Molecular processes of chromosome 9p21 deletions in human cancers

Abstract

Interstitial deletions of the chromosome 9p21 segment encoding the p16/CDKN2A tumor suppressor gene (i.e., 9p21 deletions) are frequently observed in a variety of human cancers. A majority of these deletions in lymphoid leukemia have been indicated to be mediated by illegitimate V(D)J recombination. In the present study, to elucidate the molecular processes of 9p21 deletions in nonlymphocytic malignancies, breakpoints for these deletions were analysed in 21 lung cancer cell lines and 32 nonlymphocytic cancer cell lines of nine other histological types. In all, 32 breakpoints in 21 lung cancer cell lines and 56 breakpoints in 32 nonlung cancer cell lines were mapped in a 450-kb segment encompassing the CDKN2A locus with a 10-kb resolution. The largest number of breakpoints (i.e., seven breakpoints in lung cancer and 12 breakpoints in nonlung cancers) was mapped in a 10-kb region containing the CDKN2A gene. More precise mapping of these seven and 12 breakpoints revealed that none of these breakpoints were located within 50-bp intervals to each other in this 10 kb region. Cloning and sequencing of breakpoints in 18 representative cell lines (six lung and 12 nonlung cancers) further revealed that there were no significant homologies among breakpoints in these 18 cell lines. In 11 (61%) cell lines, 1-5-bp nucleotides were overlapped at breakpoint junctions. These results indicate that DNA double-strand breaks triggering 9p21 deletions do not occur at specific DNA sequences, although they preferentially occur in or near the CDKN2A locus. It was also indicated that two broken DNA ends are rejoined by nonhomologous end-joining repair, preferentially utilizing microhomologies of DNA ends, in the occurrence of 9p21 deletions. Oncogene (2003) 22, 3792-3798. doi:10.1038/sj.onc.1206589Keywords: 9p21 deletion; p16/CDKN2A; lung cancer; DNA double-strand break; nonhomologous end joining Chromosome 9p deletions frequently occur in a variety of human cancers including lymphoid leukemia, lung cancer, esophageal cancer, glioma and melanoma, and the 9p21 segment including the p16/CDKN2A, p14/ARF and p15/CDKN2B loci has been defined as a common region for the deletions (Kamb et al., 1994;Nobori et al., 1994;Ohnishi et al., 1995;Tanaka et al., 1997;Drexler, 1998;Hamada et al., 1998Hamada et al., , 2000Park et al., 2002). The CDKN2A tumor suppressor gene and two other related genes, ARF and CDKN2B, encode critical regulators of cell cycle and/or apoptosis (Sherr, 2000). Thus, deletion of the 9p21 segment is a causative generic alteration inactivating the CDKN2A, ARF and/or CDKN2B genes, and is thought to play an important role in the development and/or progression of human cancers. Elucidation of the mechanism(s) for the deletions of the 9p21 segment is, therefore, indispensable in understanding the molecular pathways of multistage human carcinogenesis.Structural analyses of breakpoints for interstitial deletions of the 9p21 segment (i.e., 9p21 deletions) have been performed in lymphoid leukemia to predict the molecular processes of th...

Probing the chromosome 9p21 region susceptible to DNA double-strand breaks in human cells in vivo by restriction enzyme transfer

“…Therefore, the chromatin for the BCSs was thought to take a configuration(s) making them susceptible to digestion by DNA endonucleases such as restriction enzymes and RAG proteins. Breakpoints for 9p21 deletions in nonhematological malignancies were not mapped at specific DNA sequences (Florl and Schulz, 2003;Sasaki et al, 2003;Raschke et al, 2005). Therefore, genomic features enhancing DSBs were unclear.…”

“…Our previous studies indicated that DSBs triggering deletions in lymphoid leukemia are produced by the RAG complex, a DNA endonuclease used for V(D)J recombination, while those in other cancers are produced at unspecific sites by as yet unidentified factors (Kitagawa et al, 2002;Sasaki et al, 2003). Thus, the distributions were analysed not only in total but also after dividing into lymphoid leukemia and other cancers.…”

“…In particular, 16 (94%) breakpoints in lymphoid leukemia including 11 in the breakpoint cluster sites (BCSs), BCS-LL3 and -LL2 (Kitagawa et al, 2002), were mapped in the sensitive regions. A 10 kb region, where the largest number of the breakpoints in other cancers was mapped ('200-210 kb region' in Sasaki et al, 2003), was overlapped with an MspI-sensitive region including the p16 promoter region.…”

“…We previously mapped breakpoints for the interstitial 9p21 deletions removing the p16/p14ARF/p15 locus in a variety of human cancers (Kitagawa et al, 2002;Sasaki et al, 2003). To examine whether or not the chromosomal DNAs sensitive to MspI digestion are susceptible to DSBs during human carcinogenesis, these breakpoints were mapped on the MspI sensitivity map (Figure 5a).…”

“…We chose the chromosome 9p21 region containing the p16 tumor suppressor gene for this study, since this region is frequently deleted in a variety of human cancers (Kamb et al, 1994;Nobori et al, 1994;Hamada et al, 2000), and thus, the region was thought to be a representative one suffering from DSBs in human cells. In addition, we recently mapped a number of breakpoints for the deletions in a variety of human cancers (Kitagawa et al, 2002;Sasaki et al, 2003); therefore, the detailed information on breakpoint locations needed for the comparison was available.…”