The chemistry of a chitin-protein from crayfish (Astacus fluviatilis) (author's transl)]

A chitin-protein complex is obtained from crayfish (Astacus fluviatilis) by gentle decalcification with acetic acid and EDTA. The complex is treated with lithium rhodanide, urea, anhydrous formic acid, pronase, papain, anhydrous formamide or 1N NaOH. The first three of these substances have little or no effect on the stability of the chitin-protein complex. The enzymes remove most of the protein, and the last two reagents remove all of it. The protein remaining bound to the polysaccharide after treatment of the chitin-protein complex with pronase or papain is relatively rich in glycine. Quantitative analysis yielded values for the acetyl, glucosaminyl and amino acid residues which reproduce the composition of the corresponding chitin-protein complex. In these calculations however, allowance must be made for the fact that glucosamine is partly destroyed by the acid by hydrolysis and interferes with the determination of basic amino acids. The results of the present work suggest that the chitin in crayfish is present in the form of a stable complex with protein, possibly held together by covalent binding of the protein to the chitin, with glycine as the connecting amino acid.     

Precocious reprogramming of eupyrene-apyrene spermatogenesis commitment induced by allatectomy of the penultimate larval instar of the moth Actias selene

A possible causal relationship between the switch-over from eupyrene to apyrene spermatogenesis and pupation in Lepidoptera was examined in Actias selene. Precocious pupation, 11 days earlier than in the controls, was induced by allatectomy on the first day of the penultimate larval instar. The course of the eupyrene spermatogenesis, until nuclear elongation in the spermatid, is not related to pupation. In both allatectomized and control individuals, eupyrene metaphases appear 8 days after ecdysis of the fourth larval instar. Nuclear elongation, however, is triggered in the allatectomized individuals earlier than in the controls, probably by the premature decline of the juvenile hormone titre following allatectomy. Apyrene commitment, on the other hand, is directly related to pupation, as apyrene spermatogenesis begins 2 days after pupation in both control and allatectomized individuals.     

Expression of increased immunogenicity by thiol-releasing tumor variants.

Even moderate variations of the extracellular cysteine concentration were previously shown to affect T cell functions in vitro despite high concentrations of cystine. We therefore analyzed the membrane transport activities of T cells for cysteine and cystine, and the role of low molecular weight thiol in T cell-mediated host responses against a T cell tumor in vivo. A series of T cell clones and tumors including the highly malignant lymphoma L5178Y ESb and its strongly immunogenic variant ESb-D was found to express extremely weak transport activity for cystine but strong transport activity for cysteine. However, not all cells showed the expected requirement for cysteine (or 2-mercaptoethanol (2-ME)) in the culture medium. One group of clones and tumors including the malignant ESb-lymphoma did not respond to changes of extracellular cystine concentrations and was strongly thiol dependent. This group released only little acid soluble thiol (cysteine) if grown in cystine-containing cultures. The other T cell lines, in contrast, were able to maintain high intracellular GSH levels and DNA synthesis activity in cystine-containing culture medium without cystein or 2-ME and released substantial amounts of thiol. This group included the immunogenic ESb-D line. Additional thiol-releasing ESb variants were obtained by culturing large numbers of L5178Y ESb tumor cells in cultures without cysteine or 2-ME. All of these ESb variants showed a significantly decreased tumorigenicity and some of them induced cytotoxic and protective host responses even against the malignant ESb parent tumor. Taken together, our experiments suggest that the host response against a tumor may be limited in certain cases by the failure of the stimulator (i.e., the tumor) cell to deliver sufficient amounts of cysteine to the responding T cells.     

Purification and functional characterization of the human beta 2-adrenergic receptor produced in baculovirus-infected insect cells.

A human cDNA fragment bearing the complete coding region for the beta 2-adrenergic receptor was introduced into the genome of Autographa california nuclear polyhedrosis virus under the control of the polyhedrin promoter. Binding studies using [125I]iodocyanopindolol showed that Sf9 insect cells infected with the recombinant virus expressed approximately 1 x 10(6) beta 2-adrenergic receptors on their cell surface. Photoaffinity labeling of whole cells and membranes revealed a molecular weight of approximately 46,000 for the expressed receptor. The receptor produced in insect cells is glycosylated but the extent and pattern differ from that of the receptor from human tissue. The heterologously expressed receptor was purified by alprenolol affinity chromatography, and was able to activate isolated Gs-protein.     

A 45-kDa human T-cell membrane glycoprotein functions in the regulation of cell proliferative responses

A 45-kDa human T cell surface glycoprotein which is tightly bound in the membrane of the resting T cell is released into the cell medium in soluble form after cell growth activation by phytohemagglutinin or neuraminidase/galactose oxidase treatments. In limited proteolysis by Staphylococcus aureus V8 protease, two major 35-kDa and 27-kDa peptide fragments of the surface-iodinated 45-kDa protein are common to the membrane-bound and the released forms, but a third 18-kDa fragment is observed exclusively with the released protein. The apparent molecular masses of the deglycosylated peptide backbones of the membrane-bound and the released molecule are 30 +/- 1 kDa, although a small size difference cannot be excluded. A polyclonal rabbit anti-(T cell membrane protein) antiserum precipitates the 45-kDa protein. A monoclonal anti-(45-kDa protein) antibody precipitates the membrane-bound 45-kDa protein solubilized with octyl glucoside, but does not precipitate the released protein. In cell culture assays, the monoclonal anti-(45-kDa protein) antibody specifically enhances the cell proliferative responses in phytohemagglutinin-treated and mixed lymphocyte cultures. These observations suggest that the 45-kDa protein has a specific receptor function in the regulation of cell proliferative responses.     

Low concentrations of acid-soluble thiol (cysteine) in the blood plasma of HIV-1-infected patients

Blood plasma samples from HIV-1-infected persons contain elevated glutamate concentrations up to 6-fold the normal level and relatively low concentrations of acid-soluble thiol (i.e. decreased cysteine concentrations). The intracellular glutathione concentration in peripheral blood-mononuclear cells (PBMC) and monocytes from HIV antibody-positive persons are also significantly decreased. Therapy with azidothymidine (AZT) causes a substantial recovery of the plasma thiol levels; but glutamate levels remain significantly elevated and intracellular glutathione levels remain low. Cell culture experiments with approximately physiological amino-acid concentrations revealed that variations of the extracellular cysteine concentration have a strong influence on the intracellular glutathione level and the rate of DNA synthesis [( 3H]thymidine incorporation) in T cell clones and human and murine lymphocyte preparations even in the presence of several-fold higher cystine and methionine concentrations. Cysteine cannot be replaced by a corresponding increase of the extracellular cystine or methionine concentration. These experiments suggest strongly that the low cysteine concentration in the plasma of HIV-infected persons may play a role in the pathogenetic mechanism of the acquired immunodeficiency syndrome.     

Phylogenesis of spermatogenesis in Annulipalpia caddisflies: An ultrastructural analysis on philopotamidae spermiogenesis

Characters currently used in phylogenetic analyses are insufficient for determining the status of several families of the order Trichoptera (caddisflies). Comparative spermatology can contribute to solving this problem. In the suborder Integripalpia, the sperm axoneme displays the pattern of 9 + 2 pairs of parallel microtubules found in many animal species. In the suborder Annulipalpia, however, axonemes bear remarkable aberrations. Consistently, in Chimarra florida (Philopotamidae, Annulipalpia) we found that the number of central microtubules varies from zero to four in axonemes of the spermatids and that spermatozoa lack axonemes. We propose that, similarly, the axoneme pattern became unstable in the Annulipalpia ancestor, from which branched two phylogenetic lines having different kinds of axoneme aberrations: (a) families in which the number of central microtubules of the axoneme exceeds two (e.g., Philopotamidae, Polycentropodidae) and (b) families in which the number of central microtubules of the axoneme does not exceed two (e.g., Hydropsychidae).     

The Bile Acid-Sensitive Ion Channel (BASIC) Is Activated by Alterations of Its Membrane Environment

The bile acid-sensitive ion channel (BASIC) is a member of the DEG/ENaC family of ion channels. Channels of this family are characterized by a common structure, their physiological functions and modes of activation, however, are diverse. Rat BASIC is expressed in brain, liver and intestinal tract and activated by bile acids. The physiological function of BASIC and its mechanism of bile acid activation remain a puzzle. Here we addressed the question whether amphiphilic bile acids activate BASIC by directly binding to the channel or indirectly by altering the properties of the surrounding membrane. We show that membrane-active substances other than bile acids also affect the activity of BASIC and that activation by bile acids and other membrane-active substances is non-additive, suggesting that BASIC is sensitive for changes in its membrane environment. Furthermore based on results from chimeras between BASIC and ASIC1a, we show that the extracellular and the transmembrane domains are important for membrane sensitivity.