Since MIR181A1 expression was suppressed in ETV6/RUNX1-positive ALL cells,
which were stalled at the pre-B stage and could be overcome by recovery of miR-181a
expression, we propose that miR-181a may have a role in promoting pre-B cell
differentiation. However, it needs more efforts to elucidate the function of miR-181a in
normal B cell development. In vivo strategies such as using B lineage-specific knockout
mice, or a xenograft mouse model using human HSCs with induction of miR-181a
expression at specific time point will be required.
In addition, the mature products miR-181a, miR-181b, miR-181c or miR-181d are
thought to have regulatory roles at post-transcriptional level through complementarity to
target mRNAs. While these mature miRNAs have similar predicted target mRNA and
most of them are downregulated in ETV6/RUNX1 ALL, whether the members of
miR-181 family other than miR-181a also involved in such a double negative loop
would need further investigation.
In conclusion, our study enhances our understanding in the interaction of
ETV6/RUNX1 and miRNAs, increases the knowledge of MIR181A1 function in human
B-cell development, helps to unravel one of the molecular mechanism underlying
ETV6/RUNX1-mediated attenuation of B-cell differentiation, and offers the opportunity
56
to identify new targets for development of therapeutic approaches to
ETV6/RUNX1-positive leukemia.
57
Bibliography
1. Pui CH, Carroll WL, Meshinchi S, Arceci RJ. Biology, risk stratification, and therapy of pediatric acute leukemias: an update. Journal of clinical oncology : official journal of the American Society of Clinical Oncology 2011 Feb 10; 29(5):
551-565.
2. Mullighan CG. Molecular genetics of B-precursor acute lymphoblastic leukemia.
The Journal of clinical investigation 2012 Oct; 122(10): 3407-3415.
3. Lo Nigro L. Biology of childhood acute lymphoblastic leukemia. Journal of pediatric hematology/oncology 2013 May; 35(4): 245-252.
4. Pui CH, Robison LL, Look AT. Acute lymphoblastic leukaemia. Lancet 2008 Mar 22; 371(9617): 1030-1043.
5. Ghazavi F, Lammens T, Van Roy N, Poppe B, Speleman F, Benoit Y, et al.
Molecular basis and clinical significance of genetic aberrations in B-cell precursor acute lymphoblastic leukemia. Experimental hematology 2015 Aug;
43(8): 640-653.
6. Huettner CS, Zhang P, Van Etten RA, Tenen DG. Reversibility of acute B-cell leukaemia induced by BCR-ABL1. Nature genetics 2000 Jan; 24(1): 57-60.
7. van der Weyden L, Giotopoulos G, Rust AG, Matheson LS, van Delft FW, Kong J, et al. Modeling the evolution of ETV6-RUNX1-induced B-cell precursor acute lymphoblastic leukemia in mice. Blood 2011 Jul 28; 118(4):
1041-1051.
8. Roberts KG, Mullighan CG. Genomics in acute lymphoblastic leukaemia:
insights and treatment implications. Nature reviews Clinical oncology 2015 Jun;
12(6): 344-357.
9. Mullighan CG, Su X, Zhang J, Radtke I, Phillips LA, Miller CB, et al. Deletion of IKZF1 and prognosis in acute lymphoblastic leukemia. The New England journal of medicine 2009 Jan 29; 360(5): 470-480.
58
10. Den Boer ML, van Slegtenhorst M, De Menezes RX, Cheok MH, Buijs-Gladdines JG, Peters ST, et al. A subtype of childhood acute lymphoblastic leukaemia with poor treatment outcome: a genome-wide classification study. The Lancet Oncology 2009 Feb; 10(2): 125-134.
11. Moorman AV, Richards SM, Robinson HM, Strefford JC, Gibson BE, Kinsey SE, et al. Prognosis of children with acute lymphoblastic leukemia (ALL) and intrachromosomal amplification of chromosome 21 (iAMP21). Blood 2007 Mar 15; 109(6): 2327-2330.
12. Harrison CJ, Moorman AV, Schwab C, Carroll AJ, Raetz EA, Devidas M, et al.
An international study of intrachromosomal amplification of chromosome 21 (iAMP21): cytogenetic characterization and outcome. Leukemia : official journal of the Leukemia Society of America, Leukemia Research Fund, UK 2014 May; 28(5): 1015-1021.
13. Mullighan CG, Collins-Underwood JR, Phillips LA, Loudin MG, Liu W, Zhang J, et al. Rearrangement of CRLF2 in B-progenitor- and Down syndrome-associated acute lymphoblastic leukemia. Nature genetics 2009 Nov;
41(11): 1243-1246.
14. Clappier E, Auclerc MF, Rapion J, Bakkus M, Caye A, Khemiri A, et al. An intragenic ERG deletion is a marker of an oncogenic subtype of B-cell precursor acute lymphoblastic leukemia with a favorable outcome despite frequent IKZF1 deletions. Leukemia : official journal of the Leukemia Society of America, Leukemia Research Fund, UK 2014 Jan; 28(1): 70-77.
15. Mullighan CG, Miller CB, Radtke I, Phillips LA, Dalton J, Ma J, et al.
BCR-ABL1 lymphoblastic leukaemia is characterized by the deletion of Ikaros.
Nature 2008 May 1; 453(7191): 110-114.
16. Golub TR, Barker GF, Bohlander SK, Hiebert SW, Ward DC, Bray-Ward P, et al. Fusion of the TEL gene on 12p13 to the AML1 gene on 21q22 in acute lymphoblastic leukemia. Proceedings of the National Academy of Sciences of the United States of America 1995 May 23; 92(11): 4917-4921.
17. Romana SP, Mauchauffe M, Le Coniat M, Chumakov I, Le Paslier D, Berger R, et al. The t(12;21) of acute lymphoblastic leukemia results in a tel-AML1 gene
59
fusion. Blood 1995 Jun 15; 85(12): 3662-3670.
18. Zelent A, Greaves M, Enver T. Role of the TEL-AML1 fusion gene in the molecular pathogenesis of childhood acute lymphoblastic leukaemia. Oncogene 2004 May 24; 23(24): 4275-4283.
19. Shurtleff SA, Buijs A, Behm FG, Rubnitz JE, Raimondi SC, Hancock ML, et al.
TEL/AML1 fusion resulting from a cryptic t(12;21) is the most common genetic lesion in pediatric ALL and defines a subgroup of patients with an excellent prognosis. Leukemia : official journal of the Leukemia Society of America, Leukemia Research Fund, UK 1995 Dec; 9(12): 1985-1989.
20. Romana SP, Poirel H, Leconiat M, Flexor MA, Mauchauffe M, Jonveaux P, et al. High frequency of t(12;21) in childhood B-lineage acute lymphoblastic leukemia. Blood 1995 Dec 1; 86(11): 4263-4269.
21. Yang YL, Lin SR, Chen JS, Hsiao CC, Lin KH, Sheen JM, et al. Multiplex reverse transcription-polymerase chain reaction as diagnostic molecular screening of 4 common fusion chimeric genes in Taiwanese children with acute lymphoblastic leukemia. Journal of pediatric hematology/oncology 2010 Nov;
32(8): e323-330.
22. Gabert J, Beillard E, van der Velden VHJ, Bi W, Grimwade D, Pallisgaard N, et al. Standardization and quality control studies of 'real-time' quantitative reverse transcriptase polymerase chain reaction of fusion gene transcripts for residual disease detection in leukemia - A Europe Against Cancer Program. Leukemia : official journal of the Leukemia Society of America, Leukemia Research Fund, UK 2003 Dec; 17(12): 2318-2357.
23. Baens M, Peeters P, Guo C, Aerssens J, Marynen P. Genomic organization of TEL: the human ETS-variant gene 6. Genome research 1996 May; 6(5):
404-413.
24. Levanon D, Glusman G, Bangsow T, Ben-Asher E, Male DA, Avidan N, et al.
Architecture and anatomy of the genomic locus encoding the human leukemia-associated transcription factor RUNX1/AML1. Gene 2001 Jan 10;
262(1-2): 23-33.
60
25. Mukouyama Y, Chiba N, Hara T, Okada H, Ito Y, Kanamaru R, et al. The AML1 transcription factor functions to develop and maintain hematogenic precursor cells in the embryonic aorta-gonad-mesonephros region.
Developmental biology 2000 Apr 1; 220(1): 27-36.
26. Yokomizo T, Ogawa M, Osato M, Kanno T, Yoshida H, Fujimoto T, et al.
Requirement of Runx1/AML1/PEBP2alphaB for the generation of haematopoietic cells from endothelial cells. Genes to cells : devoted to molecular & cellular mechanisms 2001 Jan; 6(1): 13-23.
27. Wang LC, Swat W, Fujiwara Y, Davidson L, Visvader J, Kuo F, et al. The TEL/ETV6 gene is required specifically for hematopoiesis in the bone marrow.
Genes & development 1998 Aug 1; 12(15): 2392-2402.
28. Look AT. Oncogenic transcription factors in the human acute leukemias.
Science 1997 Nov 7; 278(5340): 1059-1064.
29. Bohlander SK. ETV6: a versatile player in leukemogenesis. Seminars in cancer biology 2005 Jun; 15(3): 162-174.
30. van Dongen JJ, Macintyre EA, Gabert JA, Delabesse E, Rossi V, Saglio G, et al.
Standardized RT-PCR analysis of fusion gene transcripts from chromosome aberrations in acute leukemia for detection of minimal residual disease. Report of the BIOMED-1 Concerted Action: investigation of minimal residual disease in acute leukemia. Leukemia : official journal of the Leukemia Society of America, Leukemia Research Fund, UK 1999 Dec; 13(12): 1901-1928.
31. Morrow M, Samanta A, Kioussis D, Brady HJ, Williams O. TEL-AML1 preleukemic activity requires the DNA binding domain of AML1 and the dimerization and corepressor binding domains of TEL. Oncogene 2007 Jun 28;
26(30): 4404-4414.
32. Gunji H, Waga K, Nakamura F, Maki K, Sasaki K, Nakamura Y, et al.
TEL/AML1 shows dominant-negative effects over TEL as well as AML1.
Biochemical and biophysical research communications 2004 Sep 17; 322(2):
623-630.
33. Guidez F, Petrie K, Ford AM, Lu H, Bennett CA, MacGregor A, et al.
61
Recruitment of the nuclear receptor corepressor N-CoR by the TEL moiety of the childhood leukemia-associated TEL-AML1 oncoprotein. Blood 2000 Oct 1;
96(7): 2557-2561.
34. Wang L, Hiebert SW. TEL contacts multiple co-repressors and specifically associates with histone deacetylase-3. Oncogene 2001 Jun 21; 20(28):
3716-3725.
35. Greaves MF, Maia AT, Wiemels JL, Ford AM. Leukemia in twins: lessons in natural history. Blood 2003 Oct 1; 102(7): 2321-2333.
36. Mori H, Colman SM, Xiao Z, Ford AM, Healy LE, Donaldson C, et al.
Chromosome translocations and covert leukemic clones are generated during normal fetal development. Proceedings of the National Academy of Sciences of the United States of America 2002 Jun 11; 99(12): 8242-8247.
37. Tsuzuki S, Seto M, Greaves M, Enver T. Modeling first-hit functions of the t(12;21) TEL-AML1 translocation in mice. Proceedings of the National Academy of Sciences of the United States of America 2004 Jun 1; 101(22):
8443-8448.
38. Fischer M, Schwieger M, Horn S, Niebuhr B, Ford A, Roscher S, et al. Defining the oncogenic function of the TEL/AML1 (ETV6/RUNX1) fusion protein in a mouse model. Oncogene 2005 Nov 17; 24(51): 7579-7591.
39. Lee RC, Feinbaum RL, Ambros V. The C. elegans heterochronic gene lin-4 encodes small RNAs with antisense complementarity to lin-14. Cell 1993 Dec 3;
75(5): 843-854.
40. Winter J, Jung S, Keller S, Gregory RI, Diederichs S. Many roads to maturity:
microRNA biogenesis pathways and their regulation. Nature cell biology 2009 Mar; 11(3): 228-234.
41. Bartel DP. MicroRNAs: genomics, biogenesis, mechanism, and function. Cell 2004 Jan 23; 116(2): 281-297.
42. Shivdasani RA. MicroRNAs: regulators of gene expression and cell differentiation. Blood 2006 Dec 1; 108(12): 3646-3653.
62
43. Hausser J, Syed AP, Bilen B, Zavolan M. Analysis of CDS-located miRNA target sites suggests that they can effectively inhibit translation. Genome research 2013 Apr; 23(4): 604-615.
44. Brummer A, Hausser J. MicroRNA binding sites in the coding region of mRNAs: extending the repertoire of post-transcriptional gene regulation.
BioEssays : news and reviews in molecular, cellular and developmental biology 2014 Jun; 36(6): 617-626.
45. Vasilatou D, Papageorgiou S, Pappa V, Papageorgiou E, Dervenoulas J. The role of microRNAs in normal and malignant hematopoiesis. European journal of haematology 2010 Jan 1; 84(1): 1-16.
46. Felli N, Fontana L, Pelosi E, Botta R, Bonci D, Facchiano F, et al. MicroRNAs 221 and 222 inhibit normal erythropoiesis and erythroleukemic cell growth via kit receptor down-modulation. Proceedings of the National Academy of Sciences of the United States of America 2005 Dec 13; 102(50): 18081-18086.
47. Gilicze AB, Wiener Z, Toth S, Buzas E, Pallinger E, Falcone FH, et al.
Myeloid-derived microRNAs, miR-223, miR27a, and miR-652, are dominant players in myeloid regulation. BioMed research international 2014; 2014:
870267.
48. Neilson JR, Zheng GX, Burge CB, Sharp PA. Dynamic regulation of miRNA expression in ordered stages of cellular development. Genes & development 2007 Mar 1; 21(5): 578-589.
49. Xiao C, Calado DP, Galler G, Thai TH, Patterson HC, Wang J, et al. MiR-150 controls B cell differentiation by targeting the transcription factor c-Myb. Cell 2007 Oct 5; 131(1): 146-159.
50. Zhou B, Wang S, Mayr C, Bartel DP, Lodish HF. miR-150, a microRNA expressed in mature B and T cells, blocks early B cell development when expressed prematurely. Proceedings of the National Academy of Sciences of the United States of America 2007 Apr 24; 104(17): 7080-7085.
51. Fazi F, Racanicchi S, Zardo G, Starnes LM, Mancini M, Travaglini L, et al.
63
Epigenetic silencing of the myelopoiesis regulator microRNA-223 by the AML1/ETO oncoprotein. Cancer cell 2007 Nov; 12(5): 457-466.
52. O'Connell RM, Rao DS, Chaudhuri AA, Baltimore D. Physiological and pathological roles for microRNAs in the immune system. Nature reviews Immunology 2010 Feb; 10(2): 111-122.
53. Shi L, Cheng Z, Zhang J, Li R, Zhao P, Fu Z, et al. hsa-mir-181a and hsa-mir-181b function as tumor suppressors in human glioma cells. Brain research 2008 Oct 21; 1236: 185-193.
54. Ouyang YB, Lu Y, Yue S, Giffard RG. miR-181 targets multiple Bcl-2 family members and influences apoptosis and mitochondrial function in astrocytes.
Mitochondrion 2012 Mar; 12(2): 213-219.
55. Gao W, Yu Y, Cao H, Shen H, Li X, Pan S, et al. Deregulated expression of miR-21, miR-143 and miR-181a in non small cell lung cancer is related to clinicopathologic characteristics or patient prognosis. Biomedicine &
pharmacotherapy = Biomedecine & pharmacotherapie 2010 Jul; 64(6):
399-408.
56. Visone R, Rassenti LZ, Veronese A, Taccioli C, Costinean S, Aguda BD, et al.
Karyotype-specific microRNA signature in chronic lymphocytic leukemia.
Blood 2009 Oct 29; 114(18): 3872-3879.
57. Pallasch CP, Patz M, Park YJ, Hagist S, Eggle D, Claus R, et al. miRNA deregulation by epigenetic silencing disrupts suppression of the oncogene PLAG1 in chronic lymphocytic leukemia. Blood 2009 Oct 8; 114(15):
3255-3264.
58. Bai H, Cao Z, Deng C, Zhou L, Wang C. miR-181a sensitizes resistant leukaemia HL-60/Ara-C cells to Ara-C by inducing apoptosis. Journal of cancer research and clinical oncology 2012 Apr; 138(4): 595-602.
59. Chen G, Zhu W, Shi D, Lv L, Zhang C, Liu P, et al. MicroRNA-181a sensitizes human malignant glioma U87MG cells to radiation by targeting Bcl-2. Oncology reports 2010 Apr; 23(4): 997-1003.
64
60. Gefen N, Binder V, Zaliova M, Linka Y, Morrow M, Novosel A, et al.
Hsa-mir-125b-2 is highly expressed in childhood ETV6/RUNX1 (TEL/AML1) leukemias and confers survival advantage to growth inhibitory signals independent of p53. Leukemia : official journal of the Leukemia Society of America, Leukemia Research Fund, UK 2010 Jan; 24(1): 89-96.
61. Diakos C, Zhong S, Xiao Y, Zhou M, Vasconcelos GM, Krapf G, et al.
TEL-AML1 regulation of survivin and apoptosis via miRNA-494 and miRNA-320a. Blood 2010 Dec 2; 116(23): 4885-4893.
62. Torrano V, Procter J, Cardus P, Greaves M, Ford AM. ETV6-RUNX1 promotes survival of early B lineage progenitor cells via a dysregulated erythropoietin receptor. Blood 2011 Nov 3; 118(18): 4910-4918.
63. Kaindl U, Morak M, Portsmouth C, Mecklenbrauker A, Kauer M, Zeginigg M, et al. Blocking ETV6/RUNX1-induced MDM2 overexpression by Nutlin-3 reactivates p53 signaling in childhood leukemia. Leukemia : official journal of the Leukemia Society of America, Leukemia Research Fund, UK 2014 Mar;
28(3): 600-608.
64. Wang X, Xuan Z, Zhao X, Li Y, Zhang MQ. High-resolution human core-promoter prediction with CoreBoost_HM. Genome research 2009 Feb;
19(2): 266-275.
65. Zaliova M, Madzo J, Cario G, Trka J. Revealing the role of TEL/AML1 for leukemic cell survival by RNAi-mediated silencing. Leukemia : official journal of the Leukemia Society of America, Leukemia Research Fund, UK 2011 Feb;
25(2): 313-320.
66. Diakos C, Krapf G, Gerner C, Inthal A, Lemberger C, Ban J, et al.
RNAi-mediated silencing of TEL/AML1 reveals a heat-shock protein- and survivin-dependent mechanism for survival. Blood 2007 Mar 15; 109(6):
2607-2610.
67. Benjamini YaH, Yosef. Controlling the false discovery rate: a practical and powerful approach to multiple testing. Journal of the Royal Statistical Society, Series B 1995; 57(1): 289-300.
65
68. Karagianni P, Wong J. HDAC3: taking the SMRT-N-CoRrect road to repression.
Oncogene 2007 Aug 13; 26(37): 5439-5449.
69. Moreno DA, Scrideli CA, Cortez MA, de Paula Queiroz R, Valera ET, da Silva Silveira V, et al. Differential expression of HDAC3, HDAC7 and HDAC9 is associated with prognosis and survival in childhood acute lymphoblastic leukaemia. British journal of haematology 2010 Sep; 150(6): 665-673.
70. Kramer OH, Zhu P, Ostendorff HP, Golebiewski M, Tiefenbach J, Peters MA, et al. The histone deacetylase inhibitor valproic acid selectively induces proteasomal degradation of HDAC2. The EMBO journal 2003 Jul 1; 22(13):
3411-3420.
71. Liu S, Klisovic RB, Vukosavljevic T, Yu J, Paschka P, Huynh L, et al.
Targeting AML1/ETO-histone deacetylase repressor complex: a novel mechanism for valproic acid-mediated gene expression and cellular differentiation in AML1/ETO-positive acute myeloid leukemia cells. The Journal of pharmacology and experimental therapeutics 2007 Jun; 321(3):
953-960.
72. Li X, Zhang J, Gao L, McClellan S, Finan MA, Butler TW, et al. MiR-181 mediates cell differentiation by interrupting the Lin28 and let-7 feedback circuit.
Cell death and differentiation 2012 Mar; 19(3): 378-386.
73. Van Dyck F, Declercq J, Braem CV, Van de Ven WJ. PLAG1, the prototype of the PLAG gene family: versatility in tumour development (review).
International journal of oncology 2007 Apr; 30(4): 765-774.
74. Mesquita Junior D, Araujo JA, Catelan TT, Souza AW, Cruvinel Wde M, Andrade LE, et al. Immune system - part II: basis of the immunological response mediated by T and B lymphocytes. Revista brasileira de reumatologia 2010 Sep-Oct; 50(5): 552-580.
75. Perez-Andres M, Paiva B, Nieto WG, Caraux A, Schmitz A, Almeida J, et al.
Human peripheral blood B-cell compartments: a crossroad in B-cell traffic.
Cytometry Part B, Clinical cytometry 2010; 78 Suppl 1: S47-60.
76. Fazi F, Nervi C. MicroRNA: basic mechanisms and transcriptional regulatory
66
networks for cell fate determination. Cardiovascular research 2008 Sep 1; 79(4):
553-561.
77. Schotte D, De Menezes RX, Akbari Moqadam F, Khankahdani LM, Lange-Turenhout E, Chen C, et al. MicroRNA characterize genetic diversity and drug resistance in pediatric acute lymphoblastic leukemia. Haematologica 2011 May; 96(5): 703-711.
78. Patz M, Pallasch CP, Wendtner CM. Critical role of microRNAs in chronic lymphocytic leukemia: overexpression of the oncogene PLAG1 by deregulated miRNAs. Leukemia & lymphoma 2010 Aug; 51(8): 1379-1381.
79. Fazi F, Rosa A, Fatica A, Gelmetti V, De Marchis ML, Nervi C, et al. A minicircuitry comprised of microRNA-223 and transcription factors NFI-A and C/EBPalpha regulates human granulopoiesis. Cell 2005 Dec 2; 123(5): 819-831.
80. Lwin T, Zhao X, Cheng F, Zhang X, Huang A, Shah B, et al. A microenvironment-mediated c-Myc/miR-548m/HDAC6 amplification loop in non-Hodgkin B cell lymphomas. The Journal of clinical investigation 2013 Nov;
123(11): 4612-4626.
81. Starkova J, Madzo J, Cario G, Kalina T, Ford A, Zaliova M, et al. The identification of (ETV6)/RUNX1-regulated genes in lymphopoiesis using histone deacetylase inhibitors in ETV6/RUNX1-positive lymphoid leukemic cells. Clinical cancer research : an official journal of the American Association for Cancer Research 2007 Mar 15; 13(6): 1726-1735.
82. Wang X, Gocek E, Liu CG, Studzinski GP. MicroRNAs181 regulate the expression of p27Kip1 in human myeloid leukemia cells induced to differentiate by 1,25-dihydroxyvitamin D3. Cell Cycle 2009 Mar 1; 8(5): 736-741.
83. Georgantas RW, 3rd, Hildreth R, Morisot S, Alder J, Liu CG, Heimfeld S, et al.
CD34+ hematopoietic stem-progenitor cell microRNA expression and function:
a circuit diagram of differentiation control. Proceedings of the National Academy of Sciences of the United States of America 2007 Feb 20; 104(8):
2750-2755.
84. Chen CZ, Li L, Lodish HF, Bartel DP. MicroRNAs modulate hematopoietic
67
lineage differentiation. Science 2004 Jan 2; 303(5654): 83-86.
85. Spierings DC, McGoldrick D, Hamilton-Easton AM, Neale G, Murchison EP, Hannon GJ, et al. Ordered progression of stage-specific miRNA profiles in the mouse B2 B-cell lineage. Blood 2011 May 19; 117(20): 5340-5349.
86. Landgraf P, Rusu M, Sheridan R, Sewer A, Iovino N, Aravin A, et al. A mammalian microRNA expression atlas based on small RNA library sequencing.
Cell 2007 Jun 29; 129(7): 1401-1414.
68
Figures
69
A
B
Figure 1. hsa-mir-181a-1 and 5p sequence of miR-181 family.
(A) Stem-loop sequence of hsa-mir-181a-1. Human precursor mir-181a-1 is in length with 110 bp arisen from MIR181A1 primary transcript. Two mature forms miR-181a (red) and miR181a-1 (blue) derived from 5ʹ and 3ʹ arm of precursor, respectively, are indicated. (B) The whole sequence of 5p and the seed sequence of each member in human miR-181 family is represented as 5ʹ to 3ʹ end.
70
Figure 2. MicroRNA expression profile in childhood B-ALL patients.
50 childhood B-ALL patients were recruited including E/R-positive, n=10 (purple); E/R-negative, n=40 (gray). 365 miRNA were analyzed by ABI Taqman qRT-PCR based miRNA arrays which were carried out by Microarray core facility, Yu’s Lab, NTU.
71
A B
C
Figure 3. Validation of individual miRNA expression.
15 childhood B-ALL patients were included (E/R-positive, n = 7; E/R-negative, n
= 8). (A) miR-181a-1 (E/R-positive samples: 0.14 ± 0.08; E/R -positive samples:
0.06 ± 0.03) and (B) miR-181a (E/R -positive samples: 6.15 ± 5.49; E/R -positive samples: 5.13 ± 4.58) levels were measured by TaqMan microRNA assays. * P ≤ 0.05 (ANOVA) (C) Association between miR-181a-1 and miR-181a expression level in primary B-ALL cells. Pearson correlation coefficient = 0.67, P = 0.0045.
72
Figure 4. Expression of ETV6/RUNX1 fusion protein and wild type RUNX1 protein in B-ALL cell lines.
REH cells (left lane) are t(12;21)-positive which express both ETV6/RUNX1 fusion protein (predicted size: 100 kDa) and RUNX1 protein (predicted size:
55kDa), and CCRF-SB cells (right lane) do not carry any common translocation which only express RUNX1 protein. Protein expression was detected by Western blot using anti-RUNX1 antibody and β-actin was used as an internal control. E/R:
ETV6/RUNX1.