RESUMEN
Interactions among ecosystem services (ESs) involve tradeoffs and synergies. Quantitatively studying the trade-off and synergistic relationships between land use/land cover change (LULC) and ESs enables the precise identification of the quality status and driving factors of ESs within the region, which is crucial for rational resource allocation and environmental protection. In this study, the spatial and temporal change characteristics of the three ESs of carbon storage (CS), soil retention (SR) and habitat quality (HQ) are explored by using the InVEST model and GIS technology in the region around Taihu Lake, and the tradeoffs and synergies among the three are determined based on the difference comparison. The results indicate that: (1) The study area has a downward trajectory in CS and HQ from 1990 to 2020, while SR experiences some fluctuations. The spatial distribution of the three ESs exhibits high levels in the southwest and low levels in the northeast. (2) The most sensitive regions where tradeoffs and synergies are most pronounced occur primarily in the newly construction land regions and the southwestern mountainous and hilly areas. In newly construction land regions, there are often tradeoffs relationships observed between CS and SR, as well as between HQ and SR. Conversely, a predominantly negative synergy is mainly observed between CS and HQ. In the southwestern hilly terrain, due to changes in landscape patterns, HQ and SR exhibit higher levels of negative synergistic relationships. (3) LULC is a significant driver of spatial and temporal changes in ESs, as well as changes in tradeoffs and synergies in the study area, necessitating integrated research from economic, social and climate change perspectives.
RESUMEN
Submergence is a major constraint on rice production in South and Southeast Asia. In this study, we determined that a gene of the Sub1A-binding protein family, SAB23, encodes a plant homeodomain (PHD)-type transcription factor that has a novel function of negatively regulating submergence tolerance in rice. The T-DNA insertion mutant sab23 displayed reduced plant height, delayed seed maturation, and lower percentage seed set. Importantly, this mutant also exhibited enhanced submergence tolerance. In addition, CRISPR/Cas9 knock out of SAB23 resulted in a significant reduction in the content of the gibberellin GA4 and a dramatic increase in the content of GA1 in the plants. SAB23 binds to the promoter of CYTOCHROME P450 714B2 (CYP714B2), which encodes a GA13-oxidase that catalyses the conversion of GA53 to GA19. Disruption of SAB23 function led to increased CYP714B2 transcription, and overexpression of CYP714B2 produced phenotypes similar to those of the SAB23-knockout plants. Taken together, our results reveal that SAB23 negatively regulates rice submergence tolerance by modulating CYP714B2 expression, which has significant potential for use in future breeding.
Asunto(s)
Oryza , Factores de Transcripción , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Oryza/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Fitomejoramiento , Sistema Enzimático del Citocromo P-450/metabolismo , MutaciónRESUMEN
Kinases have been the focus of drug discovery programs for three decades leading to over 70 therapeutic kinase inhibitors and biophysical affinity measurements for over 130,000 kinase-compound pairs. Nonetheless, the precise target spectrum for many kinases remains only partly understood. In this study, we describe a computational approach to unlocking qualitative and quantitative kinome-wide binding measurements for structure-based machine learning. Our study has three components: (i) a Kinase Inhibitor Complex (KinCo) data set comprising in silico predicted kinase structures paired with experimental binding constants, (ii) a machine learning loss function that integrates qualitative and quantitative data for model training, and (iii) a structure-based machine learning model trained on KinCo. We show that our approach outperforms methods trained on crystal structures alone in predicting binary and quantitative kinase-compound interaction affinities; relative to structure-free methods, our approach also captures known kinase biochemistry and more successfully generalizes to distant kinase sequences and compound scaffolds.
Asunto(s)
Descubrimiento de Drogas , Aprendizaje Automático , Inhibidores de Proteínas Quinasas/farmacologíaRESUMEN
BACKGROUND: Long noncoding RNAs (lncRNAs) are involved in a diverse array of biological processes. While lncRNAs are commonly upregulated in hepatocellular carcinoma (HCC), the specific regulatory roles they play in this oncogenic context require further study and clarification. Although HULC (lncRNA highly upregulated in liver cancer) is involved in disease pathogenesis, its precise role in this context remains unclear. METHODS: Here, we have explored the mechanistic relevance of HULC expression by assessing its expression in patient samples. The importance of this lncRNA in the onset and progression of HCC was investigated through in vitro approaches including Western blotting, quantitative PCR, Transwell assays, electron microscopy, wound healing assays, and real-time cell analysis (RTCA). Additionally, the in vivo functions of this lncRNA were assessed using an orthotopic HCC xenograft in nude mouse model system. RESULTS: HULC was identified as a lncRNA that is highly upregulated in human liver tumors. In vitro, HULC was able to promote HCC malignancy, although its excess overexpression also led robust autophagic induction, promoting the increased expression of autophagy-associated genes including LC3 and Beclin-1. At a mechanistic level, HULC was able to promote the phosphorylation of p65 and IkBkB thus enhancing autophagy by increasing LC3II levels in a manner dependent upon the NF-κB pathway. HULC downregulation was also linked to impaired orthotopic HCC tumor growth in vivo. The link between HULC and autophagy may play a role in disease progression. CONCLUSIONS: These results suggest that HULC is an oncogenic lncRNA, and may thus offer value as a prognostic biomarker and promoter of HCC development, in addition to being a potential therapeutic target in this cancer type.
Asunto(s)
Carcinoma Hepatocelular , Neoplasias Hepáticas , ARN Largo no Codificante , Animales , Ratones , Humanos , Neoplasias Hepáticas/patología , Carcinoma Hepatocelular/patología , ARN Largo no Codificante/genética , ARN Largo no Codificante/metabolismo , FN-kappa B/metabolismo , Pronóstico , Biomarcadores , Regulación Neoplásica de la Expresión Génica , Línea Celular Tumoral , Proliferación CelularRESUMEN
Patients diagnosed with hepatocellular carcinoma (HCC) seek a satisfactory prognosis. However, most HCC patients present a risk of recurrence, thus highlighting the lack of effectiveness of current treatments and the urgent need for improved treatment options. The purpose of this study was to identify new candidate factors in the STAT family, which is involved in hepatocellular carcinogenesis, and new targets for the treatment of HCC. Bioinformatics web resources, including Oncomine, Gene Expression Profiling Interactive Analysis (GEPIA), The Human Protein Atlas (HPA), Tumor Immune Estimation Resource (TIMER), and GSCALite, were used to identify candidate genes among the STAT family in HCC. STAT1 was significantly overexpressed in hepatocellular carcinoma. More meaningfully, the high STAT1 expression was significantly associated with poor prognosis. Therefore, STAT1 is expected to be a therapeutic target. The JAK2 inhibitor lestaurtinib was screened by the Genomics of Cancer Drug Sensitivity Project (GDSC) analysis. Pharmacological experiments showed that lestaurtinib has the ability to prevent cell migration and colony formation from single cells. We also found that STAT1 is involved in inflammatory responses and immune cell infiltration. Immune infiltration analysis revealed a strong association between STAT1 levels and immune cell abundance, immune biomarker levels, and immune checkpoints. This study suggests that STAT1 may be a key oncogene in hepatocellular carcinoma and provides evidence that the JAK2 inhibitor lestaurtinib is a potent antiproliferative agent that warrants further investigation as a targeted therapy for HCC.
RESUMEN
Due to high-intensity human disturbance and rapid climate change, optimizing the spatial pattern of land use has become a pivotal path to restoring ecosystem functions and realizing the sustainable development of human-land relationships. This review uses the literature analysis method combined with CiteSpace to determine current research progress and frontiers, challenges, and directions for further improvement in this field. The main conclusions include the following: (a) research on the optimization of spatial pattern of land use has transformed from pattern description orientation to sustainable development orientation to ecological restoration orientation. Its research paradigm has changed from pattern to function to well-being; (b) the research frontier mainly includes spatial pattern of land use that takes into account the unity of spatial structure and functional attributes, the ecological mechanism and feedback effect of change in spatial pattern of land, the theoretical framework and model construction of land use simulation and prediction based on multiple disciplines and fields, and the adaptive management of sustainable land use in the context of climate change; (c) based on current research challenges, we integrate the research on landscape ecology and ecosystem service flows to develop an "element sets-network structure-system functions-human well-being" conceptual model. We also propose the strengthening of future research on theoretical innovation, spatiotemporal mechanism selection, causal emergence mechanism, the transformation threshold, and uncertainty. We provide innovative ideas for achieving sustainable management of land systems and territorial spatial planning with the aim of improving the adaptability of land use spatial optimization. This is expected to strengthen the ability of land systems to cope with ecological security and climate risks.
Asunto(s)
Conservación de los Recursos Naturales , Ecosistema , Cambio Climático , Humanos , Modelos Teóricos , Desarrollo SostenibleRESUMEN
Soil loss caused by erosion is a global problem. Therefore, the assessment of soil erosion and the its driving mechanism are of great significance to soil conservation. However, soil erosion is affected by both climate change and human activities, which have not been quantified, and few researchers studied the differences in the driving mechanisms of soil erosion depending on the land use type. Therefore, the spatiotemporal characteristics and changing trends of soil erosion in the Dongting Lake Basin were analyzed in this study. Geographic detectors were used to identify the dominant factors affecting soil erosion in different land use types. In this study, a sensitivity experiment was conducted to clarify the relative contributions of climate change and human activities to soil erosion changes. In addition, we studied the effects of different land use types and vegetation cover restoration on soil erosion. The results show that soil erosion in the Dongting Lake Basin decreased from 2000 to 2018. Human activities represented by land use types and vegetation coverage significantly contributed to the alleviation of soil erosion in the Dongting Lake Basin, whereas climate change represented by rainfall slightly aggravated soil erosion in the study area. The restoration of grassland vegetation and transfer of cultivated land to woodlands in the study area improved the soil erosion. The slope steepness is the key factor affecting the intensity of soil erosion in dry land, paddy fields, and unused land, whereas the vegetation coverage is the key factor affecting the intensity of soil erosion in woodland, garden land, and grassland. Detailed spatiotemporally mapping of soil erosion was used to determine the connections between soil erosion and potential drivers, which have important implications for vegetation restoration and the optimization of land use planning.
Asunto(s)
Lagos , Erosión del Suelo , Conservación de los Recursos Naturales , Monitoreo del AmbienteRESUMEN
BACKGROUND: Trophoblast dysfunction during pregnancy is fundamentally involved in preeclampsia. Several studies have revealed that human chorionic villous mesenchymal stem cells (CV-MSCs) could regulate trophoblasts function. RESULTS: To understand how human chorionic villous mesenchymal stem cells (CV-MSCs) regulate trophoblast function, we treated trophoblasts with CV-MSC supernatant under hypoxic conditions. Treatment markedly enhanced proliferation and invasion and augmented autophagy. Transcriptome and pathway analyses of trophoblasts before and after treatment revealed JAK2/STAT3 signalling as an upstream regulator. In addition, STAT3 mRNA and protein levels increased during CV-MSC treatment. Consistent with these findings, JAK2/STAT3 signalling inhibition reduced the autophagy, survival and invasion of trophoblasts, even in the presence of CV-MSCs, and blocking autophagy did not affect STAT3 activation in trophoblasts treated with CV-MSCs. Importantly, STAT3 overexpression increased autophagy levels in trophoblasts; thus, it positively regulated autophagy in hypoxic trophoblasts. Human placental explants also proved our findings by showing that STAT3 was activated and that LC3B-II levels were increased by CV-MSC treatment. CONCLUSION: In summary, our data suggest that CV-MSC-dependent JAK2/STAT3 signalling activation is a prerequisite for autophagy upregulation in trophoblasts.
RESUMEN
A clear quantification and spatial mapping between supply and demand of water provision service in relation to climate change and urban expansion can provide some guidance to water resources management. Nevertheless, so far, most researches ignored the dynamic changes and influences of supply-demand coupling correlations. In this study, water yield and water demand were quantified and mapped in the Xiangjiang River Basin (XRB) from 2000 to 2018 by using the Integrated Valuation of Ecosystem Services and Trade-offs (InVEST) and water-demand models, then the spatial distribution characteristics and their matching relationship were identified by using the univariate local autocorrelation analysis and the common logarithm of water supply-demand ratio (WSDR). With that, the contributions of climate and socio-economic factors to the above-mentioned changes were explored by using geographic detector. Results showed that the annual water yield increased by 20.20% in 2000-2015 and decreased by 33.92% in 2015-2018 affected by precipitation and land use changes; Changsha-Zhuzhou- Xiangtan urban agglomeration (CZX) and Southwest of Yongzhou were the high value areas of water yield (>338 m3/hm2). Due to the urban expansion, the water demand increased by 40.50% from 2000 to 2005 and decreased by 36.39% after 2005; From 2000 to 2018, high value areas of water demand (>53566 m3/hm2) mainly appeared in midstream and downstream with high urbanization level, dense population and developed industry. Under the joint action of precipitation (prep) and urban expansion, the overall state of supply and demand in the upper reaches was surplus, and more than 90% of the regions in midstream and downstream were at the middle and high level of supply shortage, especially in Hengyang and Chenzhou. Consequently, the increasing needs of human beings should be emphasized from the overall perspective of the basin, the growth rate of construction land and the necessary green infrastructure should be controlled reasonably and configured for achieving win-win goals of coordinating environmental protection and urban development.
Asunto(s)
Ecosistema , Agua , China , Conservación de los Recursos Naturales , Humanos , Ríos , Urbanización , Abastecimiento de AguaRESUMEN
Chlorophylls function in photosynthesis, and are critical to plant developmental processes and responses to environmental stimuli. Chlorophyll b is synthesized from chlorophyll a by chlorophyll a oxygenase (CAO). Here, we characterize a yellow-green leaf (ygl) mutant and identify the causal gene which encodes a chlorophyll a oxygenase in maize (ZmCAO1). A 51 bp Popin transposon insertion in ZmCAO1 strongly disrupts its transcription. Low enzyme activity of ZmCAO1 leads to reduced concentrations of chlorophyll a and chlorophyll b, resulting in the yellow-green leaf phenotype of the ygl mutant. The net photosynthetic rate, stomatal conductance, and transpiration rate are decreased in the ygl mutant, while concentrations of δ-aminolevulinic acid (ALA), porphobilinogen (PBG) and protochlorophyllide (Pchlide) are increased. In addition, a ZmCAO1 mutation results in down-regulation of key photosynthetic genes, limits photosynthetic assimilation, and reduces plant height, ear size, kernel weight, and grain yield. Furthermore, the zmcao1 mutant shows enhanced reactive oxygen species production leading to sensitivity to waterlogging. These results demonstrate the pleiotropy of ZmCAO1 function in photosynthesis, grain yield, and waterlogging tolerance in maize.
Asunto(s)
Inundaciones , Oxigenasas/genética , Proteínas de Plantas/genética , Zea mays , Clorofila , Clorofila A , Fotosíntesis , Hojas de la Planta , Semillas/crecimiento & desarrollo , Zea mays/genéticaRESUMEN
Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) can induce substantial cytotoxicity in tumor cells but rarely exert cytotoxic activity on non-transformed cells. In the present study, we therefore evaluated interactions between TRAIL and IER3 via co-immunoprecipitation and immunofluorescence analyses, leading us to determine that these two proteins were able to drive the apoptotic death of hepatocellular carcinoma (HCC) cells and to disrupt their proliferative and migratory abilities both in vitro and in vivo. From a mechanistic perspective, we determined that TRAIL and IER3 were capable of inhibiting Wnt/ß-catenin signaling. Together, these results indicate that TRAIL can control the pathogenesis of HCC at least in part via interacting with IER3 to inhibit Wnt/ß-catenin signaling, thus indicating that this TRAIL/IER3/ß-catenin axis may be a viable therapeutic target in HCC patients.
RESUMEN
Deoxythymidilate kinase (DTYMK) has been identified as a putative oncogene associated with the incidence of hepatocellular carcinoma (HCC), but the mechanisms whereby it regulates this cancer type remain uncertain. The present study was therefore designed to explore the role of DTYMK in HCC and to evaluate the underlying molecular mechanisms. MiRNAs associated with DTYMK expression levels in HCC were identified through analyses of both clinical samples and publically available gene expression datasets. We then assessed the putative functions of DTYMK and miR-148b-3p in this oncogenic context through studies of HCC cells and a murine xenograft model system. Correlation analyses and in vitro experiments led us to confirm DTYMK as a target of miR-148b-3p. In addition, we assessed dTTP levels associated with the DTYMK pathway in HCC cells to understand the functional implications of our experimental findings. We found that HCC tissues and cells exhibited marked DTYMK upregulation and miR-148b-3p downregulation, with the expression levels of DTYMK and miR-148b-3p being negatively correlated with one another. The impact of overexpressing DTYMK in tumor cells was partially reversed upon cellular transfection with miR-148b-3p mimics, providing conclusive evidence that DTMYK is a target of this miRNA. Importantly, DTYMK-related dTTP levels were also impacted by miR-148b-3p mimic transfection. DTYMK is a key regulator of HCC progression, and its expression is suppressed by miR-148b-3p, suggesting that this miR-148b-3p/DTYMK regulatory axis may be amenable to therapeutic targeting in patients with HCC.
RESUMEN
Rare classes are usually hidden in an imbalanced dataset with the majority of the data examples from major classes. Rare-class mining (RCM) aims at extracting all the data examples belonging to rare classes. Most of the existing approaches for RCM require a certain amount of labeled data examples as input. However, they are ineffective in practice since requesting label information from domain experts is time consuming and human-labor extensive. Thus, we investigate the unsupervised RCM problem, which to the best of our knowledge is the first such attempt. To this end, we propose an efficient algorithm called Fast-RCM for unsupervised RCM, which has an approximately linear time complexity with respect to data size and data dimensionality. Given an unlabeled dataset, Fast-RCM mines out the rare class by first building a rare tree for the input dataset and then extracting data examples of the rare classes based on this rare tree. Compared with the existing approaches which have quadric or even cubic time complexity, Fast-RCM is much faster and can be extended to large-scale datasets. The experimental evaluation on both synthetic and real-world datasets demonstrate that our algorithm can effectively and efficiently extract the rare classes from an unlabeled dataset under the unsupervised settings, and is approximately five times faster than that of the state-of-the-art methods.
RESUMEN
Kinases form the backbone of numerous cell signaling pathways, with their dysfunction similarly implicated in multiple pathologies. Further facilitated by their druggability, kinases are a major focus of therapeutic development efforts in diseases such as cancer, infectious disease and autoimmune disorders. While their importance is clear, the role or biological function of nearly one-third of kinases is largely unknown. Here, we describe a data resource, the Dark Kinase Knowledgebase (DKK; https://darkkinome.org), that is specifically focused on providing data and reagents for these understudied kinases to the broader research community. Supported through NIH's Illuminating the Druggable Genome (IDG) Program, the DKK is focused on data and knowledge generation for 162 poorly studied or 'dark' kinases. Types of data provided through the DKK include parallel reaction monitoring (PRM) peptides for quantitative proteomics, protein interactions, NanoBRET reagents, and kinase-specific compounds. Higher-level data is similarly being generated and consolidated such as tissue gene expression profiles and, longer-term, functional relationships derived through perturbation studies. Associated web tools that help investigators interrogate both internal and external data are also provided through the site. As an evolving resource, the DKK seeks to continually support and enhance knowledge on these potentially high-impact druggable targets.
Asunto(s)
Internet , Bases del Conocimiento , Fosfotransferasas/metabolismo , Regulación Enzimológica de la Expresión GénicaRESUMEN
Hepatocellular carcinoma (HCC) is a major cause of tumor associated deaths globally. Annually, the prevalence of HCC is increasing and the lack of early prognostic indicators manifests a dismal prognosis for HCC patients. A deep understanding of the molecular events that promote HCC progression are required for the design of new diagnostics and therapeutics. Dermatopontin (DPT) is an extracellular matrix protein that regulates the metastatic phenotypes of many cancers. However, the effects of DPT on HCC cell growth remain undefined. In this study, we demonstrate that the exogenous expression of DPT inhibits HCC cell growth both in vitro and in vivo. Furthermore, we show that DPT regulates CXXC4, which in turn targets c-Myc, EZH2, SOX2 and ß-catenin, through its ability to impact Wnt signaling pathway. These data suggest that DPT regulates CXXC4, c-Myc, EZH2, SOX2 and ß-catenin, through Wnt signaling to repress HCC proliferation. This highlights DPT as promising target for future HCC diagnostics and therapeutic targets.
RESUMEN
Hepatocellular carcinoma (HCC) is a prevalent and highly aggressive cancer. Long non-coding RNAs (lncRNAs) are recognized as potential molecular targets for HCC and are currently under increased research focus. Here, we investigate the regulatory processes underlying the axis of the lncRNA taurine upregulated gene 1 (TUG1), Upstream Transcription Factor 1 (USF1), and reactive oxygen species modulator 1 (ROMO1) in the propagation and metastasis of HCC cells. Distribution of lncRNA TUG1 was found to be prominent in HCC cell cytoplasm and nuclei. LncRNA TUG1 conscripted the USF1 transcription factor to enhance the promoter function of ROMO1. Enlisting the USF1 transcription factor to increase ROMO1 expression following upregulation of TUG1 lncRNA enhanced HCC Huh7 cell proliferation, motility, and metastasis. Rapid tumor proliferation in nude mice provided in vivo verification. The importance of the lncRNA TUG1/USF1/ROMO1 complex as a target for HCC therapy is a key result of this investigation which is exemplified by its role in regulating the proliferation, motility, and metastasis of HCC cells.
RESUMEN
Krüppel-like factor 4 (Human Protein: KLF4; Human Gene: Klf4; Murine Protein: KLF4; Murine Gene: Klf4) is a zinc finger-containing transcription factor with diverse regulatory functions. Mouse embryonic fibroblasts (MEFs) lacking Klf4 exhibit genomic instability, increased reactive oxygen species (ROS), and decreased autophagy. Elevated ROS is linked to impairments in mitochondrial damage recovery responses and is often tied to disruption in mitochondrial-targeted autophagy known as mitophagy. In this study, we sought to identify a mechanistic connection between KLF4 and mitophagy. Using flow cytometry, we found that Klf4-null MEFs have diminished ability to recover mitochondrial health and regulate ROS levels after mitochondrial damage. Confocal microscopy indicated decreased localization of autophagy protein LC3 to mitochondria following mitochondrial damage in Klf4-null cells, suggesting decreased mitophagy. Western blotting and RT-PCR revealed decreased mRNA and protein expression of the mitophagy-associated protein Bnip3 and antioxidant protein GSTα4 in Klf4-null cells, providing a rationale for their impaired mitophagy and ROS accumulation. Inducing Bnip3 expression in these cells recovered mitophagy but did not decrease ROS accumulation. Our findings suggest that in Klf4-null cells, decreased Bnip3 expression impairs mitophagy and is associated with increased mitochondrial ROS production after mitochondrial damage, providing a rationale for their genomic instability and supports a tumor suppressive role for KLF4 in certain tumors as previously observed.
Asunto(s)
Factores de Transcripción de Tipo Kruppel/metabolismo , Mitocondrias/metabolismo , Mitofagia , Células 3T3 , Animales , Células Cultivadas , Factor 4 Similar a Kruppel , Factores de Transcripción de Tipo Kruppel/deficiencia , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Background: Hepatocellular carcinoma (HCC) is a prominent cancer type, with long non-coding RNAs (lncRNAs) being known to be relevant to its progression. We therefore investigated how a particular lncRNA known as the metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) was associated with HCC. Methods: Quantitative reverse transcriptase PCR (qPCR) was used to assess expression of MALAT1, Forkhead Box M1 (FOXM1) and miR-125a-3p in HCC tissue samples. How MALAT1 regulates HCC proliferation and metastasis was assessed through appropriate assays. FOXM1 was identified as a miR-125a-3p target using luciferase assays, and how MALAT1/miR-125a-3p alter FOXM1 expression was explored via qPCR and Western blotting. Results: HCC tissues exhibited MALAT1 upregulation. miR-125a-3p targeted FOXM1 and could be regulated by MALAT1. MALAT1 knockdown disrupted proliferation and invasion, whereas miR-125a-3p knockdown partially reversed this phenotype. Conclusions: These results indicate that MALAT1 modulates FOXM1 expression via being a miR-125a-3p sponge, thus promoting HCC progression.
RESUMEN
The target profiles of many drugs are established early in their development and are not systematically revisited at the time of FDA approval. Thus, it is often unclear whether therapeutics with the same nominal targets but different chemical structures are functionally equivalent. In this paper we use five different phenotypic and biochemical assays to compare approved inhibitors of cyclin-dependent kinases 4/6-collectively regarded as breakthroughs in the treatment of hormone receptor-positive breast cancer. We find that transcriptional, proteomic, and phenotypic changes induced by palbociclib, ribociclib, and abemaciclib differ significantly; abemaciclib in particular has advantageous activities partially overlapping those of alvocidib, an older polyselective CDK inhibitor. In cells and mice, abemaciclib inhibits kinases other than CDK4/6 including CDK2/cyclin A/E-implicated in resistance to CDK4/6 inhibition-and CDK1/cyclin B. The multifaceted experimental and computational approaches described here therefore uncover underappreciated differences in CDK4/6 inhibitor activities with potential importance in treating human patients.
Asunto(s)
Quinasa 4 Dependiente de la Ciclina/antagonistas & inhibidores , Quinasa 6 Dependiente de la Ciclina/antagonistas & inhibidores , Polifarmacología , Inhibidores de Proteínas Quinasas/farmacología , Animales , Quinasa 4 Dependiente de la Ciclina/metabolismo , Quinasa 6 Dependiente de la Ciclina/metabolismo , Femenino , Humanos , Ratones , Ratones Desnudos , Inhibidores de Proteínas Quinasas/química , Estados Unidos , United States Food and Drug AdministrationRESUMEN
BACKGROUND: Autophagy is a major cellular process by which cytoplasmic components such as damaged organelles and misfolded proteins are recycled. Although low levels of autophagy occur in cells under basal conditions, certain cellular stresses including nutrient depletion, DNA damage, and oxidative stress are known to robustly induce autophagy. Krüppel-like factor 4 (KLF4) is a zinc-finger transcription factor activated during oxidative stress to maintain genomic stability. Both autophagy and KLF4 play important roles in response to stress and function in tumor suppression. METHODS: To explore the role of KLF4 on autophagy in mouse embryonic fibroblasts (MEFs), we compared wild-type with Klf4 deficient cells. To determine the levels of autophagy, we starved MEFs for different times with Earle's balanced salts solution (EBSS). Rapamycin was used to manipulate mTOR activity and autophagy. The percentage of cells with γ-H2AX foci, a marker for DNA damage, and punctate pattern of GFP-LC3 were counted by confocal microscopy. The effects of the drug treatments, Klf4 overexpression, or Klf4 transient silencing on autophagy were analyzed using Western blot. Trypan Blue assay and flow cytometry were used to study cell viability and apoptosis, respectively. qPCR was also used to assay basal and the effects of Klf4 overexpression on Atg7 expression levels. RESULTS: Here our data suggested that Klf4 (-/-) MEFs exhibited impaired autophagy, which sensitized them to cell death under nutrient deprivation. Secondly, DNA damage in Klf4-null MEFs increased after treatment with EBSS and was correlated with increased apoptosis. Thirdly, we found that Klf4 (-/-) MEFs showed hyperactive mTOR activity. Furthermore, we demonstrated that rapamycin reduced the increased level of mTOR in Klf4 (-/-) MEFs, but did not restore the level of autophagy. Finally, re-expression of Klf4 in Klf4 deficient MEFs resulted in increased levels of LC3II, a marker for autophagy, and Atg7 expression level when compared to GFP-control transfected Klf4 (-/-) MEFs. CONCLUSION: Taken together, our results strongly suggest that KLF4 plays a critical role in the regulation of autophagy and suppression of mTOR activity. In addition, we showed that rapamycin decreased the level of mTOR in Klf4 (-/-) MEFs, but did not restore autophagy. This suggests that KLF4 regulates autophagy through both mTOR-dependent and independent mechanisms. Furthermore, for the first time, our findings provide novel insights into the mechanism by which KLF4 perhaps prevents DNA damage and apoptosis through activation of autophagy.