Molecular cloning and sequence analysis of the promoter region of mouse cyclin D1 gene: implication in phorbol ester-induced tumour promotion

Cyclin D1 is a cell cycle regulatory protein, which acts as a growth factor sensor to integrate extracellular signals with the cell cycle machinery, p articularly during G1 phase of the cell cycle. Previous study using promoti on-sensitive JB6 mouse epidermal cells, an in vitro model of the promotion stage of multistage carcinogenesis, showed that the expression of cyclin D1 is stimulated in the presence (but not in the absence) of 12-O-tetradecano ylphorbol-13-acetate (TP,A) in these cells maintained under anchorage-indep endent culture conditions. In the present study, to explore the molecular b asis of this observation, the promoter region of mouse cyclin D1 gene was c loned and sequenced (GenBank accession number AF212040). Dot matrix compari son of mouse, human and rat promoter sequences indicated that the mouse pro moter is homologous to the human and more: so to the rat promoters. The mou se promoter, like human and rat promoters, lacks canonical TATA-box or TATA -like sequence, but it has one or possibly two initiator (Inr) or Inr-like sequences. Energy dot plot analysis predicted that the mouse promoter consi sts of three domains: (1) the 3' domain contains NF-kappa B response elemen t, cAMP-response element (CRE), Inr or Inr-like elements, Spl binding site and Oct 1. (2) the middle domain contains another Sp1 binding site, E-box a nd E2F binding site and (3) the 5' domain contains TPA-response element (TR E) and a tandem silencer element. The cyclin D1 promoter sequence of either promotion-sensitive or resistant JB6 mouse epidermal cells was, except for a few minor differences, essentially identical to the sequence determined for a mouse genomic clone. Since TPA is capable of stimulating the expressi on of cyclin D1 not only through TRE but also through CRE and NF-kappa B re sponse element in the promoter, we tentatively propose a sequence of events that possibly leads to TPA-induced, anchorage-independent synthesis of cyc lins D1 and A in the promotion-sensitive JB6 mouse epidermal cells.