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Blood, Vol. 113, Issue 11, 2375-2385, March 12, 2009 This Article Abstract Full Text Services Email this article to a friend Alert me to new issues of the journal Rights and Permissions Citing Articles Citing Articles via CrossRef HOXA9 is required for survival in human MLL-rearranged acute leukemias Blood Faber et al. 113: 2375 Supplemental materials for: Faber et al Files in this Data Supplement: Document 1. Supplemental methods and results (PDF, 83.7 KB) Figure S1. Determination of GFP control shRNA transduction efficiency in 17 different MLL-rearranged and MLL-wildtype leukemia cell lines by puromycin selection (JPG, 80 KB) - When viability of puromycin selected cells was compared to transduced cells without puromycin selection (3 days after puromycin selection / 4 days after transduction) a transduction efficiency > 80% was confirmed for each cell lines tested. Figure S2. Analysis of expression of the interferon response genes OAS1, IFITM1, and MX1 (JPG, 62.9 KB) - By quantitative real-time PCR 72h after transduction with the four HOXA9 shRNA constructs, the non-targeting GFP control shRNA or empty pLKO.1 vector. Compared to untreated MOLM-14 cells all lentiviral constructs used showed normal interferon response gene expression levels and no induction whereas pI:C transfected cells revealed significant induction confirming that the interferon response pathway is inducible in this cell line. Figure S3 (JPG, 52.2 KB) - Quantitative real-time PCR analysis of HOXA9 expression 24 hours after transduction of murine leukemia stem cells (L-GMP) with the 1F3-HOXA9shRNA or 1F2-HOXA9shRNA construct demonstrated efficient HOXA9 suppression as compared to GFP-shRNA transduced controls. Figure S4. Analysis of the proliferative capacity as assessed by MTT assay in MOLM-14 cells after transduction with the 1F3-HOXA9shRNA construct (JPG, 53.8 KB) - Inhibition of the proliferative capacity was observed starting 48h after transduction. Figure S5. HOXA9 knockdown by lipofection of a 21-mer siRNA oligomer in t(9;11) THP-1 cells (JPG, 54.2 KB) - (A) Efficient HOXA9 suppression (75–80%) similar to HOXA9shRNA mediated suppression levels was confirmed by quantitative real-time PCR 72h after transfection. (B) Time course analysis of viability after HOXA9siRNA transfection revealed a progressive induction of cell death with an extent and kinetic similar to the effects observed after shRNA mediated suppression. (C) Ectopic overexpression of a non-targetable HOXA9 construct significantly rescued the phenotype (Annexin V staining). This Article Abstract Full Text Services Email this article to a friend Alert me to new issues of the journal Rights and Permissions Citing Articles Citing Articles via CrossRef

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