Joanne M. Nickol

Higher order structure of chromatin: Orientation of nucleosomes within the 30 nm chromatin solenoid is independent of species and spacer length

We have used electric dichroism to study the arrangement of nucleosomes in 30 nm chromatin solenoidal fibers prepared from a variety of sources (CHO cells, HeLa cells, rat liver, chicken erythrocytes, and sea urchin sperm) in which the nucleosome spacer length varies from approximately 10 to approximately 80 bp. Field-free relaxation times are consistent only with structures containing 6 +/- 1 nucleosomes for every 11 nm of solenoidal length. With very few assumptions about the arrangement of the spacer DNA, our dichroism data are consistent with the same orientation of the chromatosomes for every chromatin sample examined. This orientation, which maintains the faces of the radially arranged chromatosomes inclined at an angle between 20 degrees-33 degrees to the solenoid axis, thus appears to be a general structural feature of the higher order chromatin fiber.

Erythroid-specific activation and derepression of the chick β-globin promoter in vitro

Transcriptionally active extracts were prepared from chick red cells isolated at different stages of development. The template activity of cloned beta-globin genes is highest in extracts from definitive red cells, where the endogenous gene is normally expressed, and lowest in extracts from primitive red cells or nonerythroid tissues. This system has been used to identify regulatory elements and to assign functions to the proteins that bind within the beta-globin promoter. Regulation of expression is achieved, in part, by factors whose composition changes during red cell development. Two proteins, PAL and CON, bind at adjacent sites but have opposite effects on transcription in vitro. Levels of PAL, a potent repressor, are highest in mature red cells while those of CON, an activator, are highest in actively transcribing red cells. The effect of PAL can be overcome by blocking its binding site with a protein having a similar recognition sequence but a dissimilar function.

Crystallization of a DNA tridecamer d(C-G-C-A-G-A-A-T-T-C-G-C-G)☆

Crystals of the DNA tridecamer d(C-G-C-A-G-A-A-T-T-C-G-C-G) have been grown by the vapor-diffusion technique with 2-methyl-2,4-pentanediol as precipitant. They are monoclinic space group C2, with a = 79.6 A, b = 43.1 A, c = 24.9 A and beta = 98.7 degrees. Previous nuclear magnetic resonance studies predicted that this tridecamer forms a duplex similar to the B DNA dodecamer, d(C-G-C-G-A-A-T-T-C-G-C-G), except for an extra adenosine residue that is stacked within the helix but remains unpaired: (formula; see text) Preliminary X-ray diffraction studies confirmed that the tridecamer is in the B DNA conformation, consistent with the nuclear magnetic resonance results.

CHROMATIN CONFORMATION AND GENE ACTIVITY

ABSTRACT The physical properties of chromatin have been explored at the level of individual nucleosomes, extended polynucleosome filaments, and folded polynucleosome fibers. To compare the physical properties of bulk chromatin with the properties of chromatin in transcriptionally active genes, we have examined the stability of the 30 nm chromatin fiber in the neighborhood of the adult s-globin gene in chicken red cells. We find that the DNA in the neighborhood of this gene is indistinguishable from bulk chromatin in its degree of compaction and in its reaction with the HMG proteins HMG 14/17.