Spermidine-induced alteration in the gene-spacer discrimination of nucleases in protonated DNA.
The sequence preference of a Drosophila lysosomal DNase was studied on the Drosophila hsp 70 heat-shock and histone recombinants, which carry six different genes, and the surrounding spacer sequences. The distribution of cleavage sites was random in respect of the locations of gene and spacer sequences. However, in the presence of 10 mM spermidine, a major transition was observed: the coding sequences became more susceptible than the spacer regions to nuclease attack. A similar transition was induced in the sequence preference of DNase I if the digestion was performed in the presence of spermidine at pH 5.2. At pH 7.5, spermidine does not influence the sequence preference of DNase I, which indicates the involvement of DNA protonation in this transition. In the presence of spermidine, the distributions of preferred and protected sequences were almost indistinguishable for these nucleases, suggesting that the protonated DNA, and not the enzymes, is the target of spermidine. A Drosophila embryonal protein was detected and partially purified which induced the same transition as observed in the presence of spermidine. The purified protein preferentially protected the spacer DNA sequences against acid DNase or DNase I cleavage in the hsp 70 heat-shock and histone gene recombinants. The protection was concentration dependent and occurred only at pH 5.2. The transition of nuclease specificity is probably due to a conformational change in the protonated DNA, induced by the binding of either the embryonal protein or spermidine.
Gaszner
M
M
Institute of Biochemistry, Biological Research Center of the Hungarian Academy of Sciences, Szeged.
Haracska
L
L
Udvardy
A
A
D000818:Animals; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D003850:Deoxyribonuclease I / Q000378:metabolism; D003851:Deoxyribonucleases / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000196:embryology / Q000201:enzymology; D017344:Genes, Insect; D006657:Histones / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D008247:Lysosomes / Q000201:enzymology; D009690:Nucleic Acid Conformation / Q000187:drug effects; D011506:Proteins / Q000302:isolation & purification / Q000494:pharmacology; D013095:Spermidine / Q000494:pharmacology*; D013379:Substrate Specificity / Q000187:drug effects
D006657:Histones; D011506:Proteins; D004247:DNA; D003851:Deoxyribonucleases; D003850:Deoxyribonuclease I; D013095:Spermidine