Changes in structural organization of the nucleosome fiber during protooncogene activation.
Dynamics of structural changes in the chromatin of rat liver cells were studied during activation of the c-myc, c-fos, and c-jun protooncogenes. Nuclei were isolated at various stages of gene activation using cycloheximide and chromatin was subjected to partial nucleolysis by endogenous Ca2+/Mg2+-DNAse and extracted with low magnesium buffer; the nucleosome fragmentation was analyzed. DNA was isolated from the soluble chromatin and low-molecular-weight fragments were separated by electrophoresis through a composite gel system (3 and 1% agarose). This modification of DNA electrophoresis protocol improves the efficiency of separation of the low-molecular-weight DNA fragments. Increased protooncogene activity is associated with decreased content of DNAse-sensitive soluble chromatin. The fractional content of the c-fos gene is increased in the soluble chromatin. In control nuclei, endogenous Ca2+/Mg2+-DNAse cleaves mononucleosomes containing DNA fragments with variable length. Upon protooncogene activation, mono- and oligonucleosomes with elongated DNA fragments are predominantly formed. The changes involve about 40% of the nuclear chromatin. The data suggest that protooncogene activation is associated with reorganization at the nucleosome level of the structural organization of the chromatin.
Terent'ev
A A
AA
Institute of Chemical Physics, Russian Academy of Sciences, Chernogolovka, Moscow Region, 142432 Russia. boikov@icp.ac.ru
Kostyuk
G V
GV
Boikov
P Y
PY
D000818:Animals; D002843:Chromatin / Q000378:metabolism*; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D004706:Endodeoxyribonucleases / Q000378:metabolism; D005786:Gene Expression Regulation; D008099:Liver / Q000378:metabolism*; D008297:Male; D009690:Nucleic Acid Conformation; D009707:Nucleosomes / Q000378:metabolism*; D011487:Protein Conformation; D011519:Proto-Oncogenes; D051381:Rats; D017208:Rats, Wistar; D015533:Transcriptional Activation
D002843:Chromatin; D009707:Nucleosomes; D004247:DNA; D004706:Endodeoxyribonucleases; C029367:calcium magnesium dependent endodeoxyribonuclease