Glomerular lesions in Aleutian disease of mink (Mustela vison): a morphological and differential morphometrical study.

A morphological and morphometrical study has been carried out on glomerular lesions in mink with spontaneous Aleutian disease, using the WHO classification for Systemic Lupus Erytematous Nefritis. 154 renal samples from sick animals and 10 samples from uninfected mink were processed by routine histopathological techniques and metacrylate inclusions. The samples were studied quantitatively with an automatic image analyzer. 5 forms of glomerulonephritis (GN) were identified: mesangial glomerulonephritis (n = 13), focal and segmental GN (n = 10), diffuse GN (n = 99), membranous GN (n = 12) and advanced sclerosing GN (n = 10) and were associated with the degree of interstitial plasmocytosis. Glomerule morphometry was shown to be an excellent method for identifying the type of lesion, while it quantified the participation of various glomerular elements in the lesion.     

Isolation and partial characterization of a protein kinase NII from wheat germ chromatin

A protein kinase, type NII, has been purified from wheat germ chromatin. The enzyme, which uses both ATP and GTP as phosphoryl donors, catalyzes the phosphorylation of casein, phosvitin and E. coli RNA polymerase, but not of histone proteins. Polypeptide bands at 46 kDa, 37 kDa and 25 kDa were estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Autophosphorylation of the 25 kDa subunit was observed following incubation of the purified kinase with (-³²P)ATP and (-³²P)GTP.     

Analysis of X-chromosome inactivation and presumptive expression of the Wiskott-Aldrich syndrome (WAS) gene in hematopoietic cell lineages of a thrombocytopenic carrier female of WAS

Summary We report on a thrombocytopenic female belonging to a pedigree with the Wiskott-Aldrich syndrome (WAS). Restriction fragment length polymorphism (RFLP) analysis with probe M27, closely linked to the WAS gene, demonstrated that she is a carrier of WAS. Both small-sized and normal-sized platelets were present, suggesting that, unlike the vast majority of WAS carriers, she does not manifest nonrandom X-chromosome inactivation in the thrombopoietic cell lineage. Study of X-chromosome inactivation by means of RFLP and methylation analysis demonstrated that the pattern of X-chromosome inactivation was nonrandom in T lymphocytes, but random in granulocytes. While this is the first complete report on the occurrence of thrombocytopenia in a carrier female of WAS as the result of atypical lyonization, it also suggests that expression of the WAS gene occurs at (or extends up to) a later stage than the multipotent stem cell along the hematopoietic differentiation pathway.     

Human erythrocyte glucose-6-phosphate dehydrogenase. Identification of a reactive lysyl residue labelled with pyridoxal 5'-phosphate

Human erythrocyte glucose-6-phosphate dehydrogenase contains a reactive lysyl residue, which can be labelled with pyridoxal 5'-phosphate. The binding of one mole of pyridoxal 5'-phosphate per mole of enzyme subunit produces substantial inactivation. The substrate glucose-6-phosphate prevents the loss of activity, suggesting that the reaction site is close to the substrate-binding site. A tryptic peptide containing the pyridoxal-5'-phosphate-binding lysyl residue has been isolated and characterised. The reactive lysyl residue has been identified in the glucose-6-phosphate dehydrogenase amino acid sequence. Comparison with glucose-6-phosphate dehydrogenase from other sources shows a high homology with a peptide containing a reactive lysyl residue, isolated from the enzyme from Saccharomyces cerevisiae; glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides also contains a region highly homologous with the sequence around the reactive lysyl residue in the human enzyme. The results of this communication provide the first direct evidence for the association of an essential catalytic function with a specific region of the molecule of human erythrocyte glucose-6-phosphate dehydrogenase.     

Leishmania major and L. donovani: effects on proteolytic enzymes of Phlebotomus papatasi (Diptera, Psychodidae)

Phlebotomus papatasi is susceptible to Leishmania major which it transmits in nature, but is resistant to L. donovani. The present study compares the effect of L. major and L. donovani on the proteolytic activity of P. papatasi gut enzymes. The experiments measured digestion of C14-labeled globin by gut homogenates of flies. Homogenates were prepared from flies fed on serum only (controls) or from flies fed serum containing promastigotes or their dried culture overlayer. In other experiments, the promastigotes or dried culture overlayers were added in vitro to the gut homogenate of control flies. Proteolytic activity of gut homogenate from flies infected with L. major was about one-third less than that of controls, while that from flies infected with L. donovani was one-third greater. Ingestion of L. major dried culture overlayer had an effect on flies similar to that of the promastigotes, while L. donovani dried culture overlayer produced no significant effect. When added to gut homogenate in vitro, promastigotes of both species promoted proteolysis as did dried culture overlayer of L. major. Dried culture overlayer of L. donovani, however, had an opposite effect. It is suggested that the observed reduction in proteolytic activity caused by L. major infection may result from inhibition of enzyme production.     

Restriction endonuclease mapping of ribosomal RNA genes: Sequence divergence and the origin of the tetraploid treefrog Hyla versicolor

Hyla chrysoscelis (2n=24) and H. versicolor (2n=48) are a diploid-tetraploid species pair of treefrogs. Restriction endonuclease mapping of ribosomal RNA (rRNA) gene repeat units of diploids collected from eastern and western populations reveals no differences within rRNA gene coding regions but distinctive differences within the nontranscribed spacers. A minimum of two physical maps is required to construct an rRNA gene map for the tetraploid, whose repeat units appear to be a composite, with about 50% of the elements resembling the western diploid population and about 50% resembling the eastern population. These results imply that this population of the tetraploid species may have arisen from a genetically hybrid diploid. Alternatively, the dual level of sequence heterogeneity in H. versicolor may reflect some type of gene flow between the two species. The coding region of the rRNA genes in the tetraploid differs from that in either diploid in about 20% of all repeat units, as exemplified by a BamHI site located near the 5 terminus of the 28 S rRNA gene. If the 20% variant class of 28 S rRNA gene coding sequences is expressed, then there must be two structural classes of ribosomes; if only the 80% sequence class is expressed, then a genetic control mechanism must be capable of distinguishing between the two different sequence variants. It is postulated that the 20% variant sequence class may be correlated with a partial functional diploidization of rRNA genes in the tetraploid species.This research was supported, in part, by NSF Grants CDP-8002341 and PRM-8106947 and by faculty research grants from Miami University to J.C.V.     

Human chorionic gonadotropin binding to rat testis receptors is inhibited by a thymus factor

An interrelationship between immune and reproductive systems has been postulated, and involves, among others, bidirectional effects between gonads and thymus. To this effect a rat thymus fraction of about 28000 mol wt has been reported to inhibit the effect of hCG on in vitro suspension of Leydig cells. We have investigated the antigonadotropin activity of thymus extracts on rat testis receptors. Acetonic powder obtained from thymus of 14 day-old rats was separated by molecular sieve chromatography. The effect of the collected fractions on the 125I-hCG binding to receptor sites in rat testes was evaluated. A fraction corresponding to 27000-28000 mol wt named thymus factor (TF), was found to inhibit the binding activity of 125I-hCG to its testicular receptor. The inhibitory effect of TF on hCG binding is dose related. By Scatchard analysis a competitive interaction at the receptor level between TF and hCG was demonstrated. The Ka values of hCG binding were diminished in the presence of TF while no significative changes were detected in the number of receptor sites. Present results strongly suggest a modulation function of TF at the testis receptor level.     

Contrasting effects of Escherichia coli single-stranded DNA binding protein on synthesis by T7 DNA polymerase and Escherichia coli DNA polymerase I (large fragment). Evidence that binding protein inhibits trans-lesion synthesis by polymerase I

The effect of Escherichia coli single-stranded DNA binding protein (SSB) on DNA synthesis by T7 DNA polymerase and E. coli DNA polymerase I (large fragment) using native or aminofluorene-modified M13 templates was evaluated by in vitro DNA synthesis assays and polyacrylamide gel electrophoresis analysis. The two polymerase enzymes displayed differential responses to the addition of SSB. T7 DNA polymerase, a enzyme required for the replication of the T7 chromosome, was stimulated by the addition of SSB whether native or modified templates were used. On the other hand, E. coli DNA polymerase I was slightly stimulated by the addition of SSB to the native template but substantially inhibited on modified templates. This result suggests that DNA polymerase I may be able to synthesize past an aminofluorene adduct but that the presence of SSB inhibited this trans-lesion synthesis. Polyacrylamide gels of the products of DNA synthesis by polymerase I supported this inference since SSB caused a substantial increase in the accumulation of shorter DNA chains induced by blockage at the aminofluorene adduct sites.