Identification of a complex of the three forms of the rat liver asialoglycoprotein receptor

We have generated antibodies against synthetic peptides which represent the carboxyl terminus of either the major, or the two minor, forms of the rat hepatic lectin which recognizes galactose-terminated glycoproteins (asialoglycoproteins). The antibodies were shown to be specific for the form of the lectin containing the immunizing peptide sequence by the following: reaction with purified lectin after sodium dodecyl sulfate-polyacrylamide gel electrophoresis, immunoprecipitation of sodium dodecyl sulfate-denatured lectin, immunoprecipitation of lectin synthesized in vitro. These antibodies, however, precipitated all three rat hepatic lectin forms from nonionic detergent extracts of hepatocytes labeled with 125I via the lactoperoxidase catalyzed technique. A similar result was obtained if antibody was bound to intact cells prior to solubilization with detergent and collection of the immune complexes. We conclude that at least the plasma membrane-associated fraction of the rat hepatic lectin forms exists as a heterotypic complex.     

Effect of iodide on estrogen-induced uterine peroxidase

Iodide administered in the drinking water for 5–7 days increased the activity of estradiol-induced uterine peroxidase in the immature rat. This effect was specific for iodide and could not be mimicked by chloride, bromide, thiocyanate, perchlorate or iodate. Sodium iodide also increased peroxidase activity in the parotid gland but had no effect on glucose-6-phosphate dehydrogenase in the uterus, thyroid or parotid even though estradiol produced a 2-fold increase in the activity of this enzyme in the uterus. ¹²⁵I was taken up more readily by the uterus than by muscle but this process was not influenced by prior treatment of the animals with estrogen. The in vitro effect of sulfhydryl reagents on uterine peroxidase was also investigated and proposals made for possible mechanisms of action of iodide on this enzyme in the intact animal.     

Role of methyl groups in dynamics and evolution of biomolecules

Recent studies have discovered strong differences between the dynamics of nucleic acids (RNA and DNA) and proteins, especially at low hydration and low temperatures. This difference is caused primarily by dynamics of methyl groups that are abundant in proteins, but are absent or very rare in RNA and DNA. In this paper, we present a hypothesis regarding the role of methyl groups as intrinsic plasticizers in proteins and their evolutionary selection to facilitate protein dynamics and activity. We demonstrate the profound effect methyl groups have on protein dynamics relative to nucleic acid dynamics, and note the apparent correlation of methyl group content in protein classes and their need for molecular flexibility. Moreover, we note the fastest methyl groups of some enzymes appear around dynamical centers such as hinges or active sites. Methyl groups are also of tremendous importance from a hydrophobicity/folding/entropy perspective. These significant roles, however, complement our hypothesis rather than preclude the recognition of methyl groups in the dynamics and evolution of biomolecules.     

Diminished Reovirus Capsid Stability Alters Disease Pathogenesis and Littermate Transmission

Reovirus is a nonenveloped mammalian virus that provides a useful model system for studies of viral infections in the young. Following internalization into host cells, the outermost capsid of reovirus virions is removed by endosomal cathepsin proteases. Determinants of capsid disassembly kinetics reside in the viral σ3 protein. However, the contribution of capsid stability to reovirus-induced disease is unknown. In this study, we found that mice inoculated intramuscularly with a serotype 3 reovirus containing σ3-Y354H, a mutation that reduces viral capsid stability, succumbed at a higher rate than those infected with wild-type virus. At early times after inoculation, σ3-Y354H virus reached higher titers than wild-type virus at several sites within the host. Animals inoculated perorally with a serotype 1 reassortant reovirus containing σ3-Y354H developed exaggerated myocarditis accompanied by elaboration of pro-inflammatory cytokines. Surprisingly, unchallenged littermates of mice infected with σ3-Y354H virus displayed higher titers in the intestine, heart, and brain than littermates of mice inoculated with wild-type virus. Together, these findings suggest that diminished capsid stability enhances reovirus replication, dissemination, lethality, and host-to-host spread, establishing a new virulence determinant for nonenveloped viruses.     

Effect of trypsin on the cell surface proteins of hepatoma tissue culture cells. Characterization of a carbohydrate-rich glycopeptide released from a calcium binding membrane glycoprotein.

Concentrations of trypsin that bring about aggregation of hepatoma tissue culture (HTC) cells also release from the cell surface an Mr = 55,000 glycopeptide fragment. This glycopeptide fragment also accumulates in the medium, including serum-free medium, as a normal consequence of membrane protein turnover. The trypsin-released glycopeptide is labeled when cells are grown in the presence of fucose or leucine before treatment of the cells with the protease. Similarly, the glycopeptide fragment can be labeled by reacting cells in situ by lactoperoxidase-catalyzed radioiodination or by tritiated borohydride reduction of cells treated first with neuraminidase and galactose oxidase. The tryptic glycopeptide fragment was purified by concanavalin A-Sepharose chromatography, and hydroxyapatite chromatography in the presence of dodecyl sulfate. The amino acid and carbohydrate composition was determined, as was the sensitivity of the purified glycopeptide to a variety of endo- and exoglycosidases. The purified glycopeptide contains an average of 17 sialic acid residues and hence, shows charge heterogeneity after electrophoresis in isoelectric focusing gels. The charge heterogeneity can be eliminated completely by treatment with neuraminidase. The glycopeptide after this treatment is homogeneous. The trypsin-sensitive membrane glycoprotein which is the source of the Mr = 55,000 glycopeptide was identified by two-dimensional gel electrophoretic analysis of labeled cells, treated or not treated with trypsin. This glycoprotein, which has an apparent molecular weight of 85,000 and forms a homodimer in the presence of calcium ions, was purified and its identity as the parent of the Mr = 55,000 glycopeptide was confirmed by showing that the same Mr = 55,000 fragment was released by trypsin from the purified glycoprotein as was released from the intact cells.     

Potential phylogenetic utility of the low-copy nuclear gene pistillata in dicotyledonous plants: comparison to nrDNA ITS and trnL intron in Sphaerocardamum and other Brassicaceae.

We report the potential phylogenetic utility of DNA sequence data from the last 700 bp of a ca. 1-kb intron of the MADS-box gene pistillata from a sampling of Sphaerocardamum species and other Brassicaceae. These results are compared with nrDNA ITS and the chloroplast trnL intron for the same taxa to demonstrate the potential phylogenetic utility of this pistillata intron and to identify potential historically independent sequences for an ongoing study of relationships within Sphaerocardamum. Analyses of the DNA sequence data for Brassicaceae indicated that pairwise divergences and potentially informative characters were higher in the pistillata intron (0.6-30.8%, 284 characters) and ITS (0-24%, 94 characters) than in the chloroplast trnL intron (0-4.2%, 17 characters). A comparison of Sphaerocardamum sequences identified low divergences and numbers of informative characters for trnL intron (0-2.4%, 1 character) and nrDNA ITS (0-2.5%, 2 characters) and substantially more variation among the pistillata sequences (0.15-3.7%, 19 characters). Phylogenetic analyses of these pistillata sequences fully resolve ingroup relationships without character conflict. Results of pistillata PCR amplifications from a broader dicot sample showed that some primers may be useful in amplifying orthologous pistillata sequences. Ultimately this pistillata intron may be a valuable source of phylogenetic characters at lower taxonomic levels.     

Speciation and the evolution of behaviour in prosimians

A mechanism is suggested, based on a biphasic approach-withdrawal theory proposed byT. C. Schneirla, to account for how behaviours may be selected by environmental forces for transmission over generations. The basic postulates of Schneirla's theory are presented followed by an examination of some aspects of lorisid behaviour in the light of this theory.     

Structural and functional variability among DQ β alleles of DR2 subtypes

Homozygous lymphoblastoid cell lines representing various Dw subtypes of DR2 were examined for polymorphism at the DQ locus by molecular and cellular techniques. The subtypes studied included Dw2, Dw12, and a group heterogenous by cellular typing that we shall refer to as non-Dw2/non-Dw12. Restriction fragment length polymorphism analysis of cell lines representing these subtypes revealed DQ -specific patterns consistent with cellular typing. Two-dimensional gel electrophoresis of DQ molecules from representative cell lines revealed a structural polymorphism of DQ among the three subtypes. The DQ chain migrated to a position that was unique to each subtype and was consistent among various representative cell lines of each subtype. Nucleotide sequence analysis of cDNA clones of DQ from Dw2, Dw12, and non-Dw2/non-Dw12 lines confirmed that the variability resided at the genetic level. Variability was found in the form of numerous scattered nucleotide substitutions throughout the first domain of these alleles. The DQ gene of the non-Dw2/non-Dw12 cell line AZH was further found to be almost identical with the DQ gene of a DR1 line (Bell et al. 1985b), implicating a common evolutionary origin of these alleles. The only difference between these two sequences was due to an apparent gene conversion event at amino acid 57. T-cell cloning experiments resulted in the derivation of Epstein-Barr virus-specific, DQw1-restricted clones that proliferated against only those cell lines that exhibited the DQ gene common to AZH and the DR1 cell line. Thus, the polymorphism among DQ alleles within DR2 results in subtype-specific restriction.