RESUMO
Inflammatory signaling is required for hematopoietic stem and progenitor cell (HSPC) development. Here, we studied the involvement of RIG-I-like receptors (RLRs) in HSPC formation. Rig-I or Mda5 deficiency impaired, while Lgp2 deficiency enhanced, HSPC emergence in zebrafish embryos. Rig-I or Mda5 deficiency reduced HSPC numbers by inhibiting inflammatory signals that were in turn enhanced in Lgp2 deficient embryos. Simultaneous reduction of Lgp2 and either Rig-I or Mda5 rescued inflammatory signals and HSPC numbers. Modulating the expression of the signaling mediator Traf6 in RLR deficient embryos restored HSPC numbers. Repetitive element transcripts could be detected in hemogenic endothelial cells and HSPCs, suggesting a role as RLR ligands. Indeed, ectopic expression of repetitive elements enhanced HSPC formation in wild-type, but not in Rig-I or Mda5 deficient embryos. Manipulation of RLR expression in mouse fetal liver HSPCs indicated functional conservation among species. Thus, repetitive elements transcribed during development drive RLR-mediated inflammatory signals that regulate HSPC formation.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Células-Tronco Hematopoéticas/metabolismo , Sequências Repetitivas de Ácido Nucleico , Transdução de Sinais , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo , Animais , Biomarcadores , Montagem e Desmontagem da Cromatina , Elementos de DNA Transponíveis , Suscetibilidade a Doenças , Hematopoese/genética , Células-Tronco Hematopoéticas/citologia , Imunidade Inata , Imuno-Histoquímica , Inflamação/etiologia , Inflamação/metabolismo , Inflamação/patologia , RNA Helicases/deficiência , RNA Helicases/genética , Proteínas de Ligação a RNA/metabolismo , Fator 6 Associado a Receptor de TNF/metabolismo , Ácido Valproico/farmacologia , Peixe-ZebraRESUMO
The clinical benefit of MAPK pathway inhibition in melanoma patients carrying BRAF mutations is temporal. After the initial response to treatment, the majority of tumors will develop resistance and patients will relapse. Here we demonstrate that the endothelin-endothelin receptor B (ETBR) signaling pathway confers resistance to MAPK pathway inhibitors in BRAF mutated melanoma. MAPK blockade, in addition to being anti-proliferative, induces a phenotypic change which is characterized by increased expression of melanocyte-specific genes including ETBR. In the presence of MAPK inhibitors, activation of ETBR by endothelin enables the sustained proliferation of melanoma cells. In mouse models of melanoma, including patient-derived xenograft models, concurrent inhibition of the MAPK pathway and ETBR signaling resulted in a more effective anti-tumor response compared to MAPK pathway inhibition alone. The combination treatment significantly reduced tumor growth and prolonged survival compared to therapies with MAPK pathway inhibitors alone. The phosphoproteomic analysis revealed that ETBR signaling did not induce resistance towards MAPK pathway inhibitors by restoring MAPK activity, but instead via multiple alternative signaling pathways downstream of the small G proteins GNAq/11. Together these data indicate that a combination of MAPK pathway inhibitors with ETBR antagonists could have a synergistically beneficial effect in melanoma patients with hyperactivated MAPK signaling pathways.
Assuntos
Melanoma/tratamento farmacológico , Recidiva Local de Neoplasia/tratamento farmacológico , Proteínas Proto-Oncogênicas B-raf/genética , Receptor de Endotelina B/genética , Animais , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Antagonistas do Receptor de Endotelina B/farmacologia , Humanos , Melanoma/genética , Melanoma/patologia , Camundongos , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Mutação/genética , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/patologia , Inibidores de Proteínas Quinases/farmacologia , Transdução de Sinais/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Haematopoietic stem cells (HSCs) are normally quiescent, but have evolved mechanisms to respond to stress. Here, we evaluate haematopoietic regeneration induced by chemotherapy. We detect robust chromatin reorganization followed by increased transcription of transposable elements (TEs) during early recovery. TE transcripts bind to and activate the innate immune receptor melanoma differentiation-associated protein 5 (MDA5) that generates an inflammatory response that is necessary for HSCs to exit quiescence. HSCs that lack MDA5 exhibit an impaired inflammatory response after chemotherapy and retain their quiescence, with consequent better long-term repopulation capacity. We show that the overexpression of ERV and LINE superfamily TE copies in wild-type HSCs, but not in Mda5-/- HSCs, results in their cycling. By contrast, after knockdown of LINE1 family copies, HSCs retain their quiescence. Our results show that TE transcripts act as ligands that activate MDA5 during haematopoietic regeneration, thereby enabling HSCs to mount an inflammatory response necessary for their exit from quiescence.
Assuntos
Proliferação de Células/efeitos dos fármacos , Senescência Celular/efeitos dos fármacos , Elementos de DNA Transponíveis , Hematopoese/efeitos dos fármacos , Células-Tronco Hematopoéticas/efeitos dos fármacos , Helicase IFIH1 Induzida por Interferon/metabolismo , Agonistas Mieloablativos/farmacologia , Animais , Montagem e Desmontagem da Cromatina/efeitos dos fármacos , Retrovirus Endógenos/genética , Ativação Enzimática , Células HEK293 , Células-Tronco Hematopoéticas/enzimologia , Humanos , Helicase IFIH1 Induzida por Interferon/genética , Ligantes , Elementos Nucleotídeos Longos e Dispersos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de SinaisRESUMO
Chirality is a property of asymmetry between an object and its mirror image. Most biomolecules and many cell types are chiral. In the left-right organizer (LRO), cilia-driven flows transfer such chirality to the body scale. However, the existence of cellular chirality within tissues remains unknown. Here, we investigate this question in Kupffer's vesicle (KV), the zebrafish LRO. Quantitative live imaging reveals that cilia populating the KV display asymmetric orientation between the right and left sides, resulting in a chiral structure, which is different from the chiral cilia rotation. This KV chirality establishment is dynamic and depends on planar cell polarity. While its impact on left-right (LR) symmetry breaking remains unclear, we show that this asymmetry does not depend on the LR signaling pathway or flow. This work identifies a different type of tissue asymmetry and sheds light on chirality genesis in developing tissues.
Assuntos
Padronização Corporal , Cílios/metabolismo , Peixe-Zebra/embriologia , Animais , Corpos Basais/metabolismo , Organizadores Embrionários/fisiologia , Proteínas de Peixe-Zebra/metabolismoRESUMO
The H2.0-like homeobox transcription factor (HLX) regulates hematopoietic differentiation and is overexpressed in Acute Myeloid Leukemia (AML), but the mechanisms underlying these functions remain unclear. We demonstrate here that HLX overexpression leads to a myeloid differentiation block both in zebrafish and human hematopoietic stem and progenitor cells (HSPCs). We show that HLX overexpression leads to downregulation of genes encoding electron transport chain (ETC) components and upregulation of PPARδ gene expression in zebrafish and human HSPCs. HLX overexpression also results in AMPK activation. Pharmacological modulation of PPARδ signaling relieves the HLX-induced myeloid differentiation block and rescues HSPC loss upon HLX knockdown but it has no effect on AML cell lines. In contrast, AMPK inhibition results in reduced viability of AML cell lines, but minimally affects myeloid progenitors. This newly described role of HLX in regulating the metabolic state of hematopoietic cells may have important therapeutic implications.