t(15;21) translocations leading to the concurrent downregulation of RUNX1 and its transcription factor partner genes SIN3A and TCF12 in myeloid disorders

Abstract

Through a combined approach integrating RNA-Seq, SNP-array, FISH and PCR techniques, we identified two novel t(15;21) translocations leading to the inactivation of RUNX1 and its partners SIN3A and TCF12. One is a complex t(15;21)(q24;q22), with both breakpoints mapped at the nucleotide level, joining RUNX1 to SIN3A and UBL7-AS1 in a patient with myelodysplasia. The other is a recurrent t(15;21)(q21;q22), juxtaposing RUNX1 and TCF12, with an opposite transcriptional orientation, in three myeloid leukemia cases. Since our transcriptome analysis indicated a significant number of differentially expressed genes associated with both translocations, we speculate an important pathogenetic role for these alterations involving RUNX1.

Fig. 1

t(15;21)(q24;q22) translocation in case 1. a Genomic breakpoints on chromosomes 15 and 21 in case 1: Top, schematic representation of wild-type chromosomes (black lines) and involved genes (exons are shown as rectangles and introns as lines connecting exons, with arrowheads indicating the direction of transcription). The dashed black lines indicate the breakpoints within all genes. Bottom, partial chromatograms indicate fusion sequences on der(15) (left) [GenBank: KT336107] and der(21) (right) [GenBank: KT336106]. The splicing event creating the runt-related transcription factor 1 (RUNX1)/UBL7-AS1 fusion transcript is indicated in green. b RUNX1 fusion transcripts: reverse transcription polymerase chain reaction (RT-PCR) products corresponding to SIN3A/RUNX1 [GenBank: KT336104] and RUNX1/UBL7-AS1 [GenBank: KT336105] fusion transcripts (lanes 1 and 3, respectively) in case 1 are shown in the middle of the panel. Lanes 2 and 4: negative normal bone marrow samples. Lane 5: 2-log DNA ladder (New England Biolabs, Milan, Italy). Partial chromatograms (top) and structure (bottom) of SIN3A/RUNX1 and RUNX1/UBL7-AS1 PCR products are on the left and on the right, respectively. c, d RUNX1 chimeric proteins: both panels show in silico translation (ORFfinder and BlastP) of both wild-type and chimeric RUNX1 and SIN3A proteins. Arrows indicate the truncation breakpoints of wild-type proteins. e Evaluation of RUNX1 and SIN3A expression levels in case 1: exon-specific reverse transcription quantitative PCR analysis of RUNX1 (left) and SIN3A (right) was performed in case 1 vs a control pooled sample of patients with acute myeloid leukemia. Asterisks indicate statistically significant results (P < .05)

Fig. 2

t(15;21)(q21;q22) translocation in cases 2, 3, and 4. a, b Breakpoints on chromosomes 21 and 15: partial karyotypes showing fluorescence in situ hybridization (FISH) results that allowed the mapping of t(15;21) translocation breakpoints on der(21) (a) and der(15) (b), using the consistently colored probes listed for cases 2 (first column) and 4 (second column). RP11-662O10 was used only in case 4. Cases 2 and 3 shared the same breakpoints (data not shown). On the right, the map of the BAC probes used in the FISH experiments, according to GRCh37/hg19, and identifying both translocation and deletion breakpoints, is shown. The black and orange dashed lines indicate the breakpoints in cases 2 and 4, respectively. The grey rectangle encompasses the deleted region flanking the translocation breakpoint in case 4. c Evaluation of RUNX1 and TCF12 expression levels: exon-specific reverse transcription quantitative polymerase chain reaction results of runt-related transcription factor 1 (RUNX1; left) and transcription factor 12 (TCF12; right) in case 2. Asterisks indicate statistically significant results (P < .05)