Differential role for Sp1/Sp3 transcription factors in the regulation of the promoter activity of multiple cyclin-dependent kinase inhibitor genes

Abstract

Cyclin-dependent kinase inhibitors play a significant role in cell cycle progression and in cellular differentiation and their expression is regulated in different cellular settings. GC-rich regions in the promoter sequences of the cyclin-dependent kinase inhibitor genes p15INK4B and p21CIP1/WAF1 mediate the transcriptional response of these genes to extracellular stimuli. Similar GC-rich sequences in the promoter of the p15INK4A and p16INK4B gene can be targeted for transcriptional inactivation by methylation of cytosine residues. GC-rich regions represent putative target sites for binding of the ubiquitously expressed Sp1 and Sp3 transcription factors. Using a combination of functional and biochemical studies, we analyzed the potential role of the Sp1 and Sp3 factors in the regulation of CDKI p15, p16, and p21 promoter activities. Using transient reporter gene assays, we determined that Sp1 is a strong activator of these promoters, whereas Sp3 functions as a weak transactivator. We have identified multiple protein-binding sites in the proximal promoter sequences of these genes by footprinting analysis. Some of these sites are bound by Sp1 and Sp3, as demonstrated by gel-shift experiments using Sp1/Sp3-specific antibodies, permitting the demonstration that a differential role exists for Sp1 and Sp3 in the regulation of the activity of these promoters.