Figure 1
Figure 1. Expression of miR-29a is down-regulated in AML with −7/del7q and regulates expression of Ski. (A) Expression of miR-29a in AML patient samples with isolated −7/del7q karyotype or complex karyotype including −7/del7q. Expression levels of miR-29a in 12 AML samples with −7/del7q as determined by microarray analysis compared with miR-29a expression in 8 samples of normal bone marrow. Five of the original 17 values for miR-29a expression in AML samples are missing. (B) miR-29a inhibits Ski expression. HL60 cells were transfected with 29a or scrambled precursor miRNA. After 24 hours, Ski expression was analyzed in whole-cell protein extracts with Western blot. Loading control was β-actin. (C) miR-29a enhances ATRA-induced differentiation of myeloid HL60 cells. Cellular differentiation of HL60 cells transfected with 29a or scrambled control precursor miRNA additionally treated with 1.5μM ATRA for 72 hours was analyzed with flow cytometry. Black bar indicates miR-scrambled; dark gray bar, miR-scrambled plus ATRA; gray bar, miR-29a; white bar, miR-29a plus ATRA. CD11b expression of miR-scrambled transfected cells was set at 100%. Results are representative of 3 independent experiments. Values shown are mean ± SEM. (D) miR-29a reduces proliferation of myeloid HL60 cells. HL60 cells were transfected with miRNA precursor 29a or scrambled control. After 0, 24, 48, and 72 hours, proliferation was tested in an MTT assay. ■ indicates miR-29a; ▴, miR scrambled. Experiment was performed 3 times in triplicate. Values shown are mean ± SEM. (E) Expression of the Ski target gene Nr-CAM is inhibited by miR-29a. IFB melanoma cells were transfected with miR-29a or scrambled precursor miRNA. After 24 hours, Ski expression was tested in whole-cell protein lysates by Western blot (left). Loading control was β-actin. Nr-CAM expression was measured by PCR (right). Control was β-actin. (F) Inhibition of miR-29a induces Ski expression. NW1539 melanoma cells (left) or HL60 cells (right) were transfected with anti–miR-29a or scrambled oligonucleotide. After 48 hours, Western blot was performed for Ski with whole-cell protein extracts. Loading control was β-actin.

Expression of miR-29a is down-regulated in AML with −7/del7q and regulates expression of Ski. (A) Expression of miR-29a in AML patient samples with isolated −7/del7q karyotype or complex karyotype including −7/del7q. Expression levels of miR-29a in 12 AML samples with −7/del7q as determined by microarray analysis compared with miR-29a expression in 8 samples of normal bone marrow. Five of the original 17 values for miR-29a expression in AML samples are missing. (B) miR-29a inhibits Ski expression. HL60 cells were transfected with 29a or scrambled precursor miRNA. After 24 hours, Ski expression was analyzed in whole-cell protein extracts with Western blot. Loading control was β-actin. (C) miR-29a enhances ATRA-induced differentiation of myeloid HL60 cells. Cellular differentiation of HL60 cells transfected with 29a or scrambled control precursor miRNA additionally treated with 1.5μM ATRA for 72 hours was analyzed with flow cytometry. Black bar indicates miR-scrambled; dark gray bar, miR-scrambled plus ATRA; gray bar, miR-29a; white bar, miR-29a plus ATRA. CD11b expression of miR-scrambled transfected cells was set at 100%. Results are representative of 3 independent experiments. Values shown are mean ± SEM. (D) miR-29a reduces proliferation of myeloid HL60 cells. HL60 cells were transfected with miRNA precursor 29a or scrambled control. After 0, 24, 48, and 72 hours, proliferation was tested in an MTT assay. ■ indicates miR-29a; ▴, miR scrambled. Experiment was performed 3 times in triplicate. Values shown are mean ± SEM. (E) Expression of the Ski target gene Nr-CAM is inhibited by miR-29a. IFB melanoma cells were transfected with miR-29a or scrambled precursor miRNA. After 24 hours, Ski expression was tested in whole-cell protein lysates by Western blot (left). Loading control was β-actin. Nr-CAM expression was measured by PCR (right). Control was β-actin. (F) Inhibition of miR-29a induces Ski expression. NW1539 melanoma cells (left) or HL60 cells (right) were transfected with anti–miR-29a or scrambled oligonucleotide. After 48 hours, Western blot was performed for Ski with whole-cell protein extracts. Loading control was β-actin.

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