Abstract
We report the characterization of an in vitro chromatin assembly system derived from Artemia embryos and its application to the study of AluI-113 satellite DNA organization in nucleosomes. The system efficiently reconstitutes chromatin templates by associating DNA, core histones, and H1. The polynucleosomal complexes show physiological spacing of repeat length 190 +/- 5 base pairs, and the internucleosomal distances are modulated by energy-using activities that contribute to the dynamics of chromatin conformation. The assembly extract was used to reconstitute tandemly repeated AluI-113 sequences. The establishment of preferred histone octamer/satellite DNA interactions was observed. In vitro, AluI-113 elements dictated the same nucleosome translational localizations as found in vivo. Specific rotational constraints seem to be the central structural requirement for nucleosome association. Satellite dinucleosomes showed decreased translational mobility compared with mononucleosomes. This could be the consequence of interactions between rotationally positioned nucleosomes separated by linker DNA of uniform length. AluI-113 DNA led to weak cooperativity of nucleosome association in the proximal flanking regions, which decreased with distance. Moreover, the structural properties of satellite chromatin can spread, thus leading to a specific organization of adjacent nucleosomes.
Highlights
Most higher eukaryotic DNA is folded into a dynamic nucleoprotein structure, which is subject to progressive and reversible modifications of its condensation state during transitions between interphasic and metaphasic chromatin
In order to determine the structural properties of polynucleosomal complexes on multimeric AluI-113 fragments, we developed and characterized a cell-free assembly system from Artemia at the nauplius embryo stage
To investigate the chromatin organization of A. franciscana AluI-113 DNA in vivo, we performed Micrococcal Nuclease (MNase) and restriction enzyme digestions on nuclei obtained by homogenization of embryos at the nauplius stage
Summary
Most higher eukaryotic DNA is folded into a dynamic nucleoprotein structure, which is subject to progressive and reversible modifications of its condensation state during transitions between interphasic and metaphasic chromatin. There are chromosomal regions that maintain cytological properties comparable with those of the metaphase chromosome throughout the cell cycle [1] Termed heterochromatin, these highly condensed regions consist of simple DNA sequences repeated in long tandem arrays and are typically localized around centromeres and telomeres The study of the role of heterochromatic DNA on chromatin structure may help to clarify how specific heterochromatic structures are maintained To this end, we reconstituted and characterized the chromatin features of chromosomal regions that are cytologically distinguishable as Artemia franciscana (Crustacea Phyllopoda) heterochromatin. In order to determine the structural properties of polynucleosomal complexes on multimeric AluI-113 fragments, we developed and characterized a cell-free assembly system from Artemia at the nauplius embryo stage We have used this to analyze: (i) the organization of bent AluI-113 DNA into nucleosomes; (ii) the effects of interactions between consecutive physiologically spaced AluI-113 nucleosomes; and (iii) how AluI-113. Chromatin Organization of A. franciscana Satellite DNA sequences could affect the chromatin structure of non-satellite flanking regions
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
