RESUMEN
Transcriptional memory of gene expression enables adaptation to repeated stimuli across many organisms. However, the regulation and heritability of transcriptional memory in single cells and through divisions remains poorly understood. Here, we combined microfluidics with single-cell live imaging to monitor Saccharomyces cerevisiae galactokinase 1 (GAL1) expression over multiple generations. By applying pedigree analysis, we dissected and quantified the maintenance and inheritance of transcriptional reinduction memory in individual cells through multiple divisions. We systematically screened for loss- and gain-of-memory knockouts to identify memory regulators in thousands of single cells. We identified new loss-of-memory mutants, which affect memory inheritance into progeny. We also unveiled a gain-of-memory mutant, elp6Δ, and suggest that this new phenotype can be mediated through decreased histone occupancy at the GAL1 promoter. Our work uncovers principles of maintenance and inheritance of gene expression states and their regulators at the single-cell level.
Asunto(s)
Galactoquinasa/genética , Regulación Fúngica de la Expresión Génica/genética , Transcripción Genética/genética , Galactosa/metabolismo , Expresión Génica/genética , Genes Fúngicos/genética , Herencia/genética , Histonas/metabolismo , Regiones Promotoras Genéticas/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Análisis de la Célula Individual/métodosRESUMEN
Changes in histone modifications are an attractive model through which environmental signals, such as diet, could be integrated in the cell for regulating its lifespan. However, evidence linking dietary interventions with specific alterations in histone modifications that subsequently affect lifespan remains elusive. We show here that deletion of histone N-alpha-terminal acetyltransferase Nat4 and loss of its associated H4 N-terminal acetylation (N-acH4) extend yeast replicative lifespan. Notably, nat4Δ-induced longevity is epistatic to the effects of calorie restriction (CR). Consistent with this, (i) Nat4 expression is downregulated and the levels of N-acH4 within chromatin are reduced upon CR, (ii) constitutive expression of Nat4 and maintenance of N-acH4 levels reduces the extension of lifespan mediated by CR, and (iii) transcriptome analysis indicates that nat4Δ largely mimics the effects of CR, especially in the induction of stress-response genes. We further show that nicotinamidase Pnc1, which is typically upregulated under CR, is required for nat4Δ-mediated longevity. Collectively, these findings establish histone N-acH4 as a regulator of cellular lifespan that links CR to increased stress resistance and longevity.
Asunto(s)
Restricción Calórica , Regulación Fúngica de la Expresión Génica , Histonas/metabolismo , Acetiltransferasa D N-Terminal/deficiencia , Acetiltransferasa D N-Terminal/fisiología , Proteínas de Saccharomyces cerevisiae/fisiología , Saccharomyces cerevisiae/fisiología , Acetilación , Cromatina/metabolismo , Regulación hacia Abajo , Perfilación de la Expresión Génica , Histona Acetiltransferasas/metabolismo , Longevidad , Acetiltransferasa D N-Terminal/genética , Nicotinamidasa/genética , Nicotinamidasa/metabolismo , Procesamiento Proteico-Postraduccional , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Factores de Tiempo , Activación TranscripcionalRESUMEN
BACKGROUND: Transcriptome studies have revealed that many eukaryotic genomes are pervasively transcribed producing numerous long non-coding RNAs (lncRNAs). However, only a few lncRNAs have been ascribed a cellular role thus far, with most regulating the expression of adjacent genes. Even less lncRNAs have been annotated as essential hence implying that the majority may be functionally redundant. Therefore, the function of lncRNAs could be illuminated through systematic analysis of their synthetic genetic interactions (GIs). RESULTS: Here, we employ synthetic genetic array (SGA) in Saccharomyces cerevisiae to identify GIs between long intergenic non-coding RNAs (lincRNAs) and protein-coding genes. We first validate this approach by demonstrating that the telomerase RNA TLC1 displays a GI network that corresponds to its well-described function in telomere length maintenance. We subsequently performed SGA screens on a set of uncharacterised lincRNAs and uncover their connection to diverse cellular processes. One of these lincRNAs, SUT457, exhibits a GI profile associating it to telomere organisation and we consistently demonstrate that SUT457 is required for telomeric overhang homeostasis through an Exo1-dependent pathway. Furthermore, the GI profile of SUT457 is distinct from that of its neighbouring genes suggesting a function independent to its genomic location. Accordingly, we show that ectopic expression of this lincRNA suppresses telomeric overhang accumulation in sut457Δ cells assigning a trans-acting role for SUT457 in telomere biology. CONCLUSIONS: Overall, our work proposes that systematic application of this genetic approach could determine the functional significance of individual lncRNAs in yeast and other complex organisms.
Asunto(s)
Genoma Fúngico , ARN Largo no Codificante/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/genética , Telómero/genética , ADN de Hongos/genética , Exodesoxirribonucleasas/genética , Exodesoxirribonucleasas/metabolismo , Perfilación de la Expresión Génica , Ontología de Genes , Genómica , Proteínas de Saccharomyces cerevisiae/genética , Telomerasa/genética , Telomerasa/metabolismoRESUMEN
BACKGROUND AND AIMS: Human metastatic prostate cancer cell growth can be inhibited by GnRH analogs but effects on virus-immortalized prostate cells have not been investigated. METHODS: Virus-immortalized prostate cells were stably transfected with rat GnRH receptor cDNA and levels of GnRH binding were correlated with GnRH effects on signaling, cell cycle, growth, exosome production, and apoptosis. RESULTS: High levels of cell surface GnRH receptor occurred in transfected papillomavirus-immortalized WPE-1-NB26 epithelial cells but not in non-tumourigenic RWPE-1, myoepithelial WPMY-1 cells, or SV40-immortalized PNT1A. Endogenous cell surface GnRH receptor was undetectable in non-transfected cells or cancer cell lines LNCaP, PC3, and DU145. GnRH receptor levels correlated with induction of inositol phosphates, elevation of intracellular Ca(2+) , cytoskeletal actin reorganization, modulation of ERK activation and cell growth-inhibition with GnRH agonists. Hoechst 33342 DNA staining-cell sorting indicated accumulation of cells in G2 following agonist treatment. Release of exosomes from transfected WPE-1-NB26 was unaffected by agonists, unlike induction observed in HEK293([SCL60]) cells. Increased PARP cleavage and apoptotic body production were undetectable during growth-inhibition in WPE-1-NB26 cells, contrasting with HEK293([SCL60]) . EGF receptor activation inhibited GnRH-induced ERK activation in WPE-1-NB26 but growth-inhibition was not rescued by EGF or PKC inhibitor Ro320432. Growth of cells expressing low levels of GnRH receptor was not affected by agonists. CONCLUSIONS: Engineered high-level GnRH receptor activation inhibits growth of a subset of papillomavirus-immortalized prostate cells. Elucidating mechanisms leading to clone-specific differences in cell surface GnRH receptor levels is a valuable next step in developing strategies to exploit prostate cell anti-proliferation using GnRH agonists.
Asunto(s)
Hormona Liberadora de Gonadotropina/agonistas , Neoplasias de la Próstata/metabolismo , Receptores LHRH/biosíntesis , Pamoato de Triptorelina/farmacología , Alphapapillomavirus/fisiología , Apoptosis/fisiología , Ciclo Celular/fisiología , Línea Celular Tumoral , Transformación Celular Viral , Exosomas/fisiología , Citometría de Flujo , Hormona Liberadora de Gonadotropina/metabolismo , Humanos , Fosfatos de Inositol/metabolismo , Masculino , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/virología , Receptores LHRH/genética , Receptores LHRH/metabolismo , Transducción de Señal/fisiología , TransfecciónRESUMEN
Intraosseous venous malformations are rare benign lesions that account for approximately 0.5% to 1% of all osseous tumors. Involvement of the facial skeleton is rather uncommon, with the mandible and maxilla most frequently involved. In the current study, we report a case of an intraosseous venous malformation in a 52-year-old male patient with a history of mandibular keratocystic odontogenic tumor. The patient presented with an asymptomatic bony lesion in the mandible. Treatment involved surgical excision. Histopathologic examination of the excised specimen revealed an intraosseous venous malformation. Twelve months postoperatively, there was no evidence of recurrence.
Asunto(s)
Mandíbula/irrigación sanguínea , Malformaciones Vasculares/diagnóstico por imagen , Malformaciones Vasculares/cirugía , Tomografía Computarizada de Haz Cónico , Diagnóstico Diferencial , Humanos , Masculino , Persona de Mediana Edad , Radiografía PanorámicaRESUMEN
The binding of GnRH to its receptor on pituitary gonadotropes leads to the targeting of a diverse array of signalling mediators. These mediators drive multiple signal transduction pathways, which in turn regulate a variety of cellular processes, including the biosynthesis and secretion of the gonadotropins LH and FSH. Advances in our understanding of the mechanisms and signalling pathways that are recruited to regulate gonadotrope function are continually being made. This review will focus on the recent demonstration that key mediators of the canonical Wnt signalling pathway are targeted by GnRH in gonadotropes, and that these may play essential roles in regulating the expression of many of the key players in gonadotrope biology, including the GnRH receptor and the gonadotropins.