Key Role of a Downstream Specificity Protein 1 Site in Cell Cycle-regulated Transcription of the AP Endonuclease GeneAPE1/APEX in NIH3T3 Cells

• Published in: The Journal of biological chemistry Vol. 276; no. 45; p. 42011
• Main Authors: Hua Fung, Richard A. O. Bennett, Bruce Demple
• Format: Journal Article
• Language: English
• Published: American Society for Biochemistry and Molecular Biology 09-11-2001
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M106423200

Abasic (apurinic/apyrimidinic or AP) sites are a frequent type of DNA damage that threatens genetic stability. The predominant
mammalian enzyme initiating repair of AP sites is the Ape1 AP endonuclease (also called Apex or Hap1), which also facilitates
DNA binding by several transcription factors (Ref1 activity). We found that expression of the APE1 gene was coordinated with the cell cycle in murine NIH3T3 cells: APE1 mRNA levels rose after the G 1 -S transition and peaked ∼4-fold higher in early to mid-S phase. The increased APE1 mRNA was the result of transcriptional activation rather than increased mRNA stability. Fusions of various APE1 promoter fragments to the chloramphenicol acetyltransferase CAT reporter gene indicated that APE1 expression depends on two transcription factor Sp1 binding sites within the promoter region. Mutation of these sites or of
two CCAAT elements within the APE1 promoter, in conjunction with protein binding studies, demonstrated their specific roles. The Sp1 site upstream of the transcription
start, together with an adjacent CCAAT element, establishes a protein-DNA complex required for basal transcription of APE1 . The Sp1 site downstream of the transcription start was required for the response to cell growth. Because Ape1 is a dual
function enzyme, its cell cycle-dependent expression might affect both DNA repair and the activity of various transcription
factors as a function of the cell cycle.