Copy number evolution in simple and complex tandem repeats across the C57BL/6 and C57BL/10 inbred mouse lines

Simple sequence tandem repeats are among the most rapidly evolving compartments of the genome. Some repeat expansions are associated with mammalian disease or meiotic segregation distortion, yet the rates of copy number change across generations are not well known. Here, we use 14 distinct sub-lineages of the C57BL/6 and C57BL/10 inbred mouse strains, which have been evolving independently over about 300 generations, to estimate the rates of copy number changes in genome-wide tandem repeats. Rates of change varied across repeats and across lines. Notably, CAG, whose expansions in coding regions are associated with many neurological and genetic disorders, was highly stable in copy number, likely indicating stabilizing selection. Rates of change were positively correlated with copy number, but the direction and magnitude of changes varied across lines. Some mouse lines experienced consistent losses or gains across most simple repeats, but this did not correlate with copy number changes in complex repeats. Rates of copy number change were similar between simple repeats and the more abundant complex repeats after normalization by copy number. Finally, the Y-specific centromeric repeat had a 4-fold higher rate of change than the homologous centromeric repeat on other chromosomes. Structural differences in satellite complexity, or restriction to the Y chromosome and elevated mutation rates of the male germline, may explain the higher rate of change. Overall, our work underscores the mutational fluidity of long tandem arrays of repeats, and the correlations and constraints between genome-wide tandem repeats which suggest that turnover is not a completely neutral process.

Copy number evolution in simple and complex tandem repeats across the C57BL/6 and C57BL/10 inbred mouse lines

Simple sequence tandem repeats are among the most rapidly evolving compartments of the genome. Some repeat expansions are associated with mammalian disease or meiotic segregation distortion, yet the rates of copy number change across generations are not well known. Here, we use 14 distinct sub-lineages of the C57BL/6 and C57BL/10 inbred mouse strains, which have been evolving independently over about 300 generations, to estimate the rates of copy number changes in genome-wide tandem repeats. Rates of change varied across simple repeats and across lines. Notably, CAG, whose expansions in coding regions are associated with many neurological and other genetic disorders, was highly stable in copy number, likely indicating purifying selection. Rates of change were generally positively correlated with copy number, but the direction and magnitude of changes varied across lines. Some mouse lines experienced consistent losses or gains across most genome-wide simple repeats, but this did not correlate with copy number changes in complex repeats. Rates of copy number change were similar between simple repeats and the much more abundant complex repeats once they were normalized by copy number. Finally, the Y-specific centromeric repeat had a 4-fold higher rate of change than the homologous centromeric repeat on other chromosomes. Structural differences in satellite complexity, or restriction to the Y chromosome and the elevated mutation rate of the male germline, may explain the higher rate of change. Overall, our work underscores the mutational fluidity of long tandem arrays of repeats, and the correlations and constraints between genome-wide tandem repeats which suggest that turnover is not a completely neutral process.

Identification of the Novel Chimeric Gene, PVT1-WWOX, in Multiple Myeloma with 8q24 Abnormality

Abstract 1823 Genetic abnormalities play a crucial role in the pathogenesis of various malignancies, including multiple myeloma (MM). Secondary cytogenetic abnormalities implicated in MM progression include 8q24 rearrangements, gain of the long arm of chromosome 1 (1q+), and loss of the short arm of chromosome 17 (17p-). The 8q24 rearrangements, including MYC and PVT1, have been identified by conventional cytogenetic analysis in 3.5–5.0% of MM patients and by fluorescence in situ hybridization (FISH) and spectral karyotyping (SKY) in 9.5–20%. 8q24 rearrangements are frequently associated with advanced disease in MM patients and MM cell lines. Ig chromosomal translocations, such as t(8;14)(q24;q32) and t(8;22)(q24;q11), occur in approximately 25% of MMs with 8q24 rearrangements, while non-Ig chromosomal loci, including 1p13, 1p21–22, 6p21, 6q12–15, 13q14 and 16q22, in which no candidate genes have been delineated so far, have also been identified as translocation partners. MYC has long been a possible candidate target gene for 8q24 rearrangements; however, many of the breakpoints within 8q24 have been assigned to various regions that encompassed more than 2 Mb centromeric or telomeric to MYC. We have previously found frequent PVT1 rearrangements in MM. PVT1 rearrangements were detected in 7 of 12 patients (58.3%) and in 5 of 8 cell lines (62.5%) with 8q24 abnormalities. A combination of SKY, FISH, and oligonucleotide array identified several partner loci of PVT1 rearrangements, such as 4p16, 4q13, 13q13, 14q32 and 16q23–24, and identified a chimeric gene, PVT1 - NBEA, resulting in high expression of abnormal NBEA in a cell line with t(8;13)(q24;q13), AMU-MM1, suggesting PVT1 rearrangements play significant roles in myelomagenesis. In this study, we analyzed RPMI8226 cell line in detail to identify other partner genes of PVT1 in these partner loci. SKY analysis revealed a complex karyotype including der(16)t(16;22)ins(16;8)(q23;q24) in this cell line. Oligonucleotide array analysis clearly demonstrated that the copy number change at 8q24 occurred within intron 1 of PVT1, and at 16q23, the copy number change occurred within intron 8 of WWOX, indicating that the translocation breakpoints of 8q24 and 16q23 were within intron 1 of PVT1 and intron 8 of WWOX, respectively. Based on these results, RT-PCR analysis was performed to detect chimeric products and direct sequencing of this product revealed the fusion of 5'-PVT1 exon 1 with WWOX exon 9-3'. The expression level of WWOX exon 9 was higher than WWOX exon 8–9 detected by semi-quantitative RT-PCR in RPMI8226, suggesting that high expression of WWOX derived from PVT1 - WWOX chimeric transcript, like PVT1 - NBEA. WWOX is generally considered to be a candidate tumor suppressor gene, and known to have a proapoptotic effect by participating in the tumor necrosis factor (TNF) apoptotic pathway and via direct physical interaction with p53 and its homolog p73. However, immunohistochemistry revealed that WWOX protein level were rather elevated in gastric and breast carcinoma. Therefore, WWOX seemed not to act as tumor suppressor gene simply. Although both NBEA and WWOX are located at common fragile site, usually contributing to gene inactivation, FRA13A and FRA16D respectively, these genes highly express via fusion to PVT1. These findings indicate that PVT1 rearrangements play significant roles in myelomagenesis via translocation and fusion to cancer-related genes. Disclosures: No relevant conflicts of interest to declare.