RESUMO
Senescent cells are a major contributor to age-dependent cardiovascular tissue dysfunction, but knowledge of their in vivo cell markers and tissue context is lacking. To reveal tissue-relevant senescence biology, we integrate the transcriptomes of 10 experimental senescence cell models with a 224 multi-tissue gene co-expression network based on RNA-seq data of seven tissues biopsies from â¼600 coronary artery disease (CAD) patients. We identify 56 senescence-associated modules, many enriched in CAD GWAS genes and correlated with cardiometabolic traits-which supports universality of senescence gene programs across tissues and in CAD. Cross-tissue network analyses reveal 86 candidate senescence-associated secretory phenotype (SASP) factors, including COL6A3. Experimental knockdown of COL6A3 induces transcriptional changes that overlap the majority of the experimental senescence models, with cell-cycle arrest linked to modulation of DREAM complex-targeted genes. We provide a transcriptomic resource for cellular senescence and identify candidate biomarkers, SASP factors, and potential drivers of senescence in human tissues.
Assuntos
Senescência Celular , Transcriptoma , Humanos , Transcriptoma/genética , Senescência Celular/genética , Fenótipo , Biomarcadores , Colágeno , Colágeno Tipo VI/genéticaRESUMO
Attenuating pathological angiogenesis in diseases characterized by neovascularization such as diabetic retinopathy has transformed standards of care. Yet little is known about the molecular signatures discriminating physiological blood vessels from their diseased counterparts, leading to off-target effects of therapy. We demonstrate that in contrast to healthy blood vessels, pathological vessels engage pathways of cellular senescence. Senescent (p16INK4A-expressing) cells accumulate in retinas of patients with diabetic retinopathy and during peak destructive neovascularization in a mouse model of retinopathy. Using either genetic approaches that clear p16INK4A-expressing cells or small molecule inhibitors of the anti-apoptotic protein BCL-xL, we show that senolysis suppresses pathological angiogenesis. Single-cell analysis revealed that subsets of endothelial cells with senescence signatures and expressing Col1a1 are no longer detected in BCL-xL-inhibitor-treated retinas, yielding a retina conducive to physiological vascular repair. These findings provide mechanistic evidence supporting the development of BCL-xL inhibitors as potential treatments for neovascular retinal disease.
Assuntos
Senescência Celular , Doenças Retinianas/patologia , Proteína bcl-X/metabolismo , Animais , Apoptose/efeitos dos fármacos , Senescência Celular/efeitos dos fármacos , Cadeia alfa 1 do Colágeno Tipo I/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/deficiência , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Modelos Animais de Doenças , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Feminino , Flavonóis/química , Flavonóis/farmacologia , Flavonóis/uso terapêutico , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neovascularização Patológica , Doenças Retinianas/tratamento farmacológico , Doenças Retinianas/metabolismo , Tacrolimo/análogos & derivados , Tacrolimo/farmacologia , Proteína bcl-X/antagonistas & inibidoresRESUMO
We elucidated the molecular cross-talk between cartilage and synovium in osteoarthritis, the most widespread arthritis in the world, using the powerful tool of single-cell RNA-sequencing. Multiple cell types were identified based on profiling of 10,640 synoviocytes and 26,192 chondrocytes: 12 distinct synovial cell types and 7 distinct articular chondrocyte phenotypes from matched tissues. Intact cartilage was enriched for homeostatic and hypertrophic chondrocytes, while damaged cartilage was enriched for prefibro- and fibro-, regulatory, reparative and prehypertrophic chondrocytes. A total of 61 cytokines and growth factors were predicted to regulate the 7 chondrocyte cell phenotypes. Based on production by > 1% of cells, 55% of the cytokines were produced by synovial cells (39% exclusive to synoviocytes and not expressed by chondrocytes) and their presence in osteoarthritic synovial fluid confirmed. The synoviocytes producing IL-1beta (a classic pathogenic cytokine in osteoarthritis), mainly inflammatory macrophages and dendritic cells, were characterized by co-expression of surface proteins corresponding to HLA-DQA1, HLA-DQA2, OLR1 or TLR2. Strategies to deplete these pathogenic intra-articular cell subpopulations could be a therapeutic option for human osteoarthritis.
Assuntos
Biomarcadores/metabolismo , Cartilagem Articular/metabolismo , Osteoartrite/patologia , RNA-Seq/métodos , Sinoviócitos/metabolismo , Idoso , Cartilagem Articular/patologia , Estudos de Casos e Controles , Células Cultivadas , Feminino , Humanos , Masculino , Osteoartrite/genética , Osteoartrite/metabolismo , Sinoviócitos/patologiaRESUMO
BACKGROUND: Algae accumulate lipids to endure different kinds of environmental stresses including macronutrient starvation. Although this response has been extensively studied, an in depth understanding of the transcriptional regulatory network (TRN) that controls the transition into lipid accumulation remains elusive. In this study, we used a systems biology approach to elucidate the transcriptional program that coordinates the nitrogen starvation-induced metabolic readjustments that drive lipid accumulation in Chlamydomonas reinhardtii. RESULTS: We demonstrate that nitrogen starvation triggered differential regulation of 2147 transcripts, which were co-regulated in 215 distinct modules and temporally ordered as 31 transcriptional waves. An early-stage response was triggered within 12 min that initiated growth arrest through activation of key signaling pathways, while simultaneously preparing the intracellular environment for later stages by modulating transport processes and ubiquitin-mediated protein degradation. Subsequently, central metabolism and carbon fixation were remodeled to trigger the accumulation of triacylglycerols. Further analysis revealed that these waves of genome-wide transcriptional events were coordinated by a regulatory program orchestrated by at least 17 transcriptional regulators, many of which had not been previously implicated in this process. We demonstrate that the TRN coordinates transcriptional downregulation of 57 metabolic enzymes across a period of nearly 4 h to drive an increase in lipid content per unit biomass. Notably, this TRN appears to also drive lipid accumulation during sulfur starvation, while phosphorus starvation induces a different regulatory program. The TRN model described here is available as a community-wide web-resource at http://networks.systemsbiology.net/chlamy-portal. CONCLUSIONS: In this work, we have uncovered a comprehensive mechanistic model of the TRN controlling the transition from N starvation to lipid accumulation. The program coordinates sequentially ordered transcriptional waves that simultaneously arrest growth and lead to lipid accumulation. This study has generated predictive tools that will aid in devising strategies for the rational manipulation of regulatory and metabolic networks for better biofuel and biomass production.
RESUMO
We demonstrate a system for cloning and modifying the chloroplast genome from the green alga, Chlamydomonas reinhardtii. Through extensive use of sequence stabilization strategies, the ex vivo genome is assembled in yeast from a collection of overlapping fragments. The assembled genome is then moved into bacteria for large-scale preparations and transformed into C. reinhardtii cells. This system also allows for the generation of simultaneous, systematic and complex genetic modifications at multiple loci in vivo. We use this system to substitute genes encoding core subunits of the photosynthetic apparatus with orthologs from a related alga, Scenedesmus obliquus. Once transformed into algae, the substituted genome recombines with the endogenous genome, resulting in a hybrid plastome comprising modifications in disparate loci. The in vivo function of the genomes described herein demonstrates that simultaneous engineering of multiple sites within the chloroplast genome is now possible. This work represents the first steps toward a novel approach for creating genetic diversity in any or all regions of a chloroplast genome.
Assuntos
Chlamydomonas reinhardtii/genética , Genoma de Cloroplastos , Clonagem Molecular , Complexo de Proteína do Fotossistema II/genética , Subunidades Proteicas/genética , Biologia Sintética/métodos , Transformação GenéticaRESUMO
To identify genes involved in the process of germ-cell formation in Drosophila, a maternal-effect screen using the FLP/FRT-ovoD method was performed on chromosome 3R. In addition to expected mutations in the germ-cell determinant oskar and in other genes known to be involved in the process, several novel mutations caused defects in germ-cell formation. Mutations in any of three genes [l(3)malignant brain tumor, shackleton, and out of sync] affect the synchronous mitotic divisions and nuclear migration of the early embryo. The defects in nuclear migration or mitotic synchrony result in a reduction in germ-cell formation. Mutations in another gene identified in this screen, bebra, do not cause mitotic defects, but appear to act upstream of the localization of oskar. Analysis of our mutants demonstrates that two unique and independent processes must occur to form germ cells-germ-plasm formation and nuclear division/migration.