Differential protein binding in lymphocytes to a sequence in the enhancer of the mouse retrovirus SL3-3.
An electrophoretic mobility shift assay was used to characterize interactions of nuclear proteins with a DNA segment in the enhancer element of the leukemogenic murine retrovirus SL3-3. Mutation of a DNA sequence of the 5'-TGTGG-3' type decreased transcription in vivo specifically in T-lymphocyte cell lines. Extracts of nuclei from different T-lymphocyte cell lines or cells from lymphoid organs resulted in much higher amounts of complexes in vitro with this DNA sequence than did extracts from other cell lines or organs tested. Differences were also found in the sets of complexes obtained with extracts from the different types of cells. The DNA sequence specificities of the different SL3-3 enhancer factor 1 (SEF1) protein complexes were found to be distinct from those of several other previously identified DNA motifs of the TGTGG type because of differences in several nucleotides critical for binding and because these other DNA motifs could not compete with the identified DNA sequence for binding of SEF1. Limited treatment with several different proteases cleaved the SEF1 proteins such that their DNA-binding domain(s) remained and created complexes with decreased and nondistinguishable electrophoretic mobility shifts and with new properties. These results indicate that the SEF1 proteins have a structure with a flexible and relatively vulnerable hinge region linking a DNA-binding domain(s) to a more variable domain(s) with other functions. We suggest that the binding of SEF1 is an essential factor for the T-cell tropism of SL3-3 and the ability of this virus to cause T-cell lymphomas.
Thornell
A
A
Unit for Applied Cell and Molecular Biology, University of Umeå, Sweden.
Hallberg
B
B
Grundström
T
T
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004742:Enhancer Elements, Genetic; D005814:Genes, Viral; D006801:Humans; D009052:Leukemia Virus, Murine / Q000235:genetics*; D008214:Lymphocytes / Q000378:metabolism*; D008745:Methylation; D009154:Mutation; D009687:Nuclear Proteins / Q000378:metabolism*; D010957:Plasmids; D014158:Transcription, Genetic
D009687:Nuclear Proteins