Dissection Of Mechanisms Research Articles

GATA-type transcription factor MrNsdD regulates dimorphic transition, conidiation, virulence and microsclerotium formation in the entomopathogenic fungus Metarhizium rileyi.

SummaryThe GATA‐type sexual development transcription factor NsdD has been implicated in virulence, secondary metabolism and asexual development in filamentous fungi. However, little is known about its function in the yeast‐to‐hypha transition and in microsclerotium formation. In the current study, the orthologous NsdD gene MrNsdD in the entomopathogenic fungus Metarhizium rileyi was characterized. Transcriptional analysis indicated that MrNsdD was involved in yeast‐to‐hypha transition, conidiation and microsclerotium formation. After targeted deletion of MrNsdD, dimorphic transition, conidiation, fungal virulence and microsclerotium formation were all impaired. Compared with the wild‐type strain, the ΔMrNsdD mutants were hypersensitive to thermal stress. Furthermore, transcriptome sequencing analysis revealed that MrNsdD regulated a distinct signalling pathway in M. rileyi during the yeast‐to‐hypha transition or microsclerotium formation, but exhibited overlapping regulation of genes during the two distinct developmental stages. Taken together, characterization of the MrNsdD targets in this study will aid in the dissection of the molecular mechanisms of dimorphic transition and microsclerotium development.

Transcript profiling provides insights into molecular processes during shoot elongation in temperature-sensitive peach (Prunus persica)

Plant growth caused by ambient temperature is thought to be regulated by a complex transcriptional network. A temperature-sensitive peach (Prunus persica) was used to explore the mechanisms behind shoot internode elongation at elevated temperatures. There was a significantly positive correlation between the length of the terminal internode (TIL) and the maximum temperature three days prior to the measuring day. Four critical growth stages (initial period and initial elongation period at lower temperature, rapid growth period and stable growth period at higher temperature) were selected for comparative RNA-seq analysis. About 6.64G clean bases were obtained for each library, and 88.27% of the data were mapped to the reference genome. Differentially expressed gene (DEG) analysis among the three pairwise comparisons resulted in the detection of several genes related to the shoot elongation in temperature-sensitive peach. HSFAs were up-regulated in response to the elevated temperature, while the up-regulated expression of HSPs might influence hormone signaling pathways. Most of DEGs involved in auxin, abscisic acid and jasmonic acid were up-regulated, while some involved in cytokinin and brassinosteroid were down-regulated. Genes related to ethylene, salicylic acid and circadian rhythm were also differentially expressed. Genes related to aquaporins, expansins, pectinesterases and endoglucanase were up-regulated, which would promote cell elongation. These results lay a foundation for further dissection of the regulatory mechanisms underlying shoot elongation at elevated temperatures.

Nuclear-Translocation of ACLY Induced by Obesity Related Factors Enhances Pyrimidine Metabolism Through Regulating Histone Acetylation in Endometrial Cancer

Background: Endometrial cancer (EC) is a common female reproductive system cancer, and obesity is a major risk factor for the disease. ATP-citrate lyase (ACLY), a vital enzyme linking both glycolytic and lipid metabolism, plays important roles in cancer progression. However, there is little evidence of ACLY involvement in EC development. Methods: The ACLY expression in EC was evaluated by using TCGA and GEO databases and verified it using clinical samples by IHC and RT-PCR methods. The functions of ACLY was determined by both the vitro and vivo studies. Gene expression array, co-immunoprecipitation, TUNEL assay, EdU assay, chromatin immunoprecipitation and luciferase reporter assay were applied for mechanism dissection. Findings: ACLY was significantly upregulated in EC patients, particularly in the nucleus. Upregulated ACLY promoted EC cell proliferation and colony formation, and attenuated apoptosis. Mechanistically, cotreatment with obesity-related factors (estradiol, insulin and leptin) promoted nuclear translocation of ACLY through Akt-mediated phosphorylation of ACLY at Ser455 . Nuclear-localized ACLY increased histone acetylation levels, thus resulting in upregulation of pyrimidine metabolism genes, such as DHODH. Moreover, STAT3 altered the ACLY expression at the transcriptional level via directly binding to its promoter region. Interpretation: ACLY nuclear-translocation induced by obesity-related factors promotes EC progression by modulating histone acetylation levels, The ACLY axis could be served as potential therapeutic targets for the treatment of EC. Funding Statement: This research was supported by The National Natural Science Foundation of China (Grant number: 81974406) and The Sailing Program Foundation of Hunan Cancer Hospital. Declaration of Interests: The authors declare that they have no competing interests with the contents of this article. Ethics Approval Statement: The study was approved by the Research Ethics Committee of the Sixth People's Hospital, and informed consent was provided by all patients before this study.

Myelin-specific CD8 T cells require IFNγ responsiveness to optimally suppress CNS autoimmunity

Abstract Investigating the complex cellular interplay controlling immunopathogenic and immunoregulatory responses is critical for understanding multiple sclerosis (MS) and for developing successful immunotherapies. Our group has demonstrated that central nervous system (CNS) myelin-specific CD8 T cells unexpectedly harbor striking regulatory capacity in both mouse and human. In particular, PLP178–191-specific CD8 T cells (PLP-CD8) robustly suppress the MS mouse model experimental autoimmune encephalomyelitis (EAE). We have recently shown this depends on PLP-CD8 elaborating IFNγ and perforin in a coordinated suppression program over time. However, the cellular target and downstream effects of CD8 T cell-elaborated IFNγ remains poorly understood. Here, we show that while WT PLP-CD8 were robustly suppressive in IFNγR-deficient mice, IFNγR-deficient PLP-CD8 exhibited defective suppression in WT mice. Further, compared to WT counterparts, IFNγR-deficient PLP-CD8 were defective in suppressing disease in IFNγ-deficient recipients, a scenario in which the only IFNγ WT PLP-CD8 “see” is their own. These data indicate that myelin-specific CD8 T cells must sense their own IFNγ in an autocrine/paracrine manner to optimally perform autoregulatory function in vivo. Further mechanistic dissection showed that IFNγR-deficient PLP-CD8 exhibited altered degranulation/granzyme profiles, altered migration in recipients, and potential deficits in killing capacity. Collectively, this work suggests that myelin-specific CD8 T cells require IFNγ-responsiveness to optimally regulate CNS autoimmunity and may help elucidate future adoptive immunotherapeutic approaches in MS.

A Genomic Toolkit for the Mechanistic Dissection of Intractable Human Gut Bacteria

Despite the remarkable microbial diversity found within humans, our ability to link genes to phenotypes is based upon a handful of model microorganisms. We report a comparative genomics platform for Eggerthella lenta and other Coriobacteriia, a neglected taxon broadly relevant to human health and disease. We uncover extensive genetic and metabolic diversity and validate a tool for mapping phenotypes to genes and sequence variants. We also present a tool for the quantification of strains from metagenomic sequencing data, enabling the identification of genes that predict bacterial fitness. Competitive growth is reproducible under laboratory conditions and attributable to intrinsic growth rates and resource utilization. Unique signatures of invivo competition ingnotobiotic mice include an adhesin enriched in poor colonizers. Together, these computational and experimental resources represent a strong foundation for the continued mechanistic dissection of the Coriobacteriia and a template that can be applied to study other genetically intractable taxa.

Identification of QTLs associated with curd architecture in cauliflower

BackgroundCurd architecture is one of the most important characters determining the curd morphology of cauliflower. However, the genetic mechanism dissection of this complex trait at molecular level is lacking. Genes/QTLs responsible for the morphological differences between present-day loose-curd and compact-curd cauliflower haven’t been well revealed.ResultsHerein, by using a common compact-curd parent and two loose-curd parents, we developed two double haploid (DH) populations including 122 and 79 lines, respectively. For each population, we decomposed the curd architecture concept into four parameters (basal diameter, stalk length, stalk angle and curd solidity), and collected corresponding phenotypic data for each parameter across two environments. The Kosambi function and composite interval mapping algorithm were conducted to construct the linkage map and analyze the QTLs associated with curd architecture parameters. A total of 20 QTLs were detected with the minimum likelihood of odd (LOD) values ranging from 2.61 to 8.38 and the percentage of the phenotypic variance explained by each QTL (PVE) varying between 7.69 and 25.10%. Of these, two QTLs controlling stalk length (qSL.C6–1, qSL.C6–2) and two QTLs controlling curd solidity (qCS.C6–1 and qCS.C6–2) were steadily expressed in both environments. Further, qSL.C6–1, qSL.C6–2, qCS.C6–1 and qCS.C6–4 fell into the same chromosomal region of the reference genome, indicating that these loci are involved in pleiotropic effects or are tightly linked.ConclusionThe current study identified a series of QTLs associated with curd architecture parameters, which might contribute essentially to the formation of present-day loose-curd cauliflower that is widely cultivated in China. These results may pave the way for intensive deciphering the molecular mechanisms of curd development and for marker-assisted selection of curd morphology in cauliflower breeding.

Pathogenesis of peritumoral hyperexcitability in an immunocompetent CRISPR-based glioblastoma model.

Seizures often herald the clinical appearance of gliomas or appear at later stages. Dissecting their precise evolution and cellular pathogenesis in brain malignancies could inform the development of staged therapies for these highly pharmaco-resistant epilepsies. Studies in immunodeficient xenograft models have identified local interneuron loss and excess glial glutamate release as chief contributors to network disinhibition, but how hyperexcitability in the peritumoral microenvironment evolves in an immunocompetent brain is unclear. We generated gliomas in WT mice via in utero deletion of key tumor suppressor genes and serially monitored cortical epileptogenesis during tumor infiltration with in vivo electrophysiology and GCAMP7 calcium imaging, revealing a reproducible progression from hyperexcitability to convulsive seizures. Long before seizures, coincident with loss of inhibitory cells and their protective scaffolding, gain of glial glutamate antiporter xCT expression, and reactive astrocytosis, we detected local Iba1+ microglial inflammation that intensified and later extended far beyond tumor boundaries. Hitherto unrecognized episodes of cortical spreading depolarization that arose frequently from the peritumoral region may provide a mechanism for transient neurological deficits. Early blockade of glial xCT activity inhibited later seizures, and genomic reduction of host brain excitability by deleting MapT suppressed molecular markers of epileptogenesis and seizures. Our studies confirmed xenograft tumor-driven pathobiology and revealed early and late components of tumor-related epileptogenesis in a genetically tractable, immunocompetent mouse model of glioma, allowing the complex dissection of tumor versus host pathogenic seizure mechanisms.

Homeostatic Plasticity Shapes the Retinal Response to Photoreceptor Degeneration.

Homeostatic plasticity stabilizes input and activity levels during neural development, but whether it can restore connectivity and preserve circuit function during neurodegeneration is unknown. Photoreceptor degeneration is the most common cause of blindness in the industrialized world. Visual deficits are dominated by cone loss, which progresses slowly, leaving a window during which rewiring of second-order neurons (i.e., bipolar cells) could preserve function. Here we establish a transgenic model to induce cone degeneration with precise control and analyze bipolar cell responses and their effects on vision through anatomical reconstructions, invivo electrophysiology, and behavioral assays. In young retinas, we find that three bipolar cell types precisely restore input synapse numbers when 50% of cones degenerate but one does not. Of the three bipolar cell types that rewire, two contact new cones within stable dendritic territories, whereas one expands its dendrite arbors to reach new partners. In mature retinas, only one of four bipolar cell types rewires homeostatically. This steep decline in homeostatic plasticity is accompanied by reduced light responses of bipolar cells and deficits in visual behaviors. By contrast, light responses and behavioral performance are preserved when cones degenerate in young mice. Our results reveal unexpected cell type specificity and a steep maturational decline of homeostatic plasticity. The effect of homeostatic plasticity on functional outcomes identify it as a promising therapeutic target for retinal and other neurodegenerative diseases.

Transcriptional Regulation of the Proteasome: Implications for Cancer Therapy

While proteasome inhibitors are being effectively used in the clinic to treat multiple myeloma and mantle cell lymphoma, the reasons for why this strategy is not generally applicable to other tumor types is poorly understood. A detailed characterization of the cellular events triggered in response to these agents could offer vital clues to target the protein degradation pathway(s) more effectively in a broad range of tumors.Our previous studies have established the transcription factor Nrf1 as a key player in the stress‐response pathway invoked by inhibition of cellular proteasome. Nrf1, by its ability to induce de novo synthesis of proteasome subunit genes in response to proteasome inhibitors, promotes the recovery of proteasome activity, thus mitigating proteotoxic stress and enhancing cellular survival. From a therapeutic stand‐point, this suggests that disabling the Nrf1‐mediated proteasome recovery pathway could potentiate the action of proteasome inhibitor treatments. However, effective means to curb the Nrf1 pathway remain scarce. Here, using an in silico chemical biology approach, we uncovered a class of therapeutic agents that can effectively inhibit the Nrf1 pathway. Mechanistic dissection of this phenomenon provided further novel insight into the way by which Nrf1 maintains its transcriptional activity. Using triple negative breast cancer (TNBC) cell lines and patient‐derived xenograft (PDX) models, we further validated our hypothesis that attenuating the Nrf1 pathway could increase the efficacy of proteasome inhibitor therapeutics. Thus, our study sheds light on the unique biology of Nrf1 along with validating this pathway as a therapeutic target in TNBC. This could serve as a motivation for similar future efforts where Nrf1 pathway inhibitors can be tested as adjuvants for proteasome inhibitor treatments in other cancer types.Support or Funding InformationNIH grant R01GM132396

Mechanistic Dissection of Spatially Encoded GPCR Signaling Through High‐throughput CRISPR Screening

Our recent work has overturned the prevailing textbook model for GPCR signaling, which held that GPCR activation at the plasma membrane is responsible for all downstream consequences of signaling. We have demonstrated that receptor/second messenger signaling on internal compartments but not plasma membrane drives all receptor‐dependent transcriptional responses, and the site of receptor activation can dictate how cells “interpret” different ligands, including medically relevant drugs. This phenomenon of functional specialization of signaling based on receptor localization has been gaining significant support in the last few years from us and other groups, yet not much is known about its underlying mechanisms and functional consequences. To begin to address these open questions, we set up a functional genomic screen aimed at identifying factors regulating ß2‐adrenergic receptor (ß2‐AR)‐dependent transcription‐ a location‐biased GPCR signaling response. We combined pooled CRISPR interference libraries targeting all annotated transcripts in the human genome and a novel fluorescent transcriptional reporter for GPCR/cAMP signaling that we have developed with fluorescence‐activated cell sorting and deep sequencing. Through this comprehensive screen, we have identified both known and novel mediators of the GPRC pathway. We are currently defining the biochemical mechanisms and specificity of the new regulators and their contributions to location‐biased receptor signaling, and have intriguing data linking a highly‐penetrant genetic risk factor for psychosis to these pathways.Support or Funding InformationNational Institute of Mental Health [Grant MH10963301]

PD59-03 HYPOXIA INDUCE NEUROENDOCRINE DIFFERENTIATION OF PROSTATE CANCER VIA ANDROGEN RECEPTOR RELATED SIGNALS

You have accessJournal of UrologyProstate Cancer: Basic Research & Pathophysiology III (PD59)1 Apr 2020PD59-03 HYPOXIA INDUCE NEUROENDOCRINE DIFFERENTIATION OF PROSTATE CANCER VIA ANDROGEN RECEPTOR RELATED SIGNALS Yun Cui*, Yin Sun, Yang Yang, Shuai Hu, Yinong Niu, Xiaodong Zhang, Shuyuan Yeh, Chawnshang Chang, and Jie Jin Yun Cui*Yun Cui* More articles by this author , Yin SunYin Sun More articles by this author , Yang YangYang Yang More articles by this author , Shuai HuShuai Hu More articles by this author , Yinong NiuYinong Niu More articles by this author , Xiaodong ZhangXiaodong Zhang More articles by this author , Shuyuan YehShuyuan Yeh More articles by this author , Chawnshang ChangChawnshang Chang More articles by this author , and Jie JinJie Jin More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000000969.03AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Development of the neuroendocrine differentiation (NED) in prostate cancer (PCa) has been linked to the failure of androgen deprivation therapy. And it was also regarded as the main origin of neuroendocrine PCa, which is a terminal lethal cancer with little effective therapies. This study aimed to study the role of hypoxia, especially hypoxia induced androgen receptor (AR) signals, in the NED of PCa and explore the potential way to restrain NED development. METHODS: Hypoxia was achieved by maintaining the cells at 1% O2, 5% CO2 and 94% N2 in a hypoxic chamber with oxygen sensor controls. Immunohistochemistry, immunoprecipitation assay, chromatin immunoprecipitation assay, CRISPR-CAS9 mediated gene transactivation, orthotropic xenograft, cell culture, quantitative RT-PCR, immunoblotting, bioinformatics analysis, lentivirus packaging and luciferase reporter assay were used to study hypoxia induced NED of PCa in vivo and in vitro. RESULTS: PCa performed NED under hypoxia condition with morphology change and neuroendocrine-related marker increasing. In vitro studies indicated that hypoxia induce NED via suppressing the AR signals. And mechanism dissection demonstrated hypoxia could suppress AR protein via AKT/MDM2 mediated ubiquitination of the AR protein. AR could regulate miRNA-449c-5p which suppress ASCL1 expression at the transcriptional level. Importantly, human clinical data survey also revealed a positive correlation between hypoxia and NED. In vivo orthotropic xenograft indicated miR-449c-5p could suppress NED development in PCa. CONCLUSIONS: Results from human clinical data survey and preclinical studies conclude that hypoxia may increase NED via altering the AKT/MDM2/AR/miRNA-449c-5p/ASCL1 signals in PCa, and targeting these newly identified AR-mediated signals with miRNA-449c-5p may help us to better suppress PCa progression via reducing the development of NED. Source of Funding: Beijing Natural Science Foundation (No. 7184214) © 2020 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 203Issue Supplement 4April 2020Page: e1205-e1206 Advertisement Copyright & Permissions© 2020 by American Urological Association Education and Research, Inc.MetricsAuthor Information Yun Cui* More articles by this author Yin Sun More articles by this author Yang Yang More articles by this author Shuai Hu More articles by this author Yinong Niu More articles by this author Xiaodong Zhang More articles by this author Shuyuan Yeh More articles by this author Chawnshang Chang More articles by this author Jie Jin More articles by this author Expand All Advertisement PDF downloadLoading ...

MP16-09 THE ANDROGEN RECEPTOR REPRESSES PROSTATE CANCER INVASION INDUCED BY ONCOMETABOLITE R-2-HYDROXYGLUTARATE THROUGH CIRC51217/MIR-646/TGFβ1/P-SMAD2/3 SIGNALING

You have accessJournal of UrologyProstate Cancer: Basic Research & Pathophysiology I (MP16)1 Apr 2020MP16-09 THE ANDROGEN RECEPTOR REPRESSES PROSTATE CANCER INVASION INDUCED BY ONCOMETABOLITE R-2-HYDROXYGLUTARATE THROUGH CIRC51217/MIR-646/TGFβ1/P-SMAD2/3 SIGNALING Hua Xu* and Ding-wei Ye Shanghai Hua Xu*Hua Xu* More articles by this author and Ding-wei Ye ShanghaiDing-wei Ye Shanghai More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000000841.09AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: IDH1 (Isocitrate dehydrogenase 1) mutation occurring at codon 132 (R132) in prostate cancer (PCa) is considered as a classifier for a subgroup PCa with accumulation of oncometabolite R-2HG (R-2-hydroxyglutarate). Besides its association with early onset of PCa, little is known about the biological effect of IDH1 mutation and R-2HG in PCa. In this study, we focus on the biological effect of IDH1 R132H mutation, which is the most frequent hotspot mutation, and oncometabolite R-2HG in PCa cell invasion. METHODS: A retrospective cohort of 1012 PCa patients in Fudan University Shanghai Cancer Center were included for next-generation sequencing. IDH1 R132H mutation was discovered as a hot spot mutation for further research. The in vitro study was designed to explore the biological function of IDH1 R132H mutation as well as R-2HG in PCa. Transwell assay was applied to test the cell invasiveness of PC3 and C4-2 PCa cell lines. CircRNA and microRNA candidates were screened based on database and manipulation of their expression was used test their function in the signaling pathway. RESULTS: Based on the next-generation sequencing data from a retrospective cohort of 1012 PCa patients, IDH1 R132H was found to be on the most frequent mutations. We discovered 9 PCa patients (0.9%) carried IDH1 R132H mutation. In the in vitro study, we found that over-expressing mutant IDH1 R132H as well as R-2HG promotes cell invasion in PCa with negative or low expression of androgen receptor (AR). Mechanism dissection revealed that R-2HG can induce circ51217 generation through increasing N6-methyladenosine (m6A) level of TGFβ1 (Transforming growth factor-β 1) mRNA, and in turn sponge miR-646 to further increase TGFβ1 protein expression and TGFβ1/p-Smad2/3 signaling. AR can suppress R-2HG effect at multiple levels, namely repressing TGFβ1 transcription and inhibiting circ51217generation via regulating ADAR2 (Double-stranded RNA-specific editase 1) expression. Preclinical study with in vivo xenograft mouse model also shows that PCa with IDH1 R132H mutation is more invasive in experimental metastasis. CONCLUSIONS: This study demonstrates that IDH1 R132H mutation as well as oncometabolite R-2HG increases cell invasiveness of PCa with negative or low expression of AR through circ51217/miR-646/TGFβ1/p-Smad2/3 signaling. AR can negate R-2HG effect via suppressing transcription of TGFβ1 and generation of circ51217. Source of Funding: National Natural Science Foundation of China (No. 81472377 and 81802570) © 2020 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 203Issue Supplement 4April 2020Page: e217-e217 Advertisement Copyright & Permissions© 2020 by American Urological Association Education and Research, Inc.MetricsAuthor Information Hua Xu* More articles by this author Ding-wei Ye Shanghai More articles by this author Expand All Advertisement PDF downloadLoading ...

PD33-11 CD3+ T CELLS SUPPRESS ANDROGEN RECEPTOR IN BPH VIA IL-1β/MIR-15B-5P SIGNALING TO AFFECT 5 ALPHA REDUCTASE INHIBITOR TREATMENT

You have accessJournal of UrologyBenign Prostatic Hyperplasia: Medical & Non-surgical Therapy (PD33)1 Apr 2020PD33-11 CD3+ T CELLS SUPPRESS ANDROGEN RECEPTOR IN BPH VIA IL-1β/MIR-15B-5P SIGNALING TO AFFECT 5 ALPHA REDUCTASE INHIBITOR TREATMENT Hualin Song*, shuai hu, and Jie Jin Hualin Song*Hualin Song* More articles by this author , shuai hushuai hu More articles by this author , and Jie JinJie Jin More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000000904.011AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: 5 alpha reductase inhibitor (5ARI) is widely used in the treatment of benign prostatic hyperplasia (BPH), and has achieved good therapeutic effects, but 30%-40% of patients still have poor therapeutic effect, and subsequent surgery is needed. We found that patients with poor 5ARI treatment had more infiltration of CD3+ T cells than patients with good treatment. At the same time, we found that the expression of androgen receptor (AR) in the infiltrated area of CD3+ T cells in human BPH specimen epithelial cells was reduced. However, the regulatory relationship between these two events remains unclear. The aim of this study was to investigate whether CD3+ T cells can regulate AR expression and whether it can affect the therapeutic effects of 5ARI. METHODS: Paraffin-embedded tissues obtained from TURP were used for expression of CD3 and AR protein by IHC. RWPE-1, BPH-1-AR cells (transfect AR into BPH-1 cells) and PWR-1E cells were co-cultured with/without CD3+ T cells-Molt-3 and HH cells. AR expression in BPH epithelial cells after co-cultured were detected by Western blot. Cytokines in co-cultured supernatant were detected by cytokines array and qPCR. The high-throughput sequencing was used to detect miRNAs expression in RWPE-1 and BPH-1-AR cells after co-cultured. RESULTS: Patients with poor 5ARI treatment had more infiltration of CD3+ T cells than patients with good treatment. Expression of AR in epithelial cells was reduced in the area of CD3+ T cells infiltration in clinical samples. AR expression was also significantly reduced in BPH epithelial cells after co-cultured with CD3+ T cells. IL-1β was increased in co-cultured supernatant and it was secreted by CD3+T cells. Meanwhile, miRNA-15b-5p was also significantly increased in BPH epithelial cells after co-cultured. Mechanism dissection revealed that infiltrating CD3+ T cells secreted IL-1β to up-regulate miRNA-15b-5p in BPH epithelial cells to suppress AR. Targeting these newly identified signals via IL-1β neutralizing antibody or miRNA-15b-5p inhibitor both led to partially reverse the AR reduction. CONCLUSIONS: This study was concluded that infiltrating CD3+ T cells could suppress AR to in BPH via modulation of IL-1β/miR-15b-5p signaling. The AR level that has been reduced makes the effect of 5ARI no longer obvious, which is one of the reasons for patients with poor 5ARI treatment. Targeting these newly discovered signals and 5ARI combination may improve treatment outcomes to better treat patients with BPH. Source of Funding: None. © 2020 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 203Issue Supplement 4April 2020Page: e709-e710 Advertisement Copyright & Permissions© 2020 by American Urological Association Education and Research, Inc.MetricsAuthor Information Hualin Song* More articles by this author shuai hu More articles by this author Jie Jin More articles by this author Expand All Advertisement PDF downloadLoading ...

MP79-14 LNCRNA-P21 ALTERS THE ANTIANDROGEN ENZALUTAMIDE-INDUCED PROSTATE CANCER NEUROENDOCRINE DIFFERENTIATION VIA MODULATING THE EZH2/STAT3 SIGNALING

You have accessJournal of UrologyProstate Cancer: Advanced (including Drug Therapy) IV (MP79)1 Apr 2020MP79-14 LNCRNA-P21 ALTERS THE ANTIANDROGEN ENZALUTAMIDE-INDUCED PROSTATE CANCER NEUROENDOCRINE DIFFERENTIATION VIA MODULATING THE EZH2/STAT3 SIGNALING Jie Luo*, Jialin Meng, Edward M. Messing, Jean V. Joseph, Shuyuan Yeh, and Chawnshang Chang Jie Luo*Jie Luo* More articles by this author , Jialin MengJialin Meng More articles by this author , Edward M. MessingEdward M. Messing More articles by this author , Jean V. JosephJean V. Joseph More articles by this author , Shuyuan YehShuyuan Yeh More articles by this author , and Chawnshang ChangChawnshang Chang More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000000971.014AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Prostate cancer (PCa) is the most prevalent male cancer in the U.S, and the current standard androgen deprivation therapy (ADT) with the FDA-approved antiandrogen Enzalutamide (Enz, also known as MDV3100) may effectively suppress the castration resistant PCa (CRPC) and extend CRPC patients’ survival an extra 4.8 months. However, ADT with Enz treatment (ADT-Enz) may also induce some unwanted adverse effects including development of Enz-resistance and increasing the PCa cell invasion as demonstrated in several preclinical models, as well as promoting the neuroendocrine (NE) differentiation (NED). METHODS: The viabilities of cells were determined by MTT (Sigma) assays and WST-1 assays (Cayman Chemical). The wildtype (wt) or mutant (mut) lncRNA-p21 promoters were constructed into PGL3-basic plasmid (Promega) to do the luciferase assays and luciferase activity was measured by Dual-Luciferase Assay reagent (Promega). Protein-DNA complexes were detected by Chromatin immunoprecipitation (ChIP) assays. In vivo mouse models were generated by PDX implantation and treated with different compound treatments. RESULTS: Results from cell lines and human clinical sample surveys revealed that lncRNA-p21 expression is up-regulated in NE PCa and Enz treatment could increase the lncRNA-p21 to induce the NED. Mechanism dissection revealed that Enz could promote the lncRNA-p21 transcription viaaltering the androgen receptor (AR) binding to different androgen-response-elements, which switch the EZH2 function from histone-methyltransferase to non-histone methyltransferase, consequently methylating the STAT3 to promote the NED. Preclinical studies using the PDX mouse model proved that EZH2 inhibitor could block the Enz-induced NED. CONCLUSIONS: In summary, our results suggest how Enz treatment can promote NED, and provide some potential therapies of using EZH2 inhibitors to target the AR/lncRNA-p21/EZH2/STAT3 axis to reduce the Enz-induced unwanted adverse effects of promoting the NED. Source of Funding: This work was supported by NIH grants (CA122840 and CA256700), and Taiwan Department of Health Clinical Trial and Research Center of Excellence (DOH102-TD-B-111-004) to China Medical University, Taiwan. © 2020 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 203Issue Supplement 4April 2020Page: e1221-e1221 Advertisement Copyright & Permissions© 2020 by American Urological Association Education and Research, Inc.MetricsAuthor Information Jie Luo* More articles by this author Jialin Meng More articles by this author Edward M. Messing More articles by this author Jean V. Joseph More articles by this author Shuyuan Yeh More articles by this author Chawnshang Chang More articles by this author Expand All Advertisement PDF downloadLoading ...

Vesiculation of Red Blood Cells in the Blood Bank: A Multi-Omics Approach towards Identification of Causes and Consequences.

Microvesicle generation is an integral part of the aging process of red blood cells in vivo and in vitro. Extensive vesiculation impairs function and survival of red blood cells after transfusion, and microvesicles contribute to transfusion reactions. The triggers and mechanisms of microvesicle generation are largely unknown. In this study, we combined morphological, immunochemical, proteomic, lipidomic, and metabolomic analyses to obtain an integrated understanding of the mechanisms underlying microvesicle generation during the storage of red blood cell concentrates. Our data indicate that changes in membrane organization, triggered by altered protein conformation, constitute the main mechanism of vesiculation, and precede changes in lipid organization. The resulting selective accumulation of membrane components in microvesicles is accompanied by the recruitment of plasma proteins involved in inflammation and coagulation. Our data may serve as a basis for further dissection of the fundamental mechanisms of red blood cell aging and vesiculation, for identifying the cause-effect relationship between blood bank storage and transfusion complications, and for assessing the role of microvesicles in pathologies affecting red blood cells.

Network Pharmacology-Based Dissection of the Anti-diabetic Mechanism of Lobelia chinensis.

Diabetes mellitus (DM) is a chronic inflammatory disease, and the rapidly increasing DM is becoming a major problem of global public health. Traditional Chinese medicine (TCM) has a long history of treating diabetes. It has been developed and utilized because of its good efficacy and no toxic side effects. Lobelia chinensis is a traditional whole grass herbal. With the continuous deepening of pharmacological research on TCM, the active ingredients of L. chinensis are continuously revealed, which contained the alkaloids, flavonoids, flavonoid glycosides and amino acids that have the good effects of anti-inflammatory, anti-viral and anti-diabetic. In order to further explore the targets of active ingredients and its anti-diabetic mechanism, a feasible network pharmacology analysis model based on chemical, pharmacokinetic and pharmacological data was developed by network construction method to clarify the anti-diabetic mechanism of L. chinensis. The present study conducted by gas chromatography–mass spectrometer (GC/MS), which identified 208 metabolites of L. chinensis, of which 23 ingredients may have effective pharmacological effects after absorption, distribution, metabolism, and excretion (ADME) screening. Network pharmacological analysis on the active ingredients revealed that 5-hydroxymethylfurfural in L. chinensis affects the insulin resistance signaling pathway by acting on GSK3B, TNF, and MAPK1, acacetin affects the diabetic pathway by acting on INSR, DPP4, and GSK3B, that regulate type 2 diabetes, non-insulin-dependent DM, and inflammatory diseases. These results successfully indicated the potential anti-diabetic mechanism of the active ingredients of L. chinensis.

P57Kip2 is a master regulator of human adipose derived stem cell quiescence and senescence.

Although human adipose derived stem cells (hADSCs) hold great promises for regenerative medicine, their key biological properties remain poorly understood. In particular, proliferation defects resulted from deep quiescence (dormancy) and senescence represent a major hurdle in hADSC production and clinical application. We have developed a model system for mechanistic dissection of hADSC quiescence and senescence. p57Kip2, a major CDK inhibitor, was highly expressed in quiescent and senescent hADSCs but its level quickly declined upon stem cell activation. p57Kip2 overexpression induced quiescence in spite of proliferative signals and its knockdown promoted cell cycle reentry even with induction of quiescence presumably through modulating the CDK2-CyclinE1 complex. Given its key role in quiescence and senescence, p57Kip2 may be exploited for innovative strategies to amplify hADSCs of high quality for clinics.

Tumor Suppressors in Chronic Lymphocytic Leukemia: From Lost Partners to Active Targets.

Tumor suppressors play an important role in cancer pathogenesis and in the modulation of resistance to treatments. Loss of function of the proteins encoded by tumor suppressors, through genomic inactivation of the gene, disable all the controls that balance growth, survival, and apoptosis, promoting cancer transformation. Parallel to genetic impairments, tumor suppressor products may also be functionally inactivated in the absence of mutations/deletions upon post-transcriptional and post-translational modifications. Because restoring tumor suppressor functions remains the most effective and selective approach to induce apoptosis in cancer, the dissection of mechanisms of tumor suppressor inactivation is advisable in order to further augment targeted strategies. This review will summarize the role of tumor suppressors in chronic lymphocytic leukemia and attempt to describe how tumor suppressors can represent new hopes in our arsenal against chronic lymphocytic leukemia (CLL).

Characterization of Photosynthetic Phenotypes and Chloroplast Ultrastructural Changes of Soybean (Glycine max) in Response to Elevated Air Temperatures.

Heat stress negatively affects photosynthesis in crop plants. Chlorophyll fluorescence provides information about the efficiency of the light-dependent reactions of photosynthesis and can be measured non-destructively and rapidly. Four soybean (Glycine max) genotypes were grown in controlled environments at 28/20°C (control), followed by imposition of control, 38/28°C, and 45/28°C day/night temperature regimes for 7 days. Coordinated chlorophyll fluorescence, gas exchange, and chloroplast ultrastructure measurements were conducted over the course of the 7-day temperature treatments and revealed contrasting responses among the different genotypes. Although generally similar, the extent of the impact of elevated temperatures on net photosynthesis differed among genotypes. Despite dramatic effects on photosynthetic light reactions, net photosynthetic rates were not reduced by exposure to 45°C on the 1st day of treatment imposition. Temporal dynamics of light reaction characteristics over the course of the 7-day heat-wave simulation revealed distinct responses among the genotypes. Similarly, chloroplast ultrastructure examination identified contrasting responses of DT97-4290 and PI603166, particularly with respect to starch characteristics. These changes were positively associated with differences in the percent area of chloroplasts that were occupied by starch grains. Elevated temperature increased number and size of starch grains on the 1st day of DT97-4290 which was coordinated with increased minimum chlorophyll fluorescence (F0) and reduced leaf net CO2 assimilation (A). Whereas on the 7th day the elevated temperature treatment showed reduced numbers and sizes of starch grains in chloroplasts and was coordinated with similar levels of F0 and A to the control treatment. Unlike starch dynamics of PI603166 which elevated temperature had little effect on. The genotypic differences in photosynthetic and chloroplast ultrastructure responses to elevated temperatures identified here are of interest for the development of more tolerant soybean cultivars and to facilitate the dissection of molecular mechanisms underpinning heat stress tolerance of soybean photosynthesis.

Tumor-educated B cells promote renal cancer metastasis via inducing the IL-1\u03b2/HIF-2\u03b1/Notch1 signals

While B cells in the tumor microenvironment (TME) might play important roles in cancer progression, their impacts on the renal cell carcinoma (RCC) metastasis remained unclear, which drew our attention to further explore. We found that RCC tissues could recruit more B cells than the surrounding normal renal tissues from human clinical RCC samples. Wound healing assay, transwell assay and 3D invasion assays demonstrated that recruited B cells, also known as tumor-educated B cells (TEB), could significantly increase the RCC cell migration and invasion. In addition, in vivo data from xenograft RCC mouse model also confirmed that TEB could enhance RCC cell invasive and metastatic capability. Mechanism dissection revealed that TEB activated IL-1β/HIF-2α signals in RCC cells that could induce the downstream Notch1 signaling pathway. The above results demonstrated the key roles of TEB within renal cancer associated tumor microenvironment were metastasis-promotor and might help us to develop the potential therapies via targeting these newly identified IL-1β/HIF-2α/Notch1 signals in RCC progression.