RESUMEN
SAG21/LEA5 is an unusual late embryogenesis abundant protein in Arabidopsis thaliana, that is primarily mitochondrially located and may be important in regulating translation in both chloroplasts and mitochondria. SAG21 expression is regulated by a plethora of abiotic and biotic stresses and plant growth regulators indicating a complex regulatory network. To identify key transcription factors regulating SAG21 expression, yeast-1-hybrid screens were used to identify transcription factors that bind the 1685 bp upstream of the SAG21 translational start site. Thirty-three transcription factors from nine different families bound to the SAG21 promoter, including members of the ERF, WRKY and NAC families. Key binding sites for both NAC and WRKY transcription factors were tested through site directed mutagenesis indicating the presence of cryptic binding sites for both these transcription factor families. Co-expression in protoplasts confirmed the activation of SAG21 by WRKY63/ABO3, and SAG21 upregulation elicited by oligogalacturonide elicitors was partially dependent on WRKY63, indicating its role in SAG21 pathogen responses. SAG21 upregulation by ethylene was abolished in the erf1 mutant, while wound-induced SAG21 expression was abolished in anac71 mutants, indicating SAG21 expression can be regulated by several distinct transcription factors depending on the stress condition.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Factores de Transcripción/metabolismo , Arabidopsis/metabolismo , Regulación de la Expresión Génica de las Plantas , Proteínas de Arabidopsis/metabolismo , Oxidación-Reducción , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Estrés FisiológicoRESUMEN
While the acquisition of cellular plasticity in adult stem cells is essential for rapid regeneration after tissue injury, little is known about the underlying mechanisms governing this process. Our data reveal the coordination of airway progenitor differentiation plasticity by inflammatory signals during alveolar regeneration. Following damage, interleukin-1ß (IL-1ß) signalling-dependent modulation of Jag1 and Jag2 expression in ciliated cells results in the inhibition of Notch signalling in secretory cells, which drives the reprogramming and acquisition of differentiation plasticity. We identify the transcription factor Fosl2 (also known as Fra2) for secretory cell fate conversion to alveolar type 2 cells that retain the distinct genetic and epigenetic signatures of secretory lineages. We also reveal that human secretory cells positive for KDR (also known as FLK-1) display a conserved capacity to generate alveolar type 2 cells via Notch inhibition. Our results demonstrate the functional role of an IL-1ß-Notch-Fosl2 axis in the fate decision of secretory cells during injury repair, proposing a potential therapeutic target for human lung alveolar regeneration.
Asunto(s)
Diferenciación Celular/fisiología , Antígeno 2 Relacionado con Fos/metabolismo , Interleucina-1beta/metabolismo , Receptores Notch/metabolismo , Regeneración/fisiología , Animales , Antígeno 2 Relacionado con Fos/genética , Regulación de la Expresión Génica/fisiología , Interleucina-1beta/genética , Ratones , Sistema Respiratorio/metabolismo , Transducción de Señal/fisiología , Células Madre/metabolismoRESUMEN
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is the cause of a present pandemic, infects human lung alveolar type 2 (hAT2) cells. Characterizing pathogenesis is crucial for developing vaccines and therapeutics. However, the lack of models mirroring the cellular physiology and pathology of hAT2 cells limits the study. Here, we develop a feeder-free, long-term, three-dimensional (3D) culture technique for hAT2 cells derived from primary human lung tissue and investigate infection response to SARS-CoV-2. By imaging-based analysis and single-cell transcriptome profiling, we reveal rapid viral replication and the increased expression of interferon-associated genes and proinflammatory genes in infected hAT2 cells, indicating a robust endogenous innate immune response. Further tracing of viral mutations acquired during transmission identifies full infection of individual cells effectively from a single viral entry. Our study provides deep insights into the pathogenesis of SARS-CoV-2 and the application of defined 3D hAT2 cultures as models for respiratory diseases.
Asunto(s)
COVID-19 , Alveolos Pulmonares/virología , SARS-CoV-2/fisiología , Células Madre/virología , COVID-19/virología , Técnicas de Cultivo de Célula , Medios de Cultivo , Humanos , Interferones/metabolismo , Modelos Biológicos , Alveolos Pulmonares/metabolismo , Alveolos Pulmonares/ultraestructura , SARS-CoV-2/ultraestructura , Transcriptoma , Internalización del Virus , Replicación ViralRESUMEN
Diseases such as idiopathic pulmonary fibrosis, chronic obstructive pulmonary disease, and bronchopulmonary dysplasia injure the gas-exchanging alveoli of the human lung. Animal studies have indicated that dysregulation of alveolar cells, including alveolar type II stem/progenitor cells, is implicated in disease pathogenesis. Due to mouse-human differences, there has been a desperate need to develop human-relevant lung models that can more closely recapitulate the human lung during homeostasis, injury repair, and disease. Here we discuss how current single-cell RNA sequencing studies have increased knowledge of the cellular and molecular composition of human lung alveoli, including the identification of molecular heterogeneity, cellular diversity, and previously unknown cell types, some of which arise specifically during disease. For functional analysis of alveolar cells, in vitro human alveolar organoids established from human pluripotent stem cells, embryonic progenitors, and adult tissue from both healthy and diseased lungs have modeled aspects of the cellular and molecular features of alveolar epithelium. Drawbacks of such systems are highlighted, along with possible solutions. Organoid-on-a-chip and ex vivo systems including precision-cut lung slices can complement organoid studies by providing further cellular and structural complexity of lung tissues, and have been shown to be invaluable models of human lung disease, while the production of acellular and synthetic scaffolds hold promise in lung transplant efforts. Further improvements to such systems will increase understanding of the underlying biology of human alveolar stem/progenitor cells, and could lead to future therapeutic or pharmacological intervention in patients suffering from end-stage lung diseases.