Project description:The objective of the study is to caracherize the genes that are regulated by Srf in myoblasts at day 0 (J0) and in differentiated cells at day 1 (J1) and day 3 (J3). We used microarrays to investigate gene expression in Srf KO and Srf WT muscle cells at day 0 (J0), day 1 (J1) and day 3 (J3) of differentiation
Project description:Srf is a MADS-box transcription factor that is critical for muscle differentiation. Its function in hematopoiesis has not yet been revealed. Mkl1, a cofactor of Srf, is part of the t(1;22) translocation in acute megakaryoblastic leukemia, and plays a critical role in megakaryopoiesis. In order to test the role of Srf in megakaryocyte development, we crossed Pf4-Cre mice, which express Cre recombinase in cells committed to the megakaryocytic lineage, to SrfF/F mice in which functional Srf is no longer expressed after Cre-mediated excision. Pf4-Cre/SrfF/F (KO) mice are born with normal mendelian frequency, but have significant macrothrombocytopenia with approximately 50% reduction in platelet count. In contrast, the BM has increased numbers and percentages of CD41+ megakaryocytes (WT: 0.41+/-0.06%; KO: 1.92+/-0.12%) with significantly reduced ploidy. KO mice show significantly increased megakaryocyte progenitors in the BM by both FACS analysis and CFU-Mk. Megakaryocytes lacking Srf have abnormal stress fiber and demarcation membrane formation and platelets lacking Srf have abnormal actin distribution. In vitro and in vivo assays reveal platelet function defects in KO mice. Critical actin cytoskeletal genes are downregulated in KO megakaryocytes. Thus, Srf is required for normal megakaryocyte maturation and platelet production, due at least in part, to regulation of cytoskeletal genes. C-kit+CD41+ megakaryocyte progenitors from PF4-Cre/SRF C57BL/6 SRF WT (3) and C57BL/6 SRF KO (3) mice were sorted by flow cytometry and cultured for three days in thrombopoietin.
Project description:Srf is a MADS-box transcription factor that is critical for muscle differentiation. Its function in hematopoiesis has not yet been revealed. Mkl1, a cofactor of Srf, is part of the t(1;22) translocation in acute megakaryoblastic leukemia, and plays a critical role in megakaryopoiesis. In order to test the role of Srf in megakaryocyte development, we crossed Pf4-Cre mice, which express Cre recombinase in cells committed to the megakaryocytic lineage, to SrfF/F mice in which functional Srf is no longer expressed after Cre-mediated excision. Pf4-Cre/SrfF/F (KO) mice are born with normal mendelian frequency, but have significant macrothrombocytopenia with approximately 50% reduction in platelet count. In contrast, the BM has increased numbers and percentages of CD41+ megakaryocytes (WT: 0.41+/-0.06%; KO: 1.92+/-0.12%) with significantly reduced ploidy. KO mice show significantly increased megakaryocyte progenitors in the BM by both FACS analysis and CFU-Mk. Megakaryocytes lacking Srf have abnormal stress fiber and demarcation membrane formation and platelets lacking Srf have abnormal actin distribution. In vitro and in vivo assays reveal platelet function defects in KO mice. Critical actin cytoskeletal genes are downregulated in KO megakaryocytes. Thus, Srf is required for normal megakaryocyte maturation and platelet production, due at least in part, to regulation of cytoskeletal genes.
Project description:Analysis of hematopoietic LSK(Lin-Sca1+c-Kit+) cells lacking the Serum response factor (SRF) gene. Results provide insight into the role of SRF in regulating genetic programs important for hematopoietic stem cell development Comparison of the gene expression profile between murine Lin-Sca1+c-Kit+ cells from Mx-Cre C57BL/6 Srf WT (3) and C57BL/6 Srf KO (3). Cells were sorted by flow cytometry and RNA was harvested and hybridized to Affymetrix MOE430A.
Project description:Previous efforts to conditionally delete the Srf transcription factor in smooth muscle lineages were met with early demise of the mouse due to a gastrointestinal (GI) obstruction phenotype. We have developed a new, more VSMC selective Cre recombinase mouse (Itga8-CreERT2), that circumvents the GI phenotype thus affording the opportunity to evaluate, in an unambiguous manner, VSMC phenotypes where Srf is reduced. The mice live for at least 6 months post-tamoxifen and here we report the first RNA-seq experiments in mouse aortic SMC where Srf is reduced.
Project description:TMT analysis of proteomic changes in the gastrocnemius skeletal muscles of WT and Bmal1-KO mice, and Bmal1-KO mice rescued with AAV-mediated muscle-specific expression of Bmal1.
Project description:RNA expression in Srf wildtype and knockout primary bone marrow derived mature neutrophils was determined via RNA sequencing Mature Gr1 high, 7/4 high, SSC high neutrophils were sorted from Srf WT and KO mouse bone marrow and submitted for RNA sequencing
Project description:Unstimulated murine bone marrow derived macrophages cultured in L92 media from WT (4 biological replicates), Nrf2 KO (3 biological replicates) and Keap1 KD BMDM (3 biological replicates) were processed and analysed utilising DIA (label free) proteomic analysis. The Nrf2 KO mouse (DOI: 10.1006/bbrc.1997.6943 ) and Keap1 KD mouse ( DOI: 10.1128/MCB.01591-09) were previously published as noted.