Location of nucleosomes in simian virus 40 chromatin.
Over the past decade, the results of numerous indirect mappings analyses have not clarified whether or not nucleosomes occupy preferred positions in simian virus 40 (SV40) chromatin. To address this question more directly, we followed a shotgun cloning approach and determined the nucleotide sequences of over 400 cloned nucleosomal DNA fragments obtained from digestion of SV40 chromatin with micrococcal nuclease. Our results demonstrate and establish that nucleosomes do not occupy unique positions in SV40 minichromosomes and thus indicate the existence of at least several types of chromatin molecules having different nucleosome organization patterns. We developed two types of statistical analysis in order to examine the cloning data in greater detail. One type, overlap analysis, revealed the distribution of the cloned fragments with respect to SV40 DNA. The distribution exhibits an oscillating pattern, dividing the genome into regions of weak or strong nucleosome density. The other analysis determined the distribution of the midpoints of the cloned fragments and revealed potential strong and weak nucleosome location sites, and an early versus late distinction in organization of nucleosomes in SV40 chromatin. The late region appears to contain more strong nucleosome location sites (8) than the early region (4). The strongest nucleosome abuts the late side of the nuclease-hypersensitive region and includes the major transcription initiation site of the late genes. Another strong site precedes this nucleosome and includes sequences implicated in controlling the expression of the SV40 early and late genes. A strong or weak nucleosome location site is not apparent near the early side of the nucleosome-hypersensitive region. Only weak and overlapping nucleosome location sites are found in the region where replication terminates in the SV40 minichromosomes.
Ambrose
C
C
Purdue University, Department of Chemistry, West Lafayette, IN 47907.
Lowman
H
H
Rajadhyaksha
A
A
Blasquez
V
V
Bina
M
M
D000818:Animals; D002460:Cell Line; D002843:Chromatin / Q000032:analysis*; D003001:Cloning, Molecular; D004279:DNA, Viral / Q000235:genetics / Q000302:isolation & purification; D009707:Nucleosomes / Q000032:analysis*; D015183:Restriction Mapping; D013539:Simian virus 40 / Q000235:genetics*; D014158:Transcription, Genetic
D002843:Chromatin; D004279:DNA, Viral; D009707:Nucleosomes