Recurrent reciprocal RNA chimera involving YPEL5 and PPP1CB in chronic lymphocytic leukemia

Abstract

Chronic lymphocytic leukemia (CLL) is the most common form of leukemia in adults in the Western hemisphere. Tumor-specific chromosomal translocations, characteristic findings in several human malignancies that directly lead to malignant transformation, have not been identified in CLL. Using paired-end transcriptome sequencing, we identified recurrent and reciprocal RNA chimeras involving yippee like 5 (YPEL5) and serine/threonine-protein phosphatase PP1-beta-catalytic subunit (PPP1CB) in CLL. Two of seven index cases (28%) harbored the reciprocal RNA chimeras in our initial screening. Using quantitative real-time PCR (q real-time PCR), YPEL5/PPP1CB and PPP1CB/YPEL5 fusion transcripts were detected in 97 of 103 CLL samples (95%) but not in paired normal samples, benign lymphocytes, or various unrelated cancers. Whole-genome sequencing and Southern blotting demonstrated no evidence for a genomic fusion between YPEL5 and PPP1CB. YPEL5/PPP1CB chimera, when introduced into mammalian cells, expressed a truncated PPP1CB protein that demonstrated diminished phosphatase activity. PPP1CB silencing resulted in enhanced proliferation and colony formation of MEC1 and JVM3 cells, implying a role in the pathogenesis of mature B-cell leukemia. These studies uncover a potential role for recurrent RNA chimeras involving phosphatases in the pathogenesis of a common form of leukemia. chimera splicing | next-generation sequencing | serine/threonine phosphatase PP1-beta catalytic subunit | B-lymphoid malignancy B -cell chronic lymphocytic leukemia (B-CLL) is the most common form of leukemia in adults in Western countries (1). The most common recurrent cytogenetic abnormality in CLL is a deletion involving the 13q14.3 locus, which occurs in 50% of cases and targets miR-16-1, miR-15a, and DLEU2 (2-4). Twenty percent of CLLs exhibit trisomy 12 (5). Other recurrent abnormalities in CLL include del 11q22-23 (ATM) and 17p13 (targeting p53) (6, 7). Of clinical relevance, IgV mutational status and zeta-chain associated protein kinase-70 kD (ZAP-70) expression have been associated with distinct prognostic categories of B-CLL (8-10). Recently, mutations in NOTCH1 (12.2%), MYD88 (2.9%), and XPO1 (2.4%) have been identified using next-generation sequencing (11, 12). The NOTCH1 mutations occur more frequently in cases with unmutated variable regions of the Ig heavy chain genes, whereas the MYD88 mutations occur more frequently in mutated cases (11).Although the role of genomic events is well established in the pathogenesis of cancers, the contribution of posttranscriptional RNA processing, which plays a fundamental role in control of protein expression, is less well understood.Alternative splicing can affect the translation, localization, or degradation of mRNA (13) and frequently results in the production of multiple and functionally distinct protein isoforms (14). Importantly, alternative splicing and expression of abnormal splicing chimeras may contribute to cancer pathogenesis and are associated with prognostic significance (15, 16). For exam...

Transcriptome characterization by RNA sequencing identifies a major molecular and clinical subdivision in chronic lymphocytic leukemia

“…We also analyzed other cancer studies where chimeric genes have been reported. In our full set of chimeric junctions (see Methods section ''Chimeric junctions'') we found five chimeric read-through events previously identified, LAMTOR1-ANAPC15 and PTPRG-C3orf14 in melanoma (Berger et al 2010), TATDN1-MTSS1 and MAP2K7-SNAPC2 in CLL (Velusamy et al 2013), and TTTY15-USP9Y found in prostate cancer and experimentally validated (Ren et al 2012). Only the latter showed CLL specificity (see Fig.…”

“…S10a,b). Recently, Velusamy and colleagues have reported a recurrent and reciprocal chimera between YPEL5 and PPP1CB genes (Velusamy et al 2013). This recurrence was reported to be as high as 95% of the CLL samples but negative in normal samples using qPCR.…”

Oncogenic Y641 mutations in EZH2 prevent Jak2/β-TrCP-mediated degradation

“…All assays were performed in triplicate and repeated two times and results were plotted as average fold change relative to GAPDH. 39 Primers used for detecting b-TrCP, p16 Ink4a or GAPDH are as follows: b-TrCP F (5 0 -TCATAACTGCTCTGCCAGCTCGGG-3 0 ), b-TrCP R (5 0 -ATCCTCTTC GTTCTGCCAGGCC-3 0 ), CDKN2A F (5 0 -GGGCTCTCACAACTAGGAAAG-3 0 ), CDKN2A R (5 0 -GGTGTTTGGTGTCATAGGG-3 0 ), CDKN1A F (5 0 -AGGTGGACC TGGAGACTCTCAG-3 0 ), CDKN1A R (5 0 -TCCTCTTGGAGAAGATCAGCCG-3 0 ), GAPDH F (5 0 -TGCACCACCAACTGCTTAGC-3 0 ) and GAPDH R (5 0 -GGCATGG ACTGTGGTCATGAG-3 0 ).…”

A Chimeric RNA Characteristic of Rhabdomyosarcoma in Normal Myogenesis Process

“…Besides JAZF1-JJAZ1 , reports for other chimeric RNAs generated by non-chromosomal rearrangement mechanisms are also emerging. Recently, reciprocal RNA chimera involving YPEL5 and PPP1CB was identifi ed by transcriptome sequencing, and no corresponding rearrangement at the DNA level was found ( 4 ). In addition, we reported cis -splicing of adjacent genes as another mechanism to generate SLC45A3-ELK4 chimera involving neighboring genes transcribing in the same direction ( 5 ).…”