Sequence-specific binding of a c-myc nuclear-matrix-associated region shows increased nuclear matrix retention after leukemic cell (HL-60) differentiation.
HL-60 cells, a human promyelocytic leukemia cell line, contain amplified c-myc DNA sequences and mRNA transcripts. These cells can be induced to undergo macrophage differentiation by phorbol esters, which results in suppression of c-myc expression and cessation of cell proliferation. The nuclear matrix (NM), a nuclear skeleton resistant to DNase I digestion and high salt extraction, is proposed to be involved in DNA replication, gene regulation, and the correct distribution of DNA at mitosis. We have previously identified a nuclear-matrix-associated region (MAR) of the c-myc protooncogene to reside in a 1.4-kb region between Cla I and Eco RI restriction sites at the 3'-end of the gene. A 172-bp Dra I/Dra I subfragment of the 1.4-kb region was shown to be a major component of the MAR (myc-MAR), and this subfragment was demonstrated to be recognized by a nuclear protein (p25). In this report we demonstrate that phi X174 DNA, or the synthetic copolymers poly[d(G.C)] and poly[d(A.T)], are not effective suppressors of the binding of the myc-MAR to isolated NM, indicating that the binding sequence(s) are unique. We find that the addition of partially purified protein p25 increases the relative affinity of the myc-MAR for HL-60 NM in an in vitro assay system. NM isolated from HL-60 macrophages induced by phorbol esters retains significantly more myc-MAR DNA fragment in the presence of an excess amount of competitor DNA than does NM from untreated HL-60 cells. These data suggest that a change of the myc-MAR association with the NM occurs after monocytic differentiation of HL-60 cells.
Department of Anatomy, Hahnemann University, Philadelphia, Pennsylvania 19102.
R H
RH
Chou
J R
JR
Churchill
D E
DE
Mapstone
M M
MM
Flubacher
D001482:Base Composition; D001665:Binding Sites / Q000187:drug effects; D001667:Binding, Competitive; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D016259:Genes, myc; D006801:Humans; D015530:Nuclear Matrix / Q000378:metabolism*; D010703:Phorbol Esters / Q000494:pharmacology; D015183:Restriction Mapping
D010703:Phorbol Esters