Autologous antibodies to human bladder cancer

Summary The autologous serologic reactivity of 13 patients with bladder cancer was evaluated using cell lines derived from each individual's own tumor as targets. Protein A and immune adherence assays were employed to determine antibody binding to the tumor targets at varying passage numbers. Autologous reactivity was found in 6 of the 13 cell lines tested. However, the titer was usually low regardless of the passage number. Seven autologous serum/cell line combinations were tested using both low and high passage cells as targets. In six of these combinations, the degree of antibody binding was similar with both low and high passage target cells. The incidence of autologous reactivity in the 12 patients with urothelial tumors was 50%.This investigation was supported by PHS Grant number CA36933, awarded by the National Cancer Institute, DHHS.     

Family resemblance for serum uric acid in a Jerusalem sample of families

Summary Familial aggregation of serum uric acid was studied in a sample of families examined in the Jerusalem Lipid Research Clinic. We first examined homogeneity of familial correlations across the major origin groups in the Israeli population sample. In general correlations were homogeneous across origin groups, except for spouse pairs. Pooled correlations among biological relatives across the origin groups were all statistically significant. Spouse correlation upon adjustment for concomitant variables was moderately positive (r=0.115), yet significantly different from zero. Genetic and cultural determinants of uric acid were estimated utilizing a path model with 10 parameters to be estimated from a total of 16 correlations. Under a reduced model, genetic heritability (h²) was estimated to be 0.47±0.05 and cultural heritability (c²) was 0.11±0.03. However, our data gave suggestive evidence that cultural heritability was higher in parents (c²=0.28) than in children (c²=0.10). Commingling analysis and segregation analysis were also performed, and our findings imply that in the Israeli population there is no evidence for a major gene for high uric acid levels segregating in families.     

The E beta hot spot of recombination in wild-derived natural recombinant MHC haplotypes. Cross-over site mapping and the identification of a 1.0-kb E beta deletion in the p and w14 haplotypes

Detailed molecular analysis of three wild-derived MHC haplotypes provided evidence for an important role of the E beta recombinational hot spot in the recent evolution of the mouse I region. Examination of RFLP and restriction maps of cloned DNA permitted the mapping of the natural cross-over events in the haplotypes carried by strains B10.GAA37 (w21) and B10.KPB128 (w19) to a fragment of DNA not exceeding 4.1 kb, which lies almost entirely within the intron separating the beta 1 and beta 2 exons of the E beta gene. In the w14 haplotype (strain B10.STC77), which appears to be a natural recombinant between a p-like parental haplotype and another wild-derived haplotype, the site of crossing over can be mapped to a segment between the beta 2 exon of the E beta gene (left border) and the E beta 2 gene (right border). This segment containing the cross-over site in the w14 haplotype includes the E beta hot spot. In addition, the w14 haplotype as well as the standard p haplotype contain a deletion of approximately 1.0 kb in the second intron of the E beta gene, which may represent the product of an unequal cross-over event in a E beta recombinational hot spot.     

Generation of different nucleosome spacing periodicities in vitro. Possible origin of cell type specificity

We have been able to generate ordered nucleosome arrays that span the physiological range of spacing periodicities, using an in vitro system. Our system (a refinement of the procedure previously developed) uses the synthetic polynucleotide poly[d(A-T)], poly[d(A-T)], core histones, purified H1, and polyglutamic acid, a factor that increases nucleohistone solubility and greatly promotes the formation of ordered nucleosome arrays. This system has three useful features, not found in other chromatin assembly systems. First, it allowed us to examine histones from three different cell types/species (sea urchin sperm, chicken erythrocyte, and HeLa) as homologous or heterologous combinations of core and H1 histones. Second, it allowed us to control the average packing density (core histone to polynucleotide weight ratio) of nucleosomes on the polynucleotide; histone H1 is added in a second distinct step in the procedure to induce nucleosome alignment. Third, it permitted us to study nucleosome array formation in the absence of DNA base sequence effects. We show that the value of the spacing periodicity is controlled by the value of the initial average nucleosome packing density. The full range of physiological periodicities appears to be accessible to arrays generated using chicken erythrocyte (or HeLa) core histones in combination with chicken H5. However, chromatin-like structures cannot be assembled for some nucleosome packing densities in reactions involving some histone types, thus limiting the range of periodicities that can be achieved. For example, H1 histone types differ significantly in their ability to recruit disordered nucleosomes into ordered arrays at low packing densities. Sea urchin sperm H1 is more efficient than chicken H5, which is more efficient than H1 from HeLa or chicken erythrocyte. Sea urchin sperm core histones are more efficient in this respect than the other core histone types used. These findings suggest how different repeat lengths arise in different cell types and species, and provide new insights into the problems of nucleosome linker heterogeneity and how different types of chromatin structures could be generated in the same cell.     

Introns of the chicken ovalbumin gene promote nucleosome alignment in vitro.

A defined in vitro chromatin assembly system was used to examine the nucleosome alignment induced by histone H5 throughout a 12 kilobase pair chicken genomic DNA fragment containing the ovalbumin gene. In contrast with total fragmented chicken DNA and several anonymous cloned fragments, much of the gene permitted histone H5 to space nucleosomes at physiological intervals in an extended array. Nucleosomes at the 3'-end of the gene and on approximately 4 kilobase pairs of 5'-flanking ovalbumin sequence did not become aligned to appreciable extents. Analysis of cloned 2-3 kilobase pair subfragments suggested that a strong nucleosome alignment signal, specifying a 196 +/- 5 base pair repeat exists in intron E. A second discrete region of the gene, which mapped approximately to intron A, exhibited nucleosome alignment with a spacing periodicity of about 200 base pairs. The ovalbumin cDNA did not permit nucleosome alignment. These findings suggest that some of the introns contain signals that direct nucleosome alignment over the ovalbumin gene in a way conducive to its regulation.     

Isolation and identification of granule-associated proteins relevant for poly(3-hydroxyalkanoic acid) biosynthesis in Chromatium vinosum D

Abstract Poly(3-hydroxybutyric acid) granules, which harbored only four major granule-associated proteins as revealed by SDS polyacrylamide gel electrophoresis, were isolated from crude cellular extracts of Chromatium vinosum D by centrifugation in a linear sucrose gradient. N-Terminal amino acid sequence determination identified two proteins of M _r 41 000 and M _{r 40 000} as the phaE _Cv and phaC _Cv translational products, respectively, of C. vinosum D. In a previous study it was shown that both proteins are required for the expression opf poly(3-hydroxyalkanoic acid) synthase activity. The N-terminus of the third protein ( M _r 17 000) exhibited no homology to other proteins. Lysozyme, which was during purification of the granules, exhibited a strong affinity to PHB granules and was identified as the fourth protein enriched with the granules.     

A human histone H2B.1 variant gene, located on chromosome 1, utilizes alternative 3' end processing.

A variant human H2B histone gene (GL105), previously shown to encode a 2300 nt replication independent mRNA, has been cloned. We demonstrate this gene expresses alternative mRNAs regulated differentially during the HeLa S3 cell cycle. The H2B-Gl105 gene encodes both a 500 nt cell cycle dependent mRNA and a 2300 nt constitutively expressed mRNA. The 3' end of the cell cycle regulated mRNA terminates immediately following the region of hyphenated dyad symmetry typical of most histone mRNAs, whereas the constitutively expressed mRNA has a 1798 nt non-translated trailer that contains the same region of hyphenated dyad symmetry but is polyadenylated. The cap site for the H2B-GL105 mRNAs is located 42 nt upstream of the protein coding region. The H2B-GL105 histone gene was localized to chromosome region 1q21-1q23 by chromosomal in situ hybridization and by analysis of rodent-human somatic cell hybrids using an H2B-GL105 specific probe. The H2B-GL105 gene is paired with a functional H2A histone gene and this H2A/H2B gene pair is separated by a bidirectionally transcribed intergenic promoter region containing consensus TATA and CCAAT boxes and an OTF-1 element. These results demonstrate that cell cycle regulated and constitutively expressed histone mRNAs can be encoded by the same gene, and indicate that alternative 3' end processing may be an important mechanism for regulation of histone mRNA. Such control further increases the versatility by which cells can modulate the synthesis of replication-dependent as well as variant histone proteins during the cell cycle and at the onset of differentiation.     

Benzodiazepines in the brain

Great progress has been made in the last 5 yr in demonstrating the presence of benzodiazepines (BDZs) in mammalian tissues, in beginning studies on the origin of these natural compounds, and in elucidating their possible biological roles. Many unanswered questions remain regarding the sources and biosynthetic pathways responsible for the presence of BDZs in brain and their different physiological and/or biochemical actions. This essay will focus on recent findings supporting that: (1) BDZs are of natural origin; (2) mammalian brain contains BDZs in concentrations ranging between 5.10⁻¹⁰–10⁻⁸ M; (3) dietary source of BDZs might be a plausible explanation for their occurrence in animal tissues, including man; (4) the formation of BDZ-like molecules in brain is a possibility, experimentally supported; (5) BDZ-like molecules including diazepam andN-desmethyldiazepam are elevated in hepatic encephalopathy; and (6) natural BDZs in the brain are involved in the modulation of memory processes. Future studies using the full range of biochemical, physiological, behavioral, and molecular biological techniques available to the neuroscientist will hopefully continue to yield exciting and new information concerning the biological roles that BDZs might play in the normal and pathological functioning of the brain.