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C. Zhang et al.
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Figure 1. (continued from the previous page) (E) mRNA expression of ALDH1A2 in cells from three different cohorts (TARGET, Singapore, and Children’s Oncology Group/Dana-Farber Cancer Institute [COG/DFCI] cohorts) of primary T- ALL samples analyzed by RNA-sequencing. Expression values in different sub- groups are shown as transcripts per million (TPM). Bars represent the median and SD. *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001 using the two-tailed Student t test. ns: not statistically significant. (F) The RNA-sequencing gene tracks represent mRNA expression of each exon of the ALDH1A2 gene in six TAL1-positive primary T-ALL cell samples (TARGET 1 and 2; COG/DFCI 1 and 2; Singapore 1 and 2), four TAL1-positive cell lines, four TAL1-negative T-ALL cell lines and a chronic myeloid leukemia cell line (K562). Red dashed lines repre- sent the short or long transcripts of ALDH1A2. (G) Western blot analysis showing the expression of the full-length and short isoforms of the ALDH1A2 protein in various cell lines. CML: chronic myeloid leukemia.
hematopoietic cells,26 ALDH1A2 expression was not detect- ed in T cells (Online Supplementary Figure S1E). Similarly, the mouse Aldh1a2 gene was not expressed in hematopoietic cells, while it was detected in bone marrow stroma cells (Online Supplementary Figure S1F). Furthermore, recent sin- gle-cell RNA-sequencing studies confirmed that ALDH1A2 is expressed in thymic stroma cells but not in any T-cell sub- populations both in mice and human cells.27,28 These results further indicated that ALDH1A2 is aberrantly activated in cases of TAL1-positive T-ALL.
The short isoform of ALDH1A2 possesses enzymatic activity and is localized in the cytoplasm
We next investigated the function of the short isoform of the ALDH1A2 protein. The analysis of the amino acid sequence indicated that the short isoform retains function-
Cancer Cell Line Encyclopedia dataset showed the highest expression of ALDH1A2 mRNA in T-ALL samples among 40 cancer types25 (Online Supplementary Figure S1D).
Notably, RNA-sequencing analysis demonstrated that only the short isoform, which lacks the first three exons, but not the full-length (long) isoform of ALDH1A2, was expressed in the TAL1-positive T-ALL cells, including pri- mary samples and cell lines (Figure 1F). This result was consistent with that from an earlier study.12 In contrast, K562 cells, a chronic myeloid leukemia cell line, expressed the long isoform of ALDH1A2 but not the short isoform (Figure 1F, bottom). The difference in protein size was confirmed by western blot (Figure 1G).
It is noteworthy that the H3K27ac marks were not observed in normal T cells (Figure 1A). Consistent with this finding, in an analysis of a public dataset of normal human
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haematologica | 2021; 106(6)