c-Myc initiates illegitimate replication of the ribonucleotide reductase R2 gene

Abstract

The mechanisms through which the oncoprotein c-Myc initiates locus-speci®c gene ampli®cation are not understood. When analysing the initiation mechanism of cMyc-dependent ampli®cation of the mouse ribonucleotide reductase R2 (R2) gene, we observe c-Myc-dependent initiation of illegitimate DNA replication of the R2 gene. We demonstrate multiple simultaneous c-Myc-induced R2 replication forks, whereas R2 normally replicates with a single fork. In contrast, cyclin C replicates with only a single replication fork irrespective of c-Myc deregulation. In addition to de novo replication forks, cMyc also initiates bi-allelic replication of R2, abrogating its normal mono-allelic replication pattern. Moreover, several chromosomal regions also display c-Myc-induced illegitimate replication pro®les. Thus, c-Myc can act as an illegitimate replication-licensing factor that promotes de novo replication initiation and illegitimate replication timing that adversely impacts upon genomic stability. Oncogene (2002) 21, 909 ± 920.

Inhibitory effect of c-Myc on p53-induced apoptosis in leukemia cells. Microarray analysis reveals defective induction of p53 target genes and upregulation of chaperone genes

“…This function has been recently found to play a prominent role in the tumor suppressor activity of p53 in vivo (Liu et al, 2004). Also, it is becoming increasingly clear that one consequence of deregulated MYC expression is genomic instability, which results in unscheduled DNA replication, gene rearrangements and chromosomal aberrations (Li and Dang, 1999;Kuschak et al, 2002;Sheen and Dickson, 2002;Karlsson et al, 2003). Thus, our data also support the hypothesis that deregulated MYC contributes to genomic instability in cells arrested by wild-type conformation p53.…”

c-Myc-Induced Genomic Instability

“…MYC acted as an illegitimate replication-licensing factor. We concluded at this time that the types of R2 amplification observed as a result of MYC deregulation were in agreement with replication-driven instability (Kuschak et al , 2002. Other genes that undergo MYC-dependent gene amplification, such as Cyclin D2 and DHFR, also follow a replication-dependent mechanism (SF Louis and S Mai, unpubl.…”

“…Work by our group next showed that MYC was able to promote illegitimate DNA replication resulting in more than one replication firing per origin per cell cycle (Kuschak et al 2002). Interestingly, in diploid synchronized PreB mouse lymphocytes, after a single pulse of conditional MYC deregulation, the R2 gene was replicated more than once in early S phase (Kuschak et al 2002).…”

“…Interestingly, in diploid synchronized PreB mouse lymphocytes, after a single pulse of conditional MYC deregulation, the R2 gene was replicated more than once in early S phase (Kuschak et al 2002). Using two-dimensional gel electrophoresis of DNA harvested before and throughout S phase as well as mitotic chromosomes from BrdU-pulse labeled cells, we found that MYC was able to deregulate the normal "once per cycle" replication initiation of R2 that was observed in the control cells and for the control gene cyclin C. MYC forced three to four replication initiation forks and replication proceeded on both alleles, not just in a monoallelic fashion as seen in control cells (Kuschak et al 2002). MYC acted as an illegitimate replication-licensing factor.…”

“…The affected genes include ribonucleotide reductase R2 (R2) , the carbamyl-P synthetase, aspartate transcarbamylase, dihydro-orotase (CAD) enzyme-encoding gene (Miltenberger et al 1995;Fukasawa et al 1997;Chernova et al 1998;Eberhardy and Farnham 2001), ornithine decarboxylase (George et al 1996;Rounbehler et al 2009), cyclin B1, and cyclin D2 . The amplification of genes involved in DNA synthesis and cell-cycle progression will provide a proliferative advantage to cells that harbor it (Kuschak et al 2002). Not only will cells carrying such amplifications experience a growth advantage, but they will also be resistant to some of the cytotoxic drugs such as MTX (Denis et al 1991) or even radiation (Lücke-Huhle et al 1997).…”