Identification of a nuclear matrix-associated region of the c-myc protooncogene and its recognition by a nuclear protein in the human leukemia HL-60 cell line.
A nuclear matrix (NM)-associated region (MAR) of the protooncogene c-myc is identified in a human leukemia cell line (HL-60). A binding assay between isolated NM and 32P-end-labeled c-myc fragments in the presence of unlabeled competitors was used, and a 3'-end DraI/DraI fragment of 172 base pairs containing the first of the two polyadenylation [poly(A)] signals was identified as an in vitro MAR. Direct detection of endogenous c-myc fragments remaining NM bound after restriction digestion was used, and an in vivo MAR has been identified as the ClaI/EcoRI 1.4-kilobase pair fragment containing the 172-base pair in vitro MAR fragment. In addition, a nuclear protein (Mr = 25,000, p25) demonstrating preferential binding to the 172-base pair c-myc MAR has been identified and partially purified. This protein is diminished in the nuclei of the cells induced by phorbol ester to undergo macrophage differentiation. Footprint analysis shows that p25 binds to two regions of the 172-base pair fragment. One contains the first of two poly(A) addition signals and a topo II box-like sequence, and the other (AATTTCAATCCTAGTA) is 17 base pairs downstream of the first poly(A) signal.
Department of Anatomy, Hahnemann University, Philadelphia, Pennsylvania 19102.
R H
RH
Chou
J R
JR
Churchill
M M
MM
Flubacher
D E
DE
Mapstone
J
J
Jones
D001345:Autoradiography; D001483:Base Sequence; D015139:Blotting, Southern; D015153:Blotting, Western; D002460:Cell Line; D015342:DNA Probes; D004273:DNA, Neoplasm / Q000032:analysis*; D004268:DNA-Binding Proteins / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D015473:Leukemia, Promyelocytic, Acute / Q000235:genetics* / Q000473:pathology; D008969:Molecular Sequence Data; D015530:Nuclear Matrix / Q000032:analysis; D009687:Nuclear Proteins / Q000032:analysis*; D011518:Proto-Oncogene Proteins / Q000032:analysis*; D016271:Proto-Oncogene Proteins c-myc
D015342:DNA Probes; D004273:DNA, Neoplasm; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D016271:Proto-Oncogene Proteins c-myc