DNA binding and nuclease protection by the HMf histones from the hyperthermophilic archaeon Methanothermus fervidus

The DNA-binding and nuclease-protection properties of the HMf histones from the hyperthermophilic archaeon Methanothermus fervidus have been shown to be consistent with the formation of nucleosome-like structures (NLS). These proteins bind to DNA molecules as short as 20 bp and form complexes that protect DNA fragments from micrococcal nuclease (MNase) digestion that are 30 bp, ∼ 60 bp and multiples of ∼ 60 bp in length. The sequences of 49 of the ∼ 60-bp DNA fragments protected from MNase digestion by HMfA have been determined and their intrinsic curvatures calculated. A circular permutation gel mobility-shift assay was used to determine directly the curvatures for five of these sequences. HMfA bound to intrinsically curved and noncurved DNAs, but exhibited a slight preference for the model curved DNA in binding competitions with a model noncurved DNA. The results obtained are consistent with the concept that the archaeal NLS is analogous, and possibly homologous, to the central core of the eukaryal nucleosome formed by a histone (H3 + H4)2 tetramer. Received: August 11, 1996 / Accepted: November 12, 1996     

The study of cellular “microexudates” by ellipsometry and their relationship to the cell coat

The use of ellipsometry, an optical technique for the detection of very thin films, to measure the thickness of the ‘microexudate’ deposited by mammalian cells on glass and other solid surfaces is described. Significant differences were found in the thickness and the rate of formation of microexudates in different cell types. Cultivation of cells at low temperatures and in medium supplemented with actinomycin D and cycloheximide inhibited the production of cellular microexudates, indicating that these materials are actively synthesized by the cell and do not result merely from passive leakage of macromolecular material as suggested previously. Evidence is reviewed to show that cellular microexudates have a number of properties in common with the cell coat material.     

Cell Division of Escherichia coli BUG-6: Effect of Varying the Length of Growth at the Nonpermissive Temperature

When Escherichia coli BUG-6 is grown at 42 C and then returned to 30 C, the division kinetics during the recovery at 30 C are dependent on the length of time the cells were grown at 42 C. If chloramphenicol is added when the cells are shifted from 42 to 30 C, no division occurs if the period at 42 C is less than 4 min or more than 110 min. Maximum division occurs when the period at 42 C is 50 min. A discussion of these results with reference to a previously proposed model is presented.     

Minicells of Bacillus subtilis: a New Bacteriophage-Blocking Agent

Bacteriophage SP01, SP17, and phi29 rapidly absorb irreversibly to minicells of Bacillus subtilis but do not produce a lytic cycle in minicells.     

Minicells of Bacillus subtilis

After nitrosoguanidine (N-methyl-N'-nitro-N-nitrosoguanidine) mutagenesis, two Bacillus subtilis mutants (div IV-A1 and div IV-B1) were isolated that are defective in the location of division site along cell length. Both mutations were transferred into strain CU403 by transformation, and their properties were studied in the CU403 genetic background. Location of divisions in close proximity to cell pole regions in both mutants results in minicell production. Purified minicells contain a ratio of ribonucleic acid to protein comparable to that found in the parent cells. Autoradiographs of (3)H-thymine incorporation into deoxyribonucleic acid (DNA), thymine-2-(14)C incorporation into DNA, electron micrographs, and chemical analyses for DNA all fail to demonstrate DNA in the minicells. Minicells produced by both mutants are highly motile, an indication of functional energy metabolism. Electron micrographs reveal that minicells are produced by a structurally normal division mechanism and that minicells contain a normal cell surface. The div IV-A1 mutation has been mapped by PBS1 transduction linked to ura. The div IV-B1 mutation is closely linked to pheA by both PBS1 transduction and by co-transformation.     

Rapid Preparation of Thick Sections of Fleshy Plant Tissues

Methods are described for permanent micro-slide preparations of soft, large-celled plant tissues such as ripe fruit. Thick sections (200-800 [t) cut on a sliding microtome are aspirated in an aqueous killing agent; after fixing and washing, the sections are dehydrated and cleared in an alcohol-xylene series. Infiltration with 20, 30, and 40% solutions of mountant prior to mounting the sections is necessary to avoid too abrupt changes in the cleared tissues. Several staining methods have been successfully used for different purposes. The final preparations showed nearly perfect preservation of intact cells and intercellular spaces in their 3-dimension-al structure.     

OxLDL-induced gene expression patterns in CASMC are mimicked in apoE-/- mice aortas

Oxidized low density lipoprotein (oxLDL) contributes to the pathophysiology of atherosclerosis, partly by altering gene expression in vascular cells. Here, we show 221 genes differentially regulated by oxLDL in coronary artery smooth muscle cells (CASMC), using oligonucleotide microarrays. These genes were classified into 14 functional groups. A comparable gene expression pattern was detected in apoE(-/-) mice. OxLDL induced an oxidative stress response in CASMC, but not the unfolded protein response. OxLDL also caused CASMC death which was accompanied by increased expression of FasL, Bax, and p53 but was caspase-independent. This approach provides further insight into disease pathology and prognosis.