The histone H1°/H5 variant and terminal differentiation of cells during development of Xenopus laevis

The maintenance of the differentiated condition is supposed to be associated with the presence of a histone of the H1(0)/H5 subclass. If the H1(0)/H5 variant has an important role in differentiation distinct from that of H1, it should display differential expression in time and position during development. Here we report that this prediction is verified during Xenopus laevis development, in which tadpoles exhibit a very characteristic, developmentally regulated pattern of histone H1(0)/H5 expression that is different for the derivatives of each embryonic germ layer. However, the pattern of appearance of this variant during development does not reflect a simple correlation between its presence and the state of differentiation. Therefore, these results are pertinent to current ideas on differentiation and the involvement of lysine-rich histones in the repression of eukaryotic genes.     

CD3/T-cell receptor coupling to a pertussis and cholera toxin-insensitive G-protein

We have analyzed the effect of CD3/T-cell receptor stimulation on GTP hydrolysis and GTP binding. We show that stimulation of Jurkat, T-cell, membranes with OKT3 results in a 50% increase in GTP hydrolysis which is specifically inhibited by GDP. Pretreatment of the membranes with neither pertussis toxin nor cholera toxin inhibited the GTP hydrolysis. We also show that stimulation with OKT3 increases the binding of GTPγS to Jurkat membranes. These data strongly implicate the involvement of a G-protein in CD3/T-cell receptor signalling.     

Physical identification of branched intron side-products of splicing in Trypanosoma brucei.

Every mRNA in trypanosomes consists of two exons, a common 5' capped mini-exon or spliced leader and a coding-exon. All evidence suggests that the exons are joined by trans-splicing of two individual precursor RNAs, the mini-exon donor RNA or spliced leader precursor RNA (medRNA) and the pre-mRNA. We studied intermediates of the splicing reaction using denaturing two-dimensional PAGE and structurally identified a group of small (approximately 180-300 nt) non-polyadenylated, Y-shaped branched RNAs. The branched Y-shaped RNAs contain the 105 nt medRNA derived intron, joined in a 2'-5' phosphodiester bond to small heterogeneously sized RNAs. These non-polyadenylated branched Y-shaped RNA molecules are analogous to the lariat shaped introns of higher eukaryotes and presumably represent the released intron-like by-products of a trans-splicing reaction which joins the mini-exon and the major coding-exon.     

In situ hybridization as a tool to study numerical chromosome aberrations in solid bladder tumors

Summary Methods for single- and double-target in situ hybridization (ISH) to, cells isolated from solid transitional cell carcinomas (TCC's) of the urinary bladder are described. Single cell suspensions were prepared from solid tumors of the urinary bladder by mechanical disaggregation and fixed in 70% ethanol. Using two DNA probes specific for the centromeres of chromosomes #1 and #18, ISH procedures were optimized for these samples. Human lymphocytes and cells from the T24 bladder tumor cell line were used as controls. In lymphocyte nuclei and metaphase chromosome spreads, ISH showed two major spots for each of the probes. About 80% of the nuclei from T24 cells showed three spots for both the chromosome #1 and #18 specific probes. When nuclei from TCC's were analyzed, often the number of spots for chromosome #1, and to a lesser extent for chromosome #18, differed from the number expected on basis of flow cytometric ploidy measurements. The double target-ISH method in all cases allowed the correlation of numerical aberrations for chromosomes #1 and #18 in one and the same cell. By such analyses a profound heterogeneity in chromosome number was detected in most tumors. In order to optimize the reproductbility of the method and the interpretation of the ISH-signals, criteria for their analysis have been determined. This procedure can now be applied on a routine basis to solid tumor specimens.     

Localization of the nucleotide excision repair gene ERCC6 to human chromosome 10q11-q21.

We have cloned the human DNA excision repair gene ERCC6 by virtue of its ability to correct the uv sensitivity of Chinese hamster overy cell mutant UV61. This mutant is a member of complementation group 6 of the nucleotide excision repair-deficient rodent mutants. By means of in situ hybridization and Southern blot analysis of mouse x human somatic cell hybrids, the gene was localized to human chromosome 10q11-q21. An RFLP detected within the ERCC6 locus can be helpful in linkage analysis.