Chromatosome positioning on assembled long chromatin. Linker histones affect nucleosome placement on 5 S rDNA

Long chromatin containing linker histones H1 or H5 was assembled on tandemly repeated 172 or 207 base-pair nucleosome positioning sequences from a sea urchin 5 S RNA gene. The effects of H1 and H5 on spacing and positioning of nucleosomes were assessed. In the absence of linker histones, precise determinations of core particle boundaries showed that, although a large proportion of the histone octamers occupy a unique position, there is a small group of other, less populated sites located around this major site. The dominant position was found 10 to 15 base-pairs upstream from the unique position previously reported for the histone octamer on the monomer 260 base-pair sequence. Linker histones do not override the underlying DNA signals that induce the very regular spacing of nucleosomes in chromatins assembled on these strongly positioning multimer DNA sequences. They were nevertheless found to be decisive in determining the chromatosome positions and their distributions, and as such define the chromatosome as a positioning entity.     

Analysis of the slow phases of the in vivo chlorophyll fluorescence induction curve. Changes in the redox state of Photosystem II electron acceptors and fluorescence emission from Photosystems I and II

An analysis of the photo-induced decline in the in vivo chlorophyll a fluorescence emission (Kautsky phenomenon) from the bean leaf is presented. The redox state of PS II electron acceptors and the fluorescence emission from PS I and PS II were monitored during quenching of fluorescence from the maximum level at P to the steady state level at T. Simultaneous measurement of the kinetics of fluorescence emission associated with PS I and PS II indicated that the ratio of PS I/PS II emission changed in an antiparallel fashion to PS II emission throughout the induction curve. Estimation of the redox state of PS II electron acceptors at given points during P to T quenching was made by exposing the leaf to additional excitation irradiation and determining the amount of variable PS II fluorescence generated. An inverse relationship was found between the proportion of PS II electron acceptors in the oxidised state and PS II fluorescence emission. The interrelationships between the redox state of PS II electron acceptors and fluorescence emission from PS I and PS II remained similar when the shape of the induction curve from P to T was modified by increasing the excitation photon flux density. The contributions of photochemical and non-photochemical quenching to the in vivo fluorescence decline from P to T are discussed.     

Activity banding of human chromosomes as shown by histone acetylation

The expression of genes in mammalian cells depends on many factors including position in the cell cycle, stage of differentiation, age, and environmental influences. As different groups of genes are expressed, their packaging within chromatin changes and may be detected at the chromsomal level. The organization of DNA within a chromosome is determined to a large extent by the positively charged, highly conserved histones. Histone subtypes and the reversible chemical modifications of histones have been associated with gene activity. Active or potentially active genes have been associated with hyperacetylated histones and inactive genes with nonacetylated histones. Sodium butyrate increases the acetylation levels of histones in cell cultures and acts as both an inducer of gene activity and as a cell-cycle block. We describe a method to label the interphase distribution of DNA associated with various histone acetylation stages on chromosomes. Nucleosomes from untreated and butyrate-treated HeLa cells were fractionated by their acetylation level and the associated DNA labeled, and hybridized to normal human chromosomes. In the sodium butyrate-treated cells the resulting banding patterns of the high- and low-acetylated fractions were strikingly different. DNA from low-acetylated chromatin labeled several pericentric regions, whereas hybridization with DNA from highly acetylated chromatin resulted in a pattern similar to inverse G-bands on many chromsomes. The results from noninduced cells at both high and low acetylation levels were noticeably different from their induced counterparts. The capture and hybridization of DNA from interphase chromatin at different acetylation states provides a “snap-shot” of the distribution of gene activity on chromosomes at the time of cell harvest. Edited by: P.B. Moens     

Studies on the role and mode of operation of the very-lysine-rich histone H1 (F1) in eukaryote chromatin. The conformation of histone H1.

Proton magnetic resonance, circular dichroism and other studies of whole and cleaved calf thymus histone H1 (formerly F1) reveal the presence of specific folded structures in the region approximately from residue 40--115. Ionic, hydrogen-bond and hydrophobic interactions all appear to contribute to the stability of the structure, which is predicted to contain alpha-helices in regions 42--55 and 58--75. No evidence was found for beta-structures, either inter or intramolecular, or for any structure formation outside the region 40--115. At 18 degrees C and a protein concentration of 2 mM the first-order exchange rate between random-coil and structured forms is slower than 80 s-1; at 40 degrees C the exchange rate is faster than 330 s-1.     

Flux Analysis of Glucose Metabolism in Rhizopus oryzae for the Purpose of Increasing Lactate Yields

A flux analysis of glucose metabolism in the filamentous fungus Rhizopus oryzae was achieved using a specific radioactivity curve-matching program, TFLUX. Glycolytic and tricarboxylic acid cycle intermediates labeled through the addition of extracellular [U-14C]glucose were isolated and purified for specific radioactivity determinations. This information, together with pool sizes and the rates of glucose utilization and end product production, provided input for flux maps of the metabolic network under two different experimental conditions. Based upon the flux analysis of this system, a mutant of R. oryzae with higher lactate and lower ethanol yields than the parent was sought for and found.     

Nuclear magnetic resonance and optical spectroscopic studies of copolymers of polypeptides. II. Random copoly(benzyl-l-glutamate:benzyl-L-aspartate) and (benzyl-D-glutamate:benzyl-L-aspartate)

High-resolution NMR is used to study two series of benzyl-L - or D -glutamate: benzyl-L -aspartate random copolymers. The helix sense of the L -aspartate residues determined from the αCH and NH chemical shifts agrees with that obtained from ORD. The stability of the helical copolymers to TFA addition shows a minimum at the composition of helix-sense inversion for the L -glutamate:L -aspartate copolymers but no minimum for the lefthanded D -glutamate: L -aspartate series. The helix-coil transition of the glutamate and aspartate residues in each polymer is compared. For the D -glutamate: L -aspartate series no differences are found, indicating random copolymerization. In the L -glutamate:L -aspartate series significant differences between the transition midpoints are interpreted as resulting from irregular distribution of component residues along the chain.