Modulated binding of SATB1, a matrix attachment region protein, to the AT-rich sequence flanking the major breakpoint region of BCL2.
The t(14,18) chromosomal translocation that occurs in human follicular lymphoma constitutively activates the BCL2 gene and disrupts control of apoptosis. Interestingly, 70% of the t(14,18) translocations are confined to three 15-bp clusters positioned within a 150-bp region (major breakpoint region or [MBR]) in the untranslated portion of terminal exon 3. We analyzed DNA-protein interactions in the MBR, as these may play some role in targeting the translocation to this region. An 87-bp segment (87MBR) immediately 3' to breakpoint cluster 3 was essential for DNA-protein interaction monitored with mobility shift assays. We further delineated a core binding region within 87MBR: a 33-bp, very AT-rich sequence highly conserved between the human and mouse BCL2 gene (37MBR). We have purified and identified one of the core factors as the matrix attachment region (MAR) binding protein, SATB1, which is known to bind to AT-rich sequences with a high propensity to unwind. Additional factors in nuclear extracts, which we have not yet characterized further, increased SATB1 affinity for the 37MBR target four- to fivefold. Specific binding activity within 37MBR displayed cell cycle regulation in Jurkat T cells, while levels of SATB1 remained constant throughout the cell cycle. Finally, we demonstrated in vivo binding of SATB1 to the MBR, strongly suggesting the BCL2 major breakpoint region is a MAR. We discuss the potential consequences of our observations for both MBR fragility and regulatory function.
Ramakrishnan
M
M
Division of Molecular Medicine, Beckman Research Institute of the City of Hope National Medical Center, Duarte, California 91010, USA.
Liu
W M
WM
DiCroce
P A
PA
Posner
A
A
Zheng
J
J
Kohwi-Shigematsu
T
T
Krontiris
T G
TG
D000595:Amino Acid Sequence; D000818:Animals; D020029:Base Pairing; D001483:Base Sequence; D001943:Breast Neoplasms; D002883:Chromosomes, Human, Pair 14; D002887:Chromosomes, Human, Pair 18; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D005091:Exons; D005260:Female; D019254:Genes, bcl-2; D006801:Humans; D019169:Jurkat Cells; D008224:Lymphoma, Follicular / Q000235:genetics; D036961:Matrix Attachment Region Binding Proteins; D051379:Mice; D008969:Molecular Sequence Data; D015530:Nuclear Matrix / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014162:Transfection; D014178:Translocation, Genetic; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; D036961:Matrix Attachment Region Binding Proteins; D010446:Peptide Fragments; D019253:Proto-Oncogene Proteins c-bcl-2; C076189:SATB1 protein, human