Similar Datasets
Project description:Serum response factor (SRF) is a ubiquitously expressed transcription factor that binds DNA at CArG (CC[A/T]6GG) domains in association with myocardin-family proteins (eg, myocardin-related transcription factor A [MRTFA]) or the ternary complex factor family of E26 transformation-specific (ETS) proteins. In primary hematopoietic cells, knockout of either SRF or MRTFA decreases megakaryocyte (Mk) maturation causing thrombocytopenia. The human erythroleukemia (HEL) cell line mimics the effects of MRTFA on Mk maturation, and MRTFA overexpression (MRTFAOE) in HEL cells enhances megakaryopoiesis. To identify the mechanisms underlying these effects, we performed integrated analyses of anti-SRF chromatin immunoprecipitation (ChIP) and RNA-sequencing data from noninduced and phorbol ester (12-O-tetradecanoylphorbol-13-acetate [TPA])-induced HEL cells, with and without MRTFAOE We found that 11% of genes were upregulated with TPA induction, which was enhanced by MRTFAOE, resulting in an upregulation of 25% of genes. MRTFAOE increased binding of SRF to genomic sites and enhanced TPA-induced expression of SRF target genes. The TPA-induced genes are predicted to be regulated by SRF and ETS factors, whereas those upregulated by TPA plus MRTFAOE lack ETS binding motifs, and MRTFAOE skews SRF binding to genomic regions with CArG sites in regions relatively lacking in ETS binding motifs. Finally, ChIP-polymerase chain reaction using HEL cells and primary human CD34+ cell-derived subpopulations confirms that both SRF and MRTFA have increased binding during megakaryopoiesis at upregulated target genes (eg, CORO1A). We show for the first time that MRTFA increases both the genomic association and activity of SRF and upregulates genes that enhance primary human megakaryopoiesis.
| S-EPMC6199649 | biostudies-literature
Project description:SRF is a ubiquitous transcription factor that binds DNA in association with myocardin-family proteins (e.g. MKL1), or the ternary complex factor (TCF) family of ETS proteins. In primary hematopoietic cells, knockout of either SRF or MKL1 decreases megakaryocyte maturation caused thrombocytopenia, and MKL1 over-expression (MKL1OE) in the Human Erythroleukemia (HEL) cell line enhances megakaryopoiesis, but the mechanisms underlying these effects are unknown. To elucidate the role of SRF and MKL1 in megakaryopoiesis, integrated analysis was performed of anti-SRF ChIP-seq and RNA-seq data in undifferentiated and differentiated HEL cells, both with and without induction of MKL1OE. MKL1OE enhances TPA-induced megakaryopoiesis with 25% of genes upregulated, as opposed to 11% with TPA alone. MKL1OE increases SRF binding to genomic sites, and enhances expression of specific target cytoskeletal genes in response to TPA. Genes upregulated in response to TPA are predicted to be regulated by SRF and ETS factors, whereas those upregulated in TPA plus MKL1OE lack ETS binding motifs. Using ChIP-PCR, we validated that both SRF and MKL1 have increased binding at target upregulated genes, e.g. CORO1A, with MKL1OE. We show for the first time that MKL1 increases both the genomic association and activity of SRF, and upregulates genes that enhance megakaryopoiesis.
2018-10-29 | GSE112277 | GEO
Project description:SRF is a ubiquitous transcription factor that binds DNA in association with myocardin-family proteins (e.g. MKL1), or the ternary complex factor (TCF) family of ETS proteins. In primary hematopoietic cells, knockout of either SRF or MKL1 decreases megakaryocyte maturation caused thrombocytopenia, and MKL1 over-expression (MKL1OE) in the Human Erythroleukemia (HEL) cell line enhances megakaryopoiesis, but the mechanisms underlying these effects are unknown. To elucidate the role of SRF and MKL1 in megakaryopoiesis, integrated analysis was performed of anti-SRF ChIP-seq and RNA-seq data in undifferentiated and differentiated HEL cells, both with and without induction of MKL1OE. MKL1OE enhances TPA-induced megakaryopoiesis with 25% of genes upregulated, as opposed to 11% with TPA alone. MKL1OE increases SRF binding to genomic sites, and enhances expression of specific target cytoskeletal genes in response to TPA. Genes upregulated in response to TPA are predicted to be regulated by SRF and ETS factors, whereas those upregulated in TPA plus MKL1OE lack ETS binding motifs. Using ChIP-PCR, we validated that both SRF and MKL1 have increased binding at target upregulated genes, e.g. CORO1A, with MKL1OE. We show for the first time that MKL1 increases both the genomic association and activity of SRF, and upregulates genes that enhance megakaryopoiesis.
2018-10-29 | GSE112265 | GEO
Project description:MKL1 augments megakaryocyte maturation by enhancing the SRF regulatory axis
| PRJNA445459 | ENA
Project description:MKL1 augments megakaryocyte maturation by enhancing the SRF regulatory axis [RNA-seq]
| PRJNA445458 | ENA
Project description:MKL1 augments megakaryocyte maturation by enhancing the SRF regulatory axis [ChIP-seq]
| PRJNA445433 | ENA
Project description:Serum response factor and its transcriptional cofactor MKL1 are critical for megakaryocyte maturation and platelet formation. We show that MKL2, a homologue of MKL1, is expressed in megakaryocytes and plays a role in megakaryocyte maturation. Using a megakaryocyte-specific Mkl2 knockout (KO) mouse on the conventional Mkl1 KO background to produce double KO (DKO) megakaryocytes and platelets, a critical role for MKL2 is revealed. The decrease in megakaryocyte ploidy and platelet counts of DKO mice is more severe than in Mkl1 KO mice. Platelet dysfunction in DKO mice is revealed by prolonged bleeding times and ineffective platelet activation in vitro in response to adenosine 5'-diphosphate. Electron microscopy and immunofluorescence of DKO megakaryocytes and platelets indicate abnormal cytoskeletal and membrane organization with decreased granule complexity. Surprisingly, the DKO mice have a more extreme thrombocytopenia than mice lacking serum response factor (SRF) expression in the megakaryocyte compartment. Comparison of gene expression reveals approximately 4400 genes whose expression is differentially affected in DKO compared with megakaryocytes deficient in SRF, strongly suggesting that MKL1 and MKL2 have both SRF-dependent and SRF-independent activity in megakaryocytopoiesis.
| S-EPMC3447785 | biostudies-literature
Project description:Active brown adipose tissue is responsible for non-shivering thermogenesis in mammals which affects energy homeostasis. The molecular mechanisms underlying this activation as well as the formation and activation of brite adipocytes have gained increasing interest in recent years as they might be utilized to regulate systemic metabolism. We show here that the transcriptional regulators SRF and MKL1 both act as repressors of brown adipogenesis. Loss-of-function of these transcription factors leads to a significant induction of brown adipocyte differentiation, increased levels of UCP1 and other thermogenic genes as well as increased respiratory function, while SRF induction exerts the opposite effects. Interestingly, we observed that knockdown of MKL1 does not lead to a reduced expression of typical SRF target genes and that the SRF/MKL1 inhibitor CCG-1423 had no significant effects on brown adipocyte differentiation. Contrary, knockdown of MKL1 induces a significant increase in the transcriptional activity of PPARγ target genes and MKL1 interacts with PPARγ, suggesting that SRF and MKL1 independently inhibit brown adipogenesis and that MKL1 exerts its effect mainly by modulating PPARγ activity.
| S-EPMC5268445 | biostudies-literature
Project description:Megakaryoblastic leukemia 1 (MKL1), identified as part of the t(1;22) translocation specific to acute megakaryoblastic leukemia, is highly expressed in differentiated muscle cells and promotes muscle differentiation by activating serum response factor (SRF). Here we show that Mkl1 expression is up-regulated during murine megakaryocytic differentiation and that enforced overexpression of MKL1 enhances megakaryocytic differentiation. When the human erythroleukemia (HEL) cell line is induced to differentiate with 12-O-tetradecanoylphorbol 13-acetate, overexpression of MKL1 results in an increased number of megakaryocytes with a concurrent increase in ploidy. MKL1 overexpression also promotes megakaryocytic differentiation of primary human CD34(+) cells cultured in the presence of thrombopoietin. The effect of MKL1 is abrogated when SRF is knocked down, suggesting that MKL1 acts through SRF. Consistent with these findings in human cells, knockout of Mkl1 in mice leads to reduced platelet counts in peripheral blood, and reduced ploidy in bone marrow megakaryocytes. In conclusion, MKL1 promotes physiologic maturation of human and murine megakaryocytes.
| S-EPMC2661865 | biostudies-other
Project description:Laminopathies, caused by mutations in the LMNA gene encoding the nuclear envelope proteins lamins A and C, represent a diverse group of diseases that include Emery-Dreifuss muscular dystrophy (EDMD), dilated cardiomyopathy (DCM), limb-girdle muscular dystrophy, and Hutchison-Gilford progeria syndrome. Most LMNA mutations affect skeletal and cardiac muscle by mechanisms that remain incompletely understood. Loss of structural function and altered interaction of mutant lamins with (tissue-specific) transcription factors have been proposed to explain the tissue-specific phenotypes. Here we report in mice that lamin-A/C-deficient (Lmna(-/-)) and Lmna(N195K/N195K) mutant cells have impaired nuclear translocation and downstream signalling of the mechanosensitive transcription factor megakaryoblastic leukaemia 1 (MKL1), a myocardin family member that is pivotal in cardiac development and function. Altered nucleo-cytoplasmic shuttling of MKL1 was caused by altered actin dynamics in Lmna(-/-) and Lmna(N195K/N195K) mutant cells. Ectopic expression of the nuclear envelope protein emerin, which is mislocalized in Lmna mutant cells and also linked to EDMD and DCM, restored MKL1 nuclear translocation and rescued actin dynamics in mutant cells. These findings present a novel mechanism that could provide insight into the disease aetiology for the cardiac phenotype in many laminopathies, whereby lamin A/C and emerin regulate gene expression through modulation of nuclear and cytoskeletal actin polymerization.
| S-EPMC3666313 | biostudies-literature