Biology subjects

Guan, H.

Publications and source records attributed to Guan, H..

At least 19 recordsLinked to original sources

bioRxiv2026

Stability-driven multi-omics integration for reproducible latent structure

High-dimensional multi-omics data integration offers novel opportunities to characterize complex biological systems. Even though sampling variability frequently compromises findings, particularly in small cohorts, the reproducibility and generalizability of the derived latent structures are insufficiently evaluated. We propose a Stability-driven framework for multi-omics integration that combines sparse generalized canonical correlation analysis with repeated cross-validation, out-of-sample projection, and systematic evaluation of both component-level and feature-level stability. We apply this framework to untargeted metabolomic and Olink targeted inflammation proteomic profiles in a thyroid cancer case-control cohort (n = 162). Our Stability-driven integration identified reproducible metabolomic and proteomic latent components that showed consistent out-of-sample disease associations and tracked temporally structured changes relative to time to diagnosis. The proposed framework provides a generalizable strategy for identifying reproducible latent structures that improve robustness of biological inference in multi-omics studies.

bioinformatics↗

bioRxiv2026

Dysregulation of a novel autophagosome-mitochondria contact contributes to autophagy dysfunction and neurodegeneration in tauopathy

Mitochondria engage in extensive communication with other organelles through membrane contacts. Perturbed mitochondria-organelle interactions are indicated in a variety of neurodegenerative diseases, but the underlying mechanisms remain poorly understood. Here, we report a new class of mitochondria-organelle communication: autophagosome/autophagic vacuole (AV)-mitochondria (Mito) contact, which exhibits hyper-tethering in tauopathy neurons, consequently hampering AV retrograde transport. Such defects are attributed to accelerated turnover of the contact release factor TBC1D15, triggered by mitochondrial bioenergetic deficit-induced hyperactivity of the AMP-activated protein kinase (AMPK). Increasing TBC1D15 levels or repressing AMPK activity normalizes AV-Mito contact release and restores retrograde transport of AVs, thereby increasing autophagic cargo clearance and reducing tau burden in tauopathy axons. Furthermore, overexpression of TBC1D15 enhances autophagic clearance and attenuates tau pathology, alleviating neurodegeneration and cognitive dysfunction in tauopathy mice. Taken together, our study provides new insights into AV-Mito contact dysregulation in tauopathy-related autophagy failure, laying the groundwork for the development of potential therapeutics to combat tauopathy diseases.

neuroscience↗

bioRxiv2026

Progranulin deficiency induces lipid droplet accumulation in microglia via a STAT3-GPAT3 axis

Heterozygous mutations in the progranulin (PGRN) encoding gene GRN cause frontotemporal dementia (FTD), whereas homozygous GRN mutations lead to neuronal ceroid lipofuscinosis (NCL). However, the mechanisms underlying neurodegeneration due to PGRN deficiency remain unclear. In the aged brains or under neurodegenerative conditions, the accumulation of lipid droplets (LDs) in microglia contributes to cellular dysfunction and pro-inflammatory responses, exacerbating neurodegenerative pathology. Here, we investigated how PGRN deficiency induces LD accumulation in microglia. We found that PGRN ablation significantly upregulated triglycerol-3-phosphate acyltransferase 3 (GPAT3), the first and rate-limiting enzyme for triacylglycerol biosynthesis, and increased LD formation in both Grn-/- BV2 microglial cells and Grn-/-mouse brains. Mechanistic study revealed that PGRN deficiency upregulates GPAT3 via stimulating signal transduction and activator of transcription factor 3 (STAT3) signaling. Silencing Gpat3, inhibiting GPAT3 activity with the small molecule inhibitor FSG67, or PGRN restoration reduced LD accumulation and rescued the cytotoxicity of Grn-/- microglia conditional medium (MCM) toward N2a cells. Notably, intraperitoneal administration of FSG67 or AAV-MG1.2-mCx3cr1-mGrn-eGFP-mediated microglia-specific gene therapy reduced microglial LDs and ameliorated behavior phenotypes in Grn-/- mice. Our study elucidates the role of PGRN loss in microglial lipid homeostasis and identifies the STAT3-GPAT3 axis as a potential therapeutic target for LD-associated neurodegenerative diseases.

neuroscience↗

bioRxiv2025

Elevated neuronal TAF15 expression leads to FTD-like phenotype

TAF15, a DNA/RNA-binding protein whose dysfunction in alternative splicing impairs NMDA receptor signaling, can form amyloid fibrils that contribute to neurodegeneration in the neurodegenerative diseases like FTD and ALS, indicating a close involvement of TAF15 in both neuronal physiological function and activation of pathological pathways. However, the relationship between TAF15 expression levels and neurodegeneration, as well as the specific downstream pathways mediating its neurotoxicity, remains unclear. Here, we found a consistent upregulation of TAF15 in prefrontal cortex neurons from patients across multiple FTD and ALS subtypes. Both in vitro and in vivo experiments demonstrated that neuronal TAF15 overexpression triggered oxidative stress, evidenced by reactive oxygen species (ROS) production, leading to neurotoxicity and gliosis. Mice overexpressing TAF15 in medial prefrontal cortex (mPFC) neurons exhibited heightened anxiety and impaired fear memory, mimicking some key features of FTD behavioral abnormalities. Notably, these pathological and behavioral phenotypes are rescued by the antioxidant N-acetylcysteine amide (NACA), indicating that elevated TAF15 level exacerbates neurodegeneration primarily by activating oxidative stress pathways. Together, this work elucidates a novel TAF15-oxidative stress axis in FTD-associated neurodegeneration, providing a conceptual framework for future therapeutic development. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=120 SRC="FIGDIR/small/694105v1_ufig1.gif" ALT="Figure 1000"> View larger version (35K): org.highwire.dtl.DTLVardef@cc581eorg.highwire.dtl.DTLVardef@1ad2129org.highwire.dtl.DTLVardef@15c0fb5org.highwire.dtl.DTLVardef@10b57a6_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsTAF15 is upregulated in prefrontal cortex neurons of patients with FTD and ALS across different subtypes. Elevated TAF15 expression provokes ROS elevation, leading to neurotoxicity and gliosis. Neuronal TAF15 overexpression in the mouse mPFC results in FTD-like anxiety and impaired fear-memory phenotype. Reducing oxidative stress can rescue FTD-like phenotype caused by TAF15 upregulation.

neuroscience↗

bioRxiv2025

Overcoming Extrapolation Challenges of Deep Learning by Incorporating Physics in Protein Sequence-Function Modeling

Understanding protein sequence-to-function relationship is crucial to assist studies of genetic diseases, protein evolution, and protein engineering. The sequence-to-function relationship of proteins is inherently complex due to multi-site high-dimensional correlation and structural dynamics. Deep learning algorithms such as (graph) convolutional neural networks and recently transformers have become very popular for learning the protein sequence-to-function mapping from deep mutational scanning data and available structures. However, it remains very challenging for these models to achieve accurate extrapolation when predicting functional effect of variants with positions or mutation types not seen in the training data. We propose that incorporating the physics of protein interactions and dynamics can be an effective approach to overcome the extrapolation limitations. Specifically, we demonstrate that physics-based modeling can be used to quantify the energetic effects of mutations and that incorporating these physical energetics directly within the convolution and graph convolution neural networks can significantly improve the performance of positional and mutational extrapolation compared to models without biophysics-inspired features. Our results support the effectiveness of leveraging physical knowledge in overcoming the limitation of data scarcity.

bioinformatics↗

bioRxiv2025

The deubiquitinating enzyme Otu1 releases substrates from the conserved initiation complex of the Cdc48/p97 ATPase for proteasomal degradation

Many eukaryotic proteins are modified with a polyubiquitin chain and then recruited to either the Cdc48 ATPase (p97 or VCP in mammals) or the 26S proteasome by conserved cofactors. They can then shuttle between the Cdc48 ATPase and the 26S proteasome before being degraded. How substrates avoid being trapped on the Cdc48 ATPase complex is incompletely understood, as they can undergo repeated cycles of translocation through the ATPase pore. Here, we show that the deubiquitinating enzyme (DUB) Otu1 (Yod1 in mammals) can break this futile cycle. Otu1 trims the ubiquitin chain of the substrate before its translocation through the Cdc48 pore is initiated, allowing transfer to the proteasome and subsequent degradation. A cryo-EM structure shows that the mammalian homolog Yod1 binds to p97 simultaneously with other Cdc48 cofactors. As in the yeast system, polypeptide translocation through the ATPase pore is initiated by the unfolding of a ubiquitin molecule, suggesting that the mechanism of substrate processing is conserved in all eukaryotes.

biochemistry↗

bioRxiv2025

The Global Canopy Atlas: analysis-ready maps of 3D structure for the world's woody ecosystems

Woody canopies regulate exchanges of energy, water and carbon, and their three-dimensional (3D) structure supports much of terrestrial biodiversity. Remote sensing technologies such as airborne laser scanning (ALS) now enable the 3D mapping of entire landscapes. However, we lack the large, harmonized and geographically representative ALS collections needed to build a global picture of woody ecosystem structure. To address this challenge, we developed the Global Canopy Atlas (GCA): 3,458 ALS acquisitions transformed into standardized and analysis-ready maps of canopy height and elevation at 1 m2 resolution. The GCA covers 56,554 km2 across all major biomes. 19% of this area has been scanned multiple times, and 87% of all GCA products are openly available, covering 95% of the total area. To showcase its wide range of applications, we applied the GCA in three case studies. First, we validated three global satellite-derived canopy height maps, finding poor performance at native resolution (1-30 m, R2 < 0.38) and moderate performance at 250 m resolution (R2 < 0.65). Second, analyzing global patterns in canopy gap size frequency we discovered an unexpectedly large variation of power law exponents from branch to stand level ( = 1.52 to 2.38), pointing to a fundamental scale-dependence of forest structure. Third, we developed a framework to standardize forest turnover quantification from multi-source, multi-temporal ALS. In a temperate forest in North America it revealed that 21% of canopy gaps closed within 12 years of opening and would thus be missed by infrequent monitoring. As demonstrated by these case studies, the GCA provides a novel data source for ecologists, foresters, remote sensing scientists and the ecosystem modelling community that substantially advances our ability to understand the structure and dynamics of woody ecosystems at global scales.

ecology↗

bioRxiv2025

ZUGC-RNA degradation generates immunosuppressor to evade immune responses in eukaryotes

Among the hundreds of modified nucleosides identified in terrestrial life, 2-amino-6-aminopurine (Z) is widely recognized as a prominent modified purine. Recently, RNA written with the ZUGC alphabet shows significant potential in RNA therapeutics as a synthetic biosystem. Here, we demonstrate that ZUGC-RNA can evade immune recognition in eukaryotes, independent of factors such as RNA length, sequence, 5-triphosphate, modified uridine, and secondary structure. Notably, we discovered that both the degradation of ZUGC-RNA and metabolites of Z-nucleotides can function as immunosuppressors, silencing TLR7 sensing to block immune responses. This mechanism differs from that of pseudo-uridine ({Psi}) modified RNA currently in use. ZUGC-RNAs also demonstrate broad applicability across multiple neural cell types. Our findings provide valuable insights for developing more tolerable RNA-based drugs and designing immunomodulators targeting TLR7. In addition to the potential prebiotic relevance of Z, our finding not only contributes to understanding the RNA world hypothesis but also provides new insights into the exploration of the origin of life.

synthetic biology↗

bioRxiv2025

Metabolic shifts in the heart of hypothyroid mice -- insights from untargeted metabolomics

ABSTRACTSO_ST_ABSIntroductionC_ST_ABSHypothyroidism is a prevalent disorder that affect all systems of body, including the cardiovascular system and control of cardiac function. However, much remains unknown about the metabolic pathways and disorders in the heart induced by hypothyroidism, as well as the metabolic mechanisms involved. ObjectivesThis work aims to investigate the impacts of hypothyroidism on the heart and explore its underlying mechanisms through untargeted metabolomics analysis. MethodsAn experimental model of hypothyroidism was established by giving water containing 0.1% methimazole and 1% potassium perchlorate to 12-week-old C57BL/6J mice for six consecutive weeks. ResultsAfter treatment, hypothyroidism significantly reduced mices heart weight and heart rate. Metabolomics analysis revealed that the different metabolites (DEMs) primarily concentrated in Phospholipids, Lipids or Lipids-like, Carbohydrates and Carbohydrate conjugates. These specific metabolites were mainly enriched in pathways related to lipid metabolism. The hearts of hypothyroid mice exhibited significantly lower levels of lipoyl-carnitine compared to controls, along with markedly higher levels of glycolysis and pentose phosphate pathway (PPP) intermediates. Furthermore, carnitine palmitoyltransferase 1B (CPT1B) and carnitine palmitoyltransferase 2 (CPT2) expressions were significantly decreased in the hypothyroid mouse heart, while lactate dehydrogenase A (LDHA) expressions was significantly increased, according to the immunoblotting assay. ConclusionsResearch indicates that hypothyroidism triggers disruptions in both cardiac energy metabolism and phospholipid metabolism in adult mice, which in turn results in myocardial atrophy. These findings could pave the way for novel therapeutic strategies aimed at treating hypothyroidism-induced myocardial atrophy.

pathology↗

bioRxiv2024

High-dimensional mediation analysis to elucidate the role of metabolites in the association between PFAS exposure and reduced SARS-CoV-2 IgG in pregnancy

We previously found that per- and polyfluoroalkyl substances (PFAS) mixture exposure is inversely associated with SARS-CoV-2 IgG (IgG) antibody levels in pregnant individuals. Here, we aim to identify metabolites mediating this relationship to elucidate the underlying biological pathways. This cross-sectional study included 59 pregnant participants from a US-based pregnancy cohort. Untargeted metabolomic profiling was performed using Liquid Chromatography-High Resolution Sass spectrometry (LC-HRMS), and weighted Quantile Sum (WQS) regression was applied to assess the PFAS and metabolites mixture effects on IgG. Metabolite indices positively or negatively associated with IgG levels were constructed separately and their mediation effects were examined independently and jointly. The PFAS- index was negatively associated with IgG levels (beta=-0.315, p<0.001), with PFHpS and PFHxS as major contributors. Two metabolite-indices were constructed, one positively (beta=1.249, p<0.001) and one negatively (beta=-1.200, p<0.001) associated with IgG. Key contributors for these indices included trigonelline, adipate, p-octopamine, and n-acetylproline. Analysis of a single mediator showed that 74.6% (95% CI: 45.9%, 98.0%) and 68.6% (95% CI: 41.8%, 94.1%) of the PFAS index-IgG total effect were mediated by the negative and positive metabolites-indices, respectively. Joint analysis of the metabolite-indices indicated a cumulative mediation effect of 83.8% (95% CI: 58.1%, 98.7%). Enriched pathways associated with these metabolites indices were phenylalanine, tyrosine, and tryptophan biosynthesis and arginine metabolism. We observed significant mediation effects of plasma metabolites on the PFAS-IgG relationship, suggesting that PFAS is associated with alteration in the balance of plasma metabolites that contributes to reduced plasma IgG production.

bioinformatics↗

bioRxiv2024

Evolutionary analysis of paired box gene family and biological function exploration of Lr.Pax7 in lamprey (Lethenteron reissneri)

The Pax(paired box)gene family comprises highly conserved transcription factors that play a crucial regulatory role in embryonic development, tissue morphogenesis, and cell differentiation. The gene family comprises of nine members, namely Pax1 to Pax9, each exhibiting distinct functionalities and expression patterns. Among the members of the Pax family, Pax7 is particularly involved in key regulatory functions related to skeletal muscle development and satellite cell biology in vertebrates. However, the precise role of Pax7 and its downstream targets in jawless vertebrates such as lamprey remains relatively understudied. Therefore, investigating the expression and function of lampreys Pax genes holds significant value as a model organism for understanding developmental processes, evolution, and interactions with other vertebrates. In this study, we identified Pax7 in Lethentero reissneri and multiple bioinformatics analyses suggested that it may represent the ancestral gene of the Pax7 shared among higher vertebrates. To further elucidate the function and regulatory mechanism of Pax7, we performed transcriptome sequencing analysis after silencing Lr.Pax7 expression which revealed alterations in metabolic processes along with associated genes(Plcy,Jagged2,Hes,Myod,Frizzled2,etc) involved in various immune pathways(Notch, Wnt, IL-17, TNF, etc), myogenesis process, adipocyte formation process consistent with those observed in higher vertebrates. Comparative studies between lamprey and high vertebrates could provide valuable insight into understanding the functional role of Pax7 and its downstream targets across diverse biological process as well as disease states.

evolutionary biology↗

bioRxiv2024

Cervical collagen network porosity assessed by SHG endomicroscopy distinguishes preterm and normal pregnancy - a pilot study

Structured AbstractO_ST_ABSBackgroundC_ST_ABSPreterm birth (PTB) is a global public health issue affecting millions of newborns every year. Orchestrated remodeling of the cervix is essential for normal pregnancy and birth, while PTB is closely related with premature cervical ripening and loss of cervical mechanical strength. The structure and organization of fibrillar collagen in the extracellular matrix are of vital importance to the biomechanical properties of the cervix. Second harmonic generation (SHG) microscopy has proved capable of revealing the progressive changes in cervical collagen morphology over the course of pregnancy. To translate this promising imaging technology to clinical practice, a flexible SHG endomicroscope has long been envisaged for label-free, non-invasive visualization of cervical collagen architecture and for assessment of PTB risk. ObjectiveTo evaluate the potential of our newly-developed SHG endomicroscope for imaging-based differentiation of cervical collagen architecture between normal pregnant mice and RU486/mifepristone-induced PTB mouse models. Study DesignWe undertook endomicroscopy SHG imaging of cervical collagen on two types of ex vivo samples: 1) frozen cervical tissue sections ([~]50 {micro}m thick) and 2) resected intact cervices, and performed SHG-image-based quantitative collagen morphology analysis to distinguish RU486 mouse models from normal pregnant mice. ResultsEndomicroscopic SHG images of cervical tissue sections from mifepristone-treated mouse models exhibit statistically larger collagen fiber diameter, increased pore size, and reduced pore numbers than those of normal pregnant mice. Similar changes are also observed on SHG images of subepithelial collagen fibers acquired from intact cervices by the endomicroscope. ConclusionThe experiment results demonstrated that SHG endomicroscopy along with quantitative image analysis holds promising potential for clinical assessment of cervical collagen remodeling and preterm birth risk.

bioengineering↗

bioRxiv2024

Mitochondrial bioenergetics stimulates autophagy for pathological tau clearance in tauopathy neurons

Hyperphosphorylation and aggregation of microtubule-associated tau is a pathogenic hallmark of tauopathies and a defining feature of Alzheimers disease (AD). Pathological tau is targeted by autophagy for clearance, but autophagy dysfunction is indicated in tauopathy. While mitochondrial bioenergetic failure has been shown to precede the development of tau pathology, it is unclear whether energy metabolism deficiency is involved in tauopathy-related autophagy defects. Here, we reveal that stimulation of anaplerotic metabolism restores defective oxidative phosphorylation (OXPHOS) in tauopathy which, strikingly, leads to enhanced autophagy and pronounced tau clearance. OXPHOS-induced autophagy is attributed to increased ATP-dependent phosphatidylethanolamine biosynthesis in mitochondria. Excitingly, early bioenergetic stimulation boosts autophagy activity and reduces tau pathology, thereby counteracting memory impairment in tauopathy mice. Taken together, our study sheds light on a pivotal role of bioenergetic dysfunction in tauopathy-linked autophagy defects and suggests a new therapeutic strategy to prevent toxic tau buildup in AD and other tauopathies.

neuroscience↗

bioRxiv2023

Autoantibodies against eukaryotic translation elongation factor 1 delta (EEF1D) in two patients with autoimmune cerebellar ataxia

To report a novel autoantibody against eukaryotic translation elongation factor 1 delta (EEF1D) in two patients with autoimmune cerebellar ataxia (ACA). The patients on one center with cerebellar ataxia of unknown cause, who were tested positive with tissue-based indirect immunofluorescence assay (TBA) on rat cerebellum sections and negative for comprehensive anti-neural autoantibodies panel, were investigated for novel autoantibody identification. Tissue-immunoprecipitation (TIP) combined with mass spectrometric (MS) analysis was used to identify the target antigen. The EEF1D protein was identified from complex precipitated by serum and cerebrospinal fluid (CSF) of one patient, while the specific binding of autoantibodies and EEF1D were confirmed by subsequent neutralization experiment, recombinant cell-based indirect immunofluorescence assay (CBA) and western blot. In consequently test using CBA, another anti-EEF1D positive ACA patient was identified and confirmed by western blot. All enrolled 30 health doners and 15 connective tissue diseases patients without neurological disorders were anti-EEF1D negative. The two anti-EEF1D positive patients presented similar clinical manifestations, their outcomes supported the effectiveness of immunotherapy, but the cerebellar atrophy occurred before treatment may be irreversible. Symptoms of one patient worsened again after the weaning of pulse corticosteroid therapy. The identification of anti-EEF1D autoantibody provides a new potential biomarker for early diagnosis and recurrence prevention of ACA.

immunology↗

bioRxiv2021

Extraction, purification and anti-TMV effects of α (β)-2,7,11-cembratriene-4,6-diol from tobacco leaves

BackgroundAcetone ethanol extracts from tobacco leaves have antiviral activity against TMV, but the antiviral effects of their specialized metabolites have not been systematically studied yet, especially the underlying mechanism is still unexplored. ResultsThe tobacco cembranoids ({beta})-2,7,11-cembratriene-4,6-diol (({beta})-CBD) were extracted and purified with an effective and green protocol including innovatively added 5% phosphoric acid for elution, one time silica gel chromatographic column separation and impurity removal and further HPLC purification. The results of antiviral activities against tobacco mosaic virus (TMV) with the local lesion counting method showed that ({beta})-CBD have in vivo higher protective effects of 73.2% and 71.6%, at 75.0 M, respectively, than control agent Ningnanmycin (53.1%). Notably, The results of ELISA and and TMV-GFP fluorescent optical imaging assay indicated a obviously reduced viral protein and weaker GFP fluorescence signal and smaller infection area, which confirmed their anti-TMV activities at protein level. Furthermore, the enhanced production of SA and JA and the significantly increased transcription of of JA signaling pathway (COI1 and PDF1.2) and SA signaling pathway genes (PR1, NPR1 and EDS1) in treated plants further conformed that exogenously applied ({beta})-CBD can effectively elicit the tobacco plant immunity against TMV. ConclusionsThe ({beta})-CBD mainly stimulates disease resistance of tobacco plants to resist TMV and it can be used as bioagents to control TMV in the future.

plant biology↗

bioRxiv2020

Specific and conserved patterns of microbiota-structuring by maize benzoxazinoids in the field

BackgroundPlants influence their root and rhizosphere microbial communities through the secretion of root exudates. However, how specific classes of root exudate compounds impact the assembly of these root-associated microbiotas is not well understood. Maize roots secrete benzoxazinoids (BXs), a class of indole-derived defense compounds, and thereby impact the assembly of their microbiota. Here, we investigated the broader impacts of BX exudation on root and rhizosphere microbiotas of adult maize plants grown under natural conditions at different field locations in Europe and the US. We examined the microbiotas of BX-producing and multiple BX-defective lines in two genetic backgrounds across three soil types. ResultsOur analysis showed that the secretion of BXs affected community composition of rhizosphere and root microbiota, with the most pronounced effects observed for root fungi. The impact of the two genetic backgrounds was weaker than that of the presence or absence of BXs, suggesting that BX exudation is a key trait by which maize structures its associated microbiota. BX-producing plants were not consistently enriching microbial lineages across the three soil types. Instead, BX exudation consistently depleted Flavobacteriaceae and Comamonadaceae, and enriched various plant pathogenic fungi in the roots. ConclusionsThese findings reveal that BXs have a selective impact on root and rhizosphere microbiota composition across different field locations. Taken together, this study identifies the BX pathway as an interesting breeding target to manipulate plant-microbiome interactions.

plant biology↗

bioRxiv2020

LINCS gene expression signatures analysis revealed bosutinib as a radiosensitizer of breast cancer cells by targeting eIF4G1

Radioresistance represents the predominant cause for radiotherapy failure and disease progression, resulting in increased breast cancer mortality. Through gene expression signatures analyses of Library of Integrated Network-Based Cellular Signatures (LINCS) and Gene Expression Omnibus (GEO), the present study aimed to identify potential candidate radiosensitizers from known drugs systematically. The similarity of integrated gene expression signatures between irradiated eIF4G1-silenced breast cancer cells and known drugs was measured by enrichment scores. Drugs with positive enrichment scores were selected as potential radiosensitizers. The radiosensitizing effects of the candidate radiosensitizers were analyzed in breast cancer cells (MCF-7, MX-1, and MDA-MB-231) by CCK-8 assays and colony-forming capability after exposure to ionizing radiation. Cell apoptosis was detected by flow cytometry. Expressions of eIF4G1 and a series of DNA damage response proteins were analyzed by Western blot assays. Bosutinib was proposed to be a promising radiosensitizer as its administration markedly reduced the dosages of both the drug and ionizing radiation and was associated with fewer adverse drug reactions. The combined treatment with ionizing radiation and bosutinib significantly increased the cells killing potency in all three cell lines as compared to ionizing radiation or bosutinib alone. MX-1 cells were revealed to be the most sensitive to both ionizing radiation and bosutinib among the three cell lines. Bosutinib noticeably downregulated the expression of eIF4G1 in a dose-dependent manner and also reduced the expression of DNA damage response proteins (including ATM, XRCC4, ATRIP, and GADD45a). Moreover, eIF4G1 could be a key target of bosutinib through which it regulates DNA damage induced by ionizing radiation. Thus, taken together, bosutinib may serve as a potential candidate radiosensitizer for breast cancer therapy.

cancer biology↗

bioRxiv2020

Molecular basis of ubiquitination catalyzed by the bacterial transglutaminase MavC

The Legionella pneumophila effector MavC is a transglutaminase that carries out atypical ubiquitination of the ubiquitin (Ub) E2 conjugation enzyme UBE2N by catalyzing the formation of an isopeptide bond between Gln40 of Ub and Lys92 (or to a less extent, Lys94) of UBE2N, which results in inhibition of UBE2N signaling in the NF-{kappa}B pathway. In the absence of UBE2N, MavC deamidates Ub at Gln40 or catalyzes self-ubiquitination. However, the mechanisms underlying these enzymatic activities of MavC are not fully understood at molecular level. In this study, we obtained the structure of the MavC-UBE2N-Ub ternary complex that represents a snapshot of covalent cross-linking of UBE2N and Ub catalyzed by MavC. The structure reveals the unique way by which the cross-linked catalytic product UBE2N-Ub binds mainly to the Insertion and the Tail domains of MavC prior to its release. Based on our structural, biochemical and mutational analyses, we proposed the catalytic mechanism for both the deamidase and the transglutaminase activities of MavC. Finally, by comparing the structures of MavC and MvcA, the homologous protein that reverses MavC-induced UBE2N ubiquitination, we identified several key regions of the two proteins responsible for their opposite enzymatic activity. Our results provide insights into the mechanisms for substrate recognition and ubiquitination mediated by MavC as well as explanations for the opposite activity of MavC and MvcA.

microbiology↗