chironomus tentans and some closely related species

Southern gel analysis of total DNA from Chironomus tentans showed that the rRNA genes (rDNA) are homogeneous in structure. After cloning in Escherichia coli plasmid pBR313, the rDNA organisation was further studied by restriction fragment analysis and R-loop mapping. No heterogeneity could be detected by heteroduplex analysis of six different cloned rRNA cistrons. R-loop sizes of 1.69 and 3.63 kilobases (kb) were measured for the 18S and 28S rRNA coding sequences. The two spacers are 0.75 and 1.77 kb long. Southern gel analysis showed also a homogeneous rDNA structure for a Canadian population of C. tentans and C. pallidivittatus. The same technique indicated, however, that the rDNA of two other closely related species of C. thummi and C. melanotus is heterogeneous in structure. A possible correlation between this heterogeneity and the presence of heterochromatin in these species is discussed.