Enforced expression of miR-125b affects myelopoiesis by targeting multiple signaling pathways

• Published in: Blood 2011-04, Vol.117 (16), p.4338-4348
• Main Authors: Surdziel, Ewa, Cabanski, Maciej, Dallmann, Iris, Lyszkiewicz, Marcin, Krueger, Andreas, Ganser, Arnold, Scherr, Michaela, Eder, Matthias
• Format: Article
• Language: English
• Subjects: Animals; bcl-2 Homologous Antagonist-Killer Protein - genetics; bcl-2 Homologous Antagonist-Killer Protein - metabolism; Biological and medical sciences; Bone Marrow Cells - cytology; Bone Marrow Cells - metabolism; Cell Death; Cell Differentiation; Cell Line; Cell Proliferation; DNA - metabolism; Gene Expression; Granulocyte Colony-Stimulating Factor - metabolism; Granulocytes - cytology; Granulocytes - metabolism; Hematologic and hematopoietic diseases; Medical sciences; Mice; Mice, Inbred C57BL; MicroRNAs - genetics; Myeloid Cells - cytology; Myeloid Cells - metabolism; Signal Transduction; STAT3 Transcription Factor - genetics; STAT3 Transcription Factor - metabolism; Up-Regulation
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood-2010-06-289058

MicroRNAs (miRNAs) are small, noncoding RNAs that regulate gene expression by sequence-specific targeting of multiple mRNAs. Although lineage-, maturation-, and disease-specific miRNA expression has been described, miRNA-dependent phenotypes and miRNA-regulated signaling in hematopoietic cells are largely unknown. Combining functional genomics, biochemical analysis, and unbiased and hypothesis-driven miRNA target prediction, we show that lentivirally over-expressed miR-125b blocks G-CSF–induced granulocytic differentiation and enables G-CSF–dependent proliferation of murine 32D cells. In primary lineage-negative cells, miR-125b over-expression enhances colony-formation in vitro and promotes myelopoiesis in mouse bone marrow chimeras. We identified Stat3 and confirmed Bak1 as miR-125b target genes with approximately 30% and 50% reduction in protein expression, respectively. However, gene-specific RNAi reveals that this reduction, alone and in combination, is not sufficient to block G-CSF–dependent differentiation. STAT3 protein expression, DNA-binding, and transcriptional activity but not induction of tyrosine-phosphorylation and nuclear translocation are reduced upon enforced miR-125b expression, indicating miR-125b–mediated reduction of one or more STAT3 cofactors. Indeed, we identified c-Jun and Jund as potential miR-125b targets and demonstrated reduced protein expression in 32D/miR-125b cells. Interestingly, gene-specific silencing of JUND but not c-JUN partially mimics the miR-125b over-expression phenotype. These data demonstrate coordinated regulation of several signaling pathways by miR-125b linked to distinct phenotypes in myeloid cells.