Unequal nuclear Sp1/GC box DNA binding activity distinguishes proliferating from differentiated senescent or apoptotic cells.
Terminal differentiation can result in either viable, non-proliferating or apoptotic cells. In B16 melanoma, millimolar L-tyrosine induces tyrosinase, a key enzyme for terminal pigmentation concurrent with either irreversible growth arrest at low cell density, or apoptosis at high cell density. Since the promoter for melanocyte-specific tyrosinase expression contains sites for the Sp1 transcription factor, we have investigated the relationship of Sp1-mediated GC-box DNA binding activity to growth control in undifferentiated and in terminally differentiated viable or apoptotic cells. Nuclear extracts from viable, differentiated cells showed increased retardation of GC box DNA sequence compared with that seen in proliferating cells or those reversibly arrested in early G(1) or late G(1) / S. In contrast, nuclear proteins from dying, differentiated cells showed loss of nuclear GC box DNA binding activity without decrease in binding to TTTGCGCG sequences recognized by the E2F transcription factor, which is known to interact with Sp1. However, cyto-plasmic fractions from apoptotic cells revealed phos-phatase-activated retardation of GC box DNA, which was not evident in similarly treated fractions from undifferentiated cells or sparse differentiated cells. Terminal differentiation also correlated with increase in a slow-migrating phosphorylated Sp1 isoform. Our data suggests that lack of nuclear Sp1/GC box DNA binding activity, may promote apoptosis by diminishing expression of survival-associated genes regulated by GC box DNA promoter sequences in dense terminally differentiated melanoma cells.
Rieber
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M
Instituto Venezolano de Investigaciones Científicas (IVIC), Centro de Microbiología y Biología Celular, Tumor Cell Biology, Caracas, Venezuela. mrieber@pasteur.ivic.ve
Strasberg Rieber
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D000818:Animals; D017209:Apoptosis; D002454:Cell Differentiation; D002455:Cell Division; D004247:DNA / Q000378:metabolism*; D053938:DNA Fragmentation; D008546:Melanoma, Experimental / Q000473:pathology; D051379:Mice; D014442:Monophenol Monooxygenase / Q000096:biosynthesis; D010766:Phosphorylation; D016329:Sp1 Transcription Factor / Q000378:metabolism*
D016329:Sp1 Transcription Factor; D004247:DNA; D014442:Monophenol Monooxygenase