Functional Analysis of the Human Cyclin D2 and Cyclin D3 Promoters

Abstract

The D-type cyclins promote progression through the G 1 phase of the cell cycle and may provide a link between growth factors and the cell cycle machinery. We determined the nucleotide sequence of the 5-flanking region of the human cyclin D2 and cyclin D3 genes and identified the transcription start sites. Analysis of the upstream sequences required for transcription of the cyclin D2 and cyclin D3 genes in continuously dividing cells revealed marked differences in their regulatory elements. In the cyclin D2 gene positive elements were localized between positions ؊306 and ؊114 relative to the ATG codon at ؉1. Additional positive elements were localized between ؊444 and ؊345, whereas sequences that reduced transcription were identified between nucleotides ؊1624 and ؊892. In the cyclin D3 gene all of the positive elements required for maximal transcription were localized between nucleotides ؊366 and ؊167, and no negative elements were found. The activities of a reporter gene linked to the upstream regulatory sequences of the cyclin D2 gene but not the cyclin D3 gene were induced when starved cells were serum stimulated. This suggests that although the abundance of both the cyclin D2 and cyclin D3 mRNAs is increased by serum stimulation, only the cyclin D2 gene is up-regulated at the transcriptional level. Sequences between nucleotides ؊306 and ؊1624 of the cyclin D2 gene were necessary for serum inducibility.

Cell Type-specific E2F Activation and Cell Cycle Progression Induced by the Oncogene Product Tax of Human T-cell Leukemia Virus Type I

“…The KpnI-XbaI fragment was isolated and inserted into the KpnI-NheI sites of pGL2-Basic, generating pCycD3-Luc(Ϫ1319). Sequencing of these fragments identified nucleotide substitutions at nucleotides Ϫ885 (G to C), Ϫ1583 (G to C), and Ϫ1584 (C to G) in the cyclin D2 promoter and a nucleotide substitution at nucleotide Ϫ51 (T to C) and nucleotide insertions of C between nucleotides Ϫ80 and Ϫ81 and of G between nucleotides Ϫ318 and Ϫ319 in the cyclin D3 promoter, respectively, compared with sequences published previously (63).…”

Increased dosage of Runx1/AML1 acts as a positive modulator of myeloid leukemogenesis in BXH2 mice

“…To verify the implication of the candidate cooperative genes mentioned above and summarized in Figure 5a, we cloned human cyclin D3 promoter spanning À14 to À1014 bp relative to the ATG codon at þ 1 containing three putative Runx binding sites and three putative STAT binding sites (Brooks et al, 1996) in front of the luciferase gene and carried out a reporter assay. The cotransfection of RUNX1/PEBP2b and STAT3 into HL60 myeloid leukemia cells revealed more than twofold higher activation compared to that of RUNX1/PEBP2b alone (Figure 5b, lane 6 versus lane 3) and the synergy was completely lost by introduction of a point mutation in the DNA-binding domain of either gene, R174Q in RUNX1 (Michaud et al, 2002) or R414A, R417A in STAT3 (Fagerlund et al, 2002) (lanes 7 and 8, respectively).…”

A Permissive Role for Phosphatidylinositol 3-Kinase in the Stat5- mediated Expression of Cyclin D2 by the Interleukin-2 Receptor

“…In the human cyclin D2 gene, the E boxes reportedly bound by Myc-Max are located at positions Ϫ1596 bp (E box 1) and Ϫ1425 bp (E box 2) relative to the translational start site (38,40). We have previously shown via deletional analysis of a human cyclin D2 luciferase reporter gene that the region between Ϫ1624 and Ϫ1303 bp is not required for IL-2-inducible reporter gene activity (31).…”