F-box Research Articles

FBXO24 modulates mRNA alternative splicing and MIWI degradation and is required for normal sperm formation and male fertility.

Spermiogenesis is a critical, post-meiotic phase of male gametogenesis, in which the proper gene expression is essential for sperm maturation. However, the underFlying molecular mechanism that controls mRNA expression in the round spermatids remains elusive. Here, we identify that FBXO24, an orphan F-box protein, is highly expressed in the testis of humans and mice and interacts with the splicing factors (SRSF2, SRSF3, and SRSF9) to modulate the gene alternative splicing in the round spermatids. Genetic mutation of FBXO24 in mice causes many abnormal splicing events in round spermatids, thus affecting a large number of critical genes related to sperm formation that were dysregulated. Further molecular and phenotypical analyses revealed that FBXO24 deficiency results in aberrant histone retention, incomplete axonemes, oversized chromatoid body, and abnormal mitochondrial coiling along sperm flagella, ultimately leading to male sterility. In addition, we discovered that FBXO24 interacts with MIWI and SCF subunits and mediates the degradation of MIWI via K48-linked polyubiquitination. Furthermore, we show that FBXO24 depletion could lead to aberrant piRNA production in testes, which suggests FBXO24 is required for normal piRNA counts. Collectively, these data demonstrate that FBXO24 is essential for sperm formation by regulating mRNA alternative splicing and MIWI degradation during spermiogenesis.

Genome-Wide Identification, Characterization and Expression Profile of F-Box Protein Family Genes Shed Light on Lateral Branch Development in Cultivated Peanut (Arachis hypogaea L.)

F-box proteins are a large gene family in plants, and play crucial roles in plant growth, development, and stress response. To date, a comprehensive investigation of F-box family genes in peanuts, and their expression pattern in lateral branch development has not been performed. In this study, a total of 95 F-box protein family members on 18 chromosomes, named AhFBX1-AhFBX95, were identified in cultivated peanut (Arachis hypogaea L.), which were classified into four groups (Group I–IV). The gene structures and protein motifs of these peanut FBX genes were highly conserved among most FBXs. We found that significant segmental duplication events occurred between wild diploid species and the allotetraploid of peanut FBXs, and observed that AhFBXs underwent strong purifying selection throughout evolution. Cis-acting elements related to development, hormones, and stresses were identified in the promoters of AhFBX genes. In silico analysis of AhFBX genes revealed expression patterns across 22 different tissues. A total of 32 genes were predominantly expressed in leaves, pistils, and the aerial gynophore tip. Additionally, 37 genes displayed tissue-specific expression specifically at the apex of both vegetative and reproductive shoots. During our analysis of transcriptome data for lateral branch development in spreading and erect varieties, namely M130 and JH5, we identified nine deferentially expressed genes (DEGs). Quantitative real-time PCR (qRT-PCR) results further confirmed the expression patterns of these DEGs. These DEGs exhibited significant differences in their expression levels at different stages between M130 and JH5, suggesting their potential involvement in the regulation of lateral branch development. This systematic research offers valuable insights into the functional dissection of AhFBX genes in regulating plant growth habit in peanut.

Synthetically derived BiAux modulates auxin co-receptor activity to stimulate lateral root formation.

The root system plays an essential role in plant growth and adaptation to the surrounding environment. The root clock periodically specifies lateral root prebranch sites (PBS), where a group of pericycle founder cells (FC) is primed to become lateral root founder cells and eventually give rise to lateral root primordia or lateral roots (LRs). This clock-driven organ formation process is tightly controlled by modulation of auxin content and signaling. Auxin perception entails the physical interaction of TRANSPORT INHIBITOR RESPONSE 1 (TIR1) or AUXIN SIGNALING F-BOX (AFBs) proteins with AUXIN/INDOLE-3-ACETIC ACID (Aux/IAA) repressors to form a co-receptor system. Despite the apparent simplicity, the understanding of how specific auxin co-receptors are assembled remains unclear. We identified the compound bis-methyl auxin conjugated with N-glucoside, or BiAux, in Arabidopsis (Arabidopsis thaliana) that specifically induces the formation of PBS and the emergence of LR, with a slight effect on root elongation. Docking analyses indicated that BiAux binds to F-box proteins, and we showed that BiAux function depends on TIR1 and AFB2 F-box proteins and AUXIN RESPONSE FACTOR 7 activity, which is involved in FC specification and LR formation. Finally, using a yeast (Saccharomyces cerevisiae) heterologous expression system, we showed that BiAux favors the assemblage of specific co-receptors subunits involved in LR formation and enhances AUXIN/INDOLE-3-ACETIC ACID 28 protein degradation. These results indicate that BiAux acts as an allosteric modulator of specific auxin co-receptors. Therefore, BiAux exerts a fine-tune regulation of auxin signaling aimed to the specific formation of LR among the many development processes regulated by auxin.

Cullin-RING ligases employ geometrically optimized catalytic partners for substrate targeting

Cullin-RING ligases (CRLs) ubiquitylate specific substrates selected from other cellular proteins. Substrate discrimination and ubiquitin transferase activity were thought to be strictly separated. Substrates are recognized by substrate receptors, such as Fbox or BCbox proteins. Meanwhile, CRLs employ assorted ubiquitin-carrying enzymes (UCEs, which are a collection of E2 and ARIH-family E3s) specialized for either initial substrate ubiquitylation (priming) or forging poly-ubiquitin chains. We discovered specific human CRL-UCE pairings governing substrate priming. The results reveal pairing of CUL2-based CRLs and UBE2R-family UCEs in cells, essential for efficient PROTAC-induced neo-substrate degradation. Despite UBE2R2's intrinsic programming to catalyze poly-ubiquitylation, CUL2 employs this UCE for geometrically precise PROTAC-dependent ubiquitylation of a neo-substrate and for rapid priming of substrates recruited to diverse receptors. Cryo-EM structures illuminate how CUL2-based CRLs engage UBE2R2 to activate substrate ubiquitylation. Thus, pairing with a specific UCE overcomes E2 catalytic limitations to drive substrate ubiquitylation and targeted protein degradation.

BFP1: One of 700 Arabidopsis F-box proteins mediates degradation of JA oxidases to promote plant immunity

BFP1: One of 700 Arabidopsis F-box proteins mediates degradation of JA oxidases to promote plant immunity

Endoribonuclease DNE1 Promotes Ethylene Response by Modulating EBF1/2 mRNA Processing in Arabidopsis.

The gaseous phytohormone ethylene plays a crucial role in plant growth, development, and stress responses. In the ethylene signal transduction cascade, the F-box proteins EIN3-BINDING F-BOX 1 (EBF1) and EBF2 are identified as key negative regulators governing ethylene sensitivity. The translation and processing of EBF1/2 mRNAs are tightly controlled, and their 3' untranslated regions (UTRs) are critical in these regulations. However, despite their significance, the exact mechanisms modulating the processing of EBF1/2 mRNAs remain poorly understood. In this work, we identified the gene DCP1-ASSOCIATED NYN ENDORIBONUCLEASE 1 (DNE1), which encodes an endoribonuclease and is induced by ethylene treatment, as a positive regulator of ethylene response. The loss of function mutant dne1-2 showed mild ethylene insensitivity, highlighting the importance of DNE1 in ethylene signaling. We also found that DNE1 colocalizes with ETHYLENE INSENSITIVE 2 (EIN2), the core factor manipulating the translation of EBF1/2, and targets the P-body in response to ethylene. Further analysis revealed that DNE1 negatively regulates the abundance of EBF1/2 mRNAs by recognizing and cleaving their 3'UTRs, and it also represses their translation. Moreover, the dne1 mutant displays hypersensitivity to 1,4-dithiothreitol (DTT)-induced ER stress and oxidative stress, indicating the function of DNE1 in stress responses. This study sheds light on the essential role of DNE1 as a modulator of ethylene signaling through regulation of EBF1/2 mRNA processing. Our findings contribute to the understanding of the intricate regulatory process of ethylene signaling and provide insights into the significance of ribonuclease in stress responses.

The F-box protein RhSAF destabilizes the gibberellic acid receptor RhGID1 to mediate ethylene-induced petal senescence in rose.

Roses are among the most popular ornamental plants cultivated worldwide for their great economic, symbolic, and cultural importance. Nevertheless, rapid petal senescence markedly reduces rose (Rosa hybrida) flower quality and value. Petal senescence is a developmental process tightly regulated by various phytohormones. Ethylene accelerates petal senescence, while gibberellic acid (GA) delays this process. However, the molecular mechanisms underlying the crosstalk between these phytohormones in the regulation of petal senescence remain largely unclear. Here, we identified SENESCENCE-ASSOCIATED F-BOX (RhSAF), an ethylene-induced F-box protein gene encoding a recognition subunit of the SCF-type E3 ligase. We demonstrated that RhSAF promotes degradation of the GA receptor GIBBERELLIN INSENSITIVE DWARF1 (RhGID1) to accelerate petal senescence. Silencing RhSAF expression delays petal senescence, while suppressing RhGID1 expression accelerates petal senescence. RhSAF physically interacts with RhGID1s and targets them for ubiquitin/26S proteasome-mediated degradation. Accordingly, ethylene-induced RhGID1C degradation and RhDELLA3 accumulation are compromised in RhSAF-RNAi lines. Our results demonstrate that ethylene antagonizes GA activity through RhGID1 degradation mediated by the E3 ligase RhSAF. These findings enhance our understanding of the phytohormone crosstalk regulating petal senescence and provide insights for improving flower longevity.

Knockdown of TRIP13 inhibits gastric cancer stemness and cisplatin resistance by regulating FBXW7/ENO1 axis

Purpose: To explore the role of thyroid hormone receptor interactor 13 (TRIP13) in gastric cancer (GC) and the associated mechanism.
 Methods: TRIP13 expression in GC was found in The Cancer Genome Atlas (TCGA) database and siTRIP13 was transfected in GC cells in order to generate cell lines with low TRIP13 expression. Cell proliferation was determined by colony formation assay, while cell migration and invasion were evaluated by scratch test and Transwell assay, respectively. Sphere formation assay was used to assess cell stemness characteristics whereas half-inhibitory concentration (IC50) value was evaluated by Cell Counting Kit-8 (CCK8). F-box and WD repeat domain containing 7 (FBXW7) and enolase 1 (ENO1) expression were determined by western-blot assay.
 Results: TRIP13 was significantly expressed in GC based on TCGA database (p < 0.001) and high expression predicted poor prognosis in GC patients (p < 0.05). Based on the findings of the cell function assay, si-TRIP13 inhibited the proliferation, migration, invasion and stemness of GC cells (p < 0.001) and decreased the IC50 value. Mechanically, knockdown of TRIP13 decreased ENO1 expression level by stabilizing FBXW7.
 Conclusion: TRIP13 functions as an oncogene in GC and stimulates growth and stemness via FBXW7/ENO1 pathway in GC. Thus, a potential therapeutic target for the treatment of gastric cancer is provided by this study.

The F-box protein ZmFBL41 negatively regulates disease resistance to Rhizoctonia solani by degrading the abscisic acid synthase ZmNCED6 in maize.

The maize F-box protein ZmFBL41 targets abscisic acid synthase 9-cis-epoxycarotenoid dioxygenase 6 for degradation, and this regulatory module is exploited by Rhizoctonia solani to promote infection. F-box proteins are crucial regulators of plant growth, development, and responses to abiotic and biotic stresses. Previous research identified the F-box gene ZmFBL41 as a negative regulator of maize (Zea mays) defenses against Rhizoctonia solani. However, the precise mechanisms by which F-box proteins mediate resistance to R. solani remain poorly understood. In this study, we show that ZmFBL41 interacts with an abscisic acid (ABA) synthase, 9-cis-epoxycarotenoid dioxygenase 6 (ZmNCED6), promoting its degradation via the ubiquitination pathway. We discovered that the ectopic overexpression of ZmNCED6 in rice (Oryza sativa) inhibited R. solani infection by activating stomatal closure, callose deposition, and jasmonic acid (JA) biosynthesis, indicating that ZmNCED6 enhances plant immunity against R. solani. Natural variation at ZmFBL41 across different maize haplotypes did not affect the ZmFBL41-ZmNCED6 interaction. These findings suggest that ZmFBL41 targets ZmNCED6 for degradation, leading to a decrease in ABA levels in maize, in turn, inhibiting ABA-mediated disease resistance pathways, such as stomatal closure, callose deposition, and JA biosynthesis, ultimately facilitating R. solani infection.

A Sorghum F-Box Protein Induces an Oxidative Burst in the Defense Against Colletotrichum sublineola.

The hemibiotrophic fungal pathogen Colletotrichum sublineola is the causal agent of anthracnose in sorghum (Sorghum bicolor), resulting in leaf blight, stalk rot, and head blight in susceptible genotypes, with yield losses of up to 50%. The development of anthracnose-resistant cultivars can reduce reliance on fungicides and provide a more sustainable and economical means for disease management. A previous genome-wide association study of the sorghum association panel identified the candidate resistance gene Sobic.005G172300 encoding an F-box protein. To better understand the role of this gene in the defense against C. sublineola, gene expression following infection with C. sublineola was monitored by RNA sequencing in seedlings of sorghum accession SC110, which harbored the resistance allele, and three accessions that harbored a susceptible allele. Only in SC110 did the expression of Sobic.005G172300 increase during the biotrophic phase of infection. Subsequent transcriptome analysis, gene co-expression networks, and gene regulatory networks of inoculated and mock-inoculated seedlings of resistant and susceptible accessions suggest that the increase in expression of Sobic.005G172300 induces an oxidative burst by lowering the concentration of ascorbic acid during the biotrophic phase of infection. Based on gene regulatory network analysis, the protein encoded by Sobic.005G172300 is proposed to target proteins involved in the biosynthesis of ascorbic acid for polyubiquitination through the SCF E3 ubiquitin ligase, causing their degradation via the proteasome.

DNMT1-mediated epigenetic suppression of FBXO32 expression promoting cyclin dependent kinase 9 (CDK9) survival and esophageal cancer cell growth

ABSTRACT Esophageal cancer (EC) is a common and serious form of cancer, and while DNA methyltransferase-1 (DNMT1) promotes DNA methylation and carcinogenesis, the role of F-box protein 32 (FBXO32) in EC and its regulation by DNMT1-mediated methylation is still unclear. FBXO32 expression was examined in EC cells with high DNMT1 expression using GSE163735 dataset. RT-qPCR assessed FBXO32 expression in normal and EC cells, and impact of higher FBXO32 expression on cell proliferation, migration, and invasion was evaluated, along with EMT-related proteins. The xenograft model established by injecting EC cells transfected with FBXO32 was used to evaluate tumor growth, apoptosis, and tumor cells proliferation and metastasis. Chromatin immunoprecipitation (ChIP) assay was employed to study the interaction between DNMT1 and FBXO32. HitPredict, co-immunoprecipitation (Co-IP), and Glutathione-S-transferase (GST) pulldown assay analyzed the interaction between FBXO32 and cyclin dependent kinase 9 (CDK9). Finally, the ubiquitination assay identified CDK9 ubiquitination, and its half-life was measured using cycloheximide and confirmed through western blotting. DNMT1 negatively correlated with FBXO32 expression in esophageal cells. High FBXO32 expression was associated with better overall survival in patients. Knockdown of DNMT1 in EC cells increased FBXO32 expression and suppressed malignant phenotypes. FBXO32 repressed EC tumor growth and metastasis in mice. Enrichment of DNMT1 in FBXO32 promoter region led to increased DNA methylation and reduced transcription. Mechanistically, FBXO32 degraded CDK9 through promoting its ubiquitination.

FBXO8 is a novel prognostic biomarker in different molecular subtypes of breast cancer and suppresses breast cancer progression by targeting c-MYC

F-box only protein 8 (FBXO8) is a recently identified member of the F-box proteins, showcasing its novelty in this protein family. Extensive research has established FBXO8's role as a tumor suppressor in various cancers, including hepatocellular carcinoma, and colorectal cancer, Nevertheless, its functional, mechanistic, and prognostic roles in primary and metastatic breast cancer, particularly in different molecular subtypes of breast cancer, various stages, as well as its potential implications in immunotherapy, tumor microenvironment, and prognostic survival among breast cancer patients, remain unexplored. In this article, we employed a multi-dimensional investigation leveraging TCGA, TIMER, TISIDB, STRING, MEXPRESS, UALCAN, and cBioPortal databases to explore the underlying suppression mechanism of FBXO8 in breast cancer. FBXO8 negatively correlates with MYC, NOTCH, WNT and inflammatory signaling pathways in breast tumor microenvironment. Furthermore we conducted RT-PCR, western blot, cell proliferation, cell migration, and mRNA target gene RT-PCR analyses to elucidate the role of FBXO8 in breast cancer progression. Mechanistically, PTEN and FBXW7 expression were down-regulated and MYC, IL10, IL6, NOTCH1, WNT6 mRNA expressions were up-regulated in FBXO8 knockdown cell lines. c-MYC silenced cells showed an increase in FBXO8 protein level, which suggests a negative feedback loop between FBXO8 and c-MYC to control breast cancer metastasis. These findings illuminate the novel role of FBXO8 as a prognostic and therapeutic target across different molecular subtypes of breast cancer. Finally, through the utilization of virtual screening and Molecular Dynamics simulations, we successfully identified two FDA-approved medications, Ledipasvir and Paritaprevir, that demonstrated robust binding capabilities and interactions with FBXO8

FBXW7 polymorphism asserts susceptibility to colorectal cancer

FBXW7, belonging to the F-Box protein family, is considered a candidate cancer susceptibility gene. Our findings indicate that single nucleotide polymorphisms (SNPs) in the FBXW7 gene are linked to cancer risk, strengthening FBXW7′s role in the pathogenesis of colorectal cancer. Our case-control study comprised of 450 patients diagnosed with colorectal cancer (CRC) and an equal number of 450 healthy subjects. FBXW7 SNPs rs2255137C>T and rs6842544C>T were genotyped using PCR-Restriction Fragment Length Polymorphism (PCR-RFLP) and Single-Stranded Conformation Polymorphism (SSCP) techniques and further cross-checked by direct sequencing. Linkage disequilibrium and haplotype analyses of these SNPs were also assessed. The in-silico approach was used to reveal the functional analysis between the nonsynonymous variation (rs6842544) and CRC followed by its validation at the protein level by western blotting and reverse transcription-PCR. A significant association of colorectal cancer was detected with rs6842544 SNP. However, there was no association between FBXW7 rs2255137 polymorphism and CRC. The homozygous individuals carrying the C variant in FBXW7 rs6842544 showed a slightly higher risk for colorectal cancer (OR = 1.590, 95%CI = 0.39 ∼ 2.89, p = 0.011). The haplotype CC identified in this study seemed to be associated with good prognosis (OR = 1.22, 95% CI = 1.00 ∼ 1.47, p = 0.0013) whereas the TT haplotype was found to reduce the CRC risk (OR = 0.642, 95%CI = 0.48 ∼ 0.84, p = 0.039). In-silico prediction proposed that the variant R133G is responsible for the lower expression of FBXW7. Additionally, the expression profiling of FBXW7 nonsynonymous SNP was significantly lower in primary CRC tissues than in the paired non-cancerous tissues at protein and mRNA levels. The study indicates that the FBXW7 rs6842544 is associated with the risk of development of CRC and could serve as a molecular biological marker to screen high-risk groups for CRC.

Identification and expression analysis of genomic regions associated with the traits contributing to lodging tolerance in wheat (Triticum aestivum L.)

Crop lodging, either stem breaking/stem bending or root lodging, is a major concern for farmers worldwide. In the present study, meta-QTLs, ortho-MQTLs and candidate genes related to lodging and its associated traits were identified using 836 of the 1269 known QTLs retrieved from 107 QTL mapping studies published during 1998–2022. The study resulted in the identification of 86 MQTLs with an average confidence interval (CI) of 1.66 cM (4.26-fold reduction) as against a CI of > 7.08 cM in the initial QTLs. Out of 86 MQTLs, a subset of 12 MQTLs demonstrated stability and robustness, earning them the title of breeders’ MQTLs. These selections were made based on criteria such as the number of initial QTLs involved, small CI, high LOD scores, and PVE values. Furthermore, leveraging synteny and collinearity facilitated the identification of 10 ortho-MQTLs shared between wheat and maize. Further, in silico expression analysis allowed the detection of 53 significantly expressed candidate genes encoding different proteins/products including the following: F-box protein, GDSL lipase/esterases, SANT/Myb domain transcription factor, pectin esterase inhibitor domain, glycoside hydrolase family, protein kinase domain, zinc finger domain, cytochrome P450, UDP-glucuronosyl/UDP-glucosyltransferase, glycosyltransferase 61 etc. In addition, 26 lodging tolerance genes from Arabidopsis, rice, and maize were also utilized to identify 34 wheat homologues in wheat MQTL regions. Among these, three genes TraesCS2D02G256600 (TaEREB114-2D), TraesCS7B02G340100 (TaIRX14-7B), and TraesCS1B02G347600 (TaGA2ox4-1B) were prioritized for qRT-PCR-based expression analysis in one lodging-tolerant (i.e., WH-1105) and one lodging-susceptible (i.e., WH-1184) variety. When comparing these two varieties, we found higher relative expression levels of these three genes in the second basal internode at the milky endosperm stage of the lodging-tolerant variety. The MQTLs, ortho-MQTLs, and candidate genes (CGs) identified in this study can be utilized for marker-assisted selection to improve lodging tolerance and enhance our understanding of the molecular basis of lodging tolerance in wheat.

FBXO5-mediated RNF183 degradation prevents endoplasmic reticulum stress-induced apoptosis and promotes colon cancer progression

Endoplasmic reticulum (ER) stress induces the unfolded protein response (UPR), and prolonged ER stress leads to cell apoptosis. Despite increasing research in this area, the underlying molecular mechanisms remain unclear. Here, we discover that ER stress upregulates the UPR signaling pathway while downregulating E2F target gene expression and inhibiting the G2/M phase transition. Prolonged ER stress decreases the mRNA levels of E2F2, which specifically regulates the expression of F-Box Protein 5(FBXO5), an F-box protein that functions as an inhibitor of the anaphase-promoting complex/cyclosome (APC/C) ubiquitin ligase complex. Depletion of FBXO5 results in increased ER stress-induced apoptosis and decreased expression of proteins related to PERK/IRE1α/ATF6 signaling. Overexpression of FBXO5 wild-type (not its ΔF-box mutant) alleviates apoptosis and the expression of the C/EBP Homologous Protein (CHOP)/ATF. Mechanistically, we find that FBXO5 directly binds to and promotes the ubiquitin-dependent degradation of RNF183, which acts as a ubiquitin E3 ligase in regulating ER stress-induced apoptosis. Reversal of the apoptosis defects caused by FBXO5 deficiency in colorectal cancer cells can be achieved by knocking down RNF183 in FBXO5-deficient cells. Functionally, we observed significant upregulation of FBXO5 in colon cancer tissues, and its silencing suppresses tumor occurrence in vivo. Therefore, our study highlights the critical role of the FBXO5/RNF183 axis in ER stress regulation and identifies a potential therapeutic target for colon cancer treatment.

The LRR receptor-like kinase ALR1 is a plant aluminum ion sensor

Plant survival requires an ability to adapt to differing concentrations of nutrient and toxic soil ions, yet ion sensors and associated signaling pathways are mostly unknown. Aluminum (Al) ions are highly phytotoxic, and cause severe crop yield loss and forest decline on acidic soils which represent ∼30% of land areas worldwide. Here we found an Arabidopsis mutant hypersensitive to Al. The gene encoding a leucine-rich-repeat receptor-like kinase, was named Al Resistance1 (ALR1). Al ions binding to ALR1 cytoplasmic domain recruits BAK1 co-receptor kinase and promotes ALR1-dependent phosphorylation of the NADPH oxidase RbohD, thereby enhancing reactive oxygen species (ROS) generation. ROS in turn oxidatively modify the RAE1 F-box protein to inhibit RAE1-dependent proteolysis of the central regulator STOP1, thus activating organic acid anion secretion to detoxify Al. These findings establish ALR1 as an Al ion receptor that confers resistance through an integrated Al-triggered signaling pathway, providing novel insights into ion-sensing mechanisms in living organisms, and enabling future molecular breeding of acid-soil-tolerant crops and trees, with huge potential for enhancing both global food security and forest restoration.

FBXL20 promotes synaptic impairment in depression disorder via degrading vesicle-associated proteins

BackgroundSynaptic plasticity changes in presynaptic terminals or postsynaptic membranes play a critical role in cognitive impairments and emotional disorders, but the underlying molecular mechanisms in depression remain largely unknown. MethodsThe regulation effects of F-box and leucine-rich repeat protein 20 (FBXL20), vesicular glutamate transporter 1 (VGLUT1) and vesicle-associated membrane protein 1 (VAMP1) on synaptic plasticity and depressive-like behaviors examined by proteomics analysis, viral stereotaxic injection, transmission electron microscope and biochemical methods. The glutamate release detected by fluorescent probe in cultured primary pyramidal neurons. ResultsWe found that chronic unpredictable mild stress (CUMS) induced significant synaptic deficits within hippocampus of depressed rats, accompanied with the decreased expression of VGLUT1 and VAMP1. Moreover, knockdown of VGLUT1 or VAMP1 in hippocampal pyramidal neurons resulted in abnormal glutamatergic neurotransmitter release. In addition, we found that the E3 ubiquitin ligase FBXL20 was increased within hippocampus, which may promote ubiquitination and degradation of VGLUT1 and VAMP1, and thus resulted in the reduction of glutamatergic neurotransmitter release, the disruptions of synaptic transmission and the induction of depression-like behaviors in rats. In contrast, shRNA knockdown of FBXL20 within the hippocampus of depressed rats significantly ameliorated synaptic damage and depression-like behaviors. LimitationOnly one type of depression model was used in the present study, while other animal models should be used in the future to confirm the underlying mechanisms reported here. ConclusionsThis study provides new insights that inhibiting FBXL20 pathway in depressed rats may be an effective strategy to rescue synaptic transmission and depression-like behaviors.

Potential role of Fbxo22 in resistance to endocrine therapy in breast cancer with invasive lobular carcinoma

BackgroundInvasive lobular carcinoma (ILC) is distinct from invasive ductal carcinoma (IDC) in terms of their hormonal microenvironments that may require different therapeutic strategies. We previously reported that selective estrogen receptor modulator (SERM) function requires F-box protein 22 (Fbxo22). Here, we investigated the role of Fbxo22 as a potential biomarker contributing to the resistance to endocrine therapy in ILC.MethodsA total of 302 breast cancer (BC) patients including 150 ILC were recruited in the study. Fbxo22 expression and clinical information were analyzed to elucidate whether Fbxo22 negativity could be a prognostic factor or there were any correlations among clinical variables and SERM efficacy.ResultsFbxo22 negativity was significantly higher in ILC compared with IDC (58.0% vs. 27.0%, P < 0.001) and higher in postmenopausal patients than premenopausal patients (64.1% vs. 48.2%, P = 0.041). In the ILC cohort, Fbxo22-negative patients had poorer overall survival (OS) than Fbxo22-positive patients, with 10-year OS rates of 77.4% vs. 93.6% (P = 0.055). All patients treated with SERMs, Fbxo22 negativity resulted in a poorer outcome, with 10-year OS rates of 81.3% vs. 92.3% (P = 0.032). In multivariate analysis regarding recurrence-free survival (RFS) in ILC patients, Fbxo22 status was independently predictive of survival as well as lymph node metastasis.ConclusionFbxo22 negativity significantly impacts on survival in BC patients with IDC and ILC, and the disadvantage was enhanced among ILC postmenopausal women or patients treated with SERMs. The findings suggest that different therapeutic strategies might be needed according to the different histopathological types when considering adjuvant endocrine therapy.

Integrated Transcriptomics and Metabolomics Analysis Promotes the Understanding of Adventitious Root Formation in Eucommia ulmoides Oliver.

As a primary approach to nutrient propagation for many woody plants, cutting roots is essential for the breeding and production of Eucommia ulmoides Oliver. In this study, hormone level, transcriptomics, and metabolomics analyses were performed on two E. ulmoides varieties with different adventitious root (AR) formation abilities. The higher JA level on the 0th day and the lower JA level on the 18th day promoted superior AR development. Several hub genes executed crucial roles in the crosstalk regulation of JA and other hormones, including F-box protein (EU012075), SAUR-like protein (EU0125382), LOB protein (EU0124232), AP2/ERF transcription factor (EU0128499), and CYP450 protein (EU0127354). Differentially expressed genes (DEGs) and metabolites of AR formation were enriched in phenylpropanoid biosynthesis, flavonoid biosynthesis, and isoflavonoid biosynthesis pathways. The up-regulated expression of PAL, CCR, CAD, DFR, and HIDH genes on the 18th day could contribute to AR formation. The 130 cis-acting lncRNAs had potential regulatory functions on hub genes in the module that significantly correlated with JA and DEGs in three metabolism pathways. These revealed key molecules, and vital pathways provided more comprehensive insight for the AR formation mechanism of E. ulmoides and other plants.

COP9 signalosome-mediated deneddylation of CULLIN1 is necessary for SCFEBF1 assembly in Arabidopsis thaliana

Functions of the SKP1-CUL1-F box (SCF) ubiquitin E3 ligases are essential in plants. The F box proteins (FBPs) are substrate receptors that recruit substrates and assemble an active SCF complex, but the regulatory mechanism underlying the FBPs binding to CUL1 to activate the SCF cycle is not fully understood. We show that Arabidopsis csn1-10 is defective in SCFEBF1-mediated PIF3 degradation during de-etiolation, due to impaired association of EBF1 with CUL1 in csn1-10. EBF1 preferentially associates with un-neddylated CUL1 that is deficient in csn1-10 and the EBF1-CUL1 binding is rescued by the neddylation inhibitor MLN4924. Furthermore, we identify a subset of FBPs with impaired binding to CUL1 in csn1-10, indicating their assembly to form SCF complexes may depend on COP9 signalosome (CSN)-mediated deneddylation of CUL1. This study reports that a key role of CSN-mediated CULLIN deneddylation is to gate the binding of the FBP-substrate module to CUL1, thus initiating the SCF cycle of substrate ubiquitination.