Biology subjects

Zhang, L.

Publications and source records attributed to Zhang, L..

At least 1,045 records · Page 58Linked to original sources

bioRxiv2022

Redifferentiated cardiomyocytes retain residual dedifferentiation signatures and are protected against ischaemic injury

Cardiomyocyte renewal by dedifferentiation and proliferation has fueled the field of regenerative cardiology in recent years, while the reverse process of redifferentiation remains largely unexplored. Redifferentiation is characterised by the restoration of function that is lost during dedifferentiation and is key to the healing process following injury. Previously, we showed that ERBB2-mediated heart regeneration has these two distinct phases: dedifferentiation, followed by redifferentiation. Here, using temporal RNAseq and proteomics, we survey the landscape of the dedifferentiation-redifferentiation process in the adult mouse heart. We find well characterised dedifferentiation pathways, such as reduced oxphos, increased proliferation and increased EMT-like features, largely return to normal, though elements of residual dedifferentiation remain, even after contractile function is restored. These hearts appeared rejuvenated and showed robust resistance to ischaemic injury. We find that redifferentiation is driven by negative feedback signalling, notably through LATS1/2 Hippo pathway activity. Disabling LATS1/2 in dedifferentiated cardiomyocytes augments dedifferentiation in vitro and prevents redifferentiation in vivo. Taken together, our data reveal the non-trivial nature of redifferentiation, whereby elements of dedifferentiation linger in a surprisingly beneficial manner. This cycle of dedifferentiation-redifferentiation protects against future insult, in what could become a novel prophylactic treatment against ischemic heart disease for at-risk patients.

cell biology↗

bioRxiv2022

Identification of a small molecule, Mebendazole, as a novel ethylene signaling activator reveals a role of ethylene signaling on the regulation of lateral root angles

The lateral root angle or gravitropic set-point angle (GSA) is an important trait for root system architecture (RSA) that determines the radial expansion of the root system. The GSA therefore plays a crucial role for the ability of plants to access nutrients and water in the soil. Despite its importance, only few regulatory pathways and mechanisms that determine GSA are known, and these mostly relate to auxin and cytokinin pathways. Here, we report the identification of a small molecule, Mebendazole (MBZ) that modulates GSA in Arabidopsis thaliana roots and acts via the activation of ethylene signaling. We uncover that MBZ directly acts on the serine/threonine protein kinase CTR1, which is a negative regulator of ethylene signaling. Our study not only reveals that the ethylene signaling pathway is essential for GSA regulation, but it also identifies a small molecular modulator of RSA and the first small molecule that acts downstream of ethylene receptors and that directly activates ethylene signaling. In briefHe et al. identify a small molecule that regulates lateral root angle. They show that the compound increases lateral root angle by inhibiting CTR1 kinase activity, which in turn activates ethylene signaling. Therefore, they uncover that the ethylene pathway is involved in lateral root angle regulation. HighlightsO_LIMBZ increases lateral root angle C_LIO_LIMBZ regulates lateral root angle by activating ethylene signaling C_LIO_LIMBZ inhibits CTR1 kinase activity C_LIO_LIThe ethylene pathway is involved in lateral root angle regulation C_LI

plant biology↗

bioRxiv2022

On Correlation between Structural Properties and Viral Escape Measurements from Deep Mutational Scanning

Encouraged by recent efforts to map responses of SARS-CoV-2 mutations to various antibody treatments with deep mutational scanning, we explored the possibility of tying measurable structural contact information from the binding complexes of antibodies and their targets to experimentally determined viral escape responses. With just a single crystal structure for each binding complex, we find that the average correlation coefficient R is surprisingly high at 0.76. Our two methods for calculating contact information use binary contacts measured between all residues of two proteins. By varying the parameters to obtain binary contacts, we find that 3.6 [A] and 7 [A] are pivotal distances to toggle the binary step function when tallying the contacts for each method. The correlations are improved by short simulations ([~]25 ns), which increase average R to 0.78. With blind tests using the random forest model, we can further improve average R to 0.84. These easy-to-implement measurements can be utilized in computational screening of viral mutations that escape antibody treatments and potentially other protein-protein interaction problems.

bioinformatics↗

bioRxiv2022

Phosphoregulation of DSB-1 mediates control of meiotic double-strand break activity

In the first meiotic cell division, proper segregation of chromosomes in most organisms depends on chiasmata, exchanges of continuity between homologous chromosomes that originate from the repair of programmed double-strand breaks (DSBs) catalyzed by the Spo11 endonuclease. Since DSBs can lead to irreparable damage in germ cells, while chromosomes lacking DSBs also lack chiasmata, the number of DSBs must be carefully regulated, so as to be neither too high nor too low. Here, we show that in Caenorhabditis elegans, meiotic DSB levels are controlled by the phosphoregulation of DSB-1, a homolog of the yeast Spo11 cofactor Rec114, by the opposing activities of PP4PPH-4.1 phosphatase and ATRATL-1 kinase. Increased DSB-1 phosphorylation in pph-4.1 mutants correlates with reduction in DSB formation, while prevention of DSB-1 phosphorylation drastically increases the number of meiotic DSBs both in pph-4.1 mutants as well as in the wild type background. C. elegans and its close relatives also possess a diverged paralog of DSB-1, called DSB-2, and loss of dsb-2 is known to reduce DSB formation in oocytes with increasing age. We show that the proportion of the phosphorylated, and thus inactivated, form of DSB-1 increases with age and also upon loss of DSB-2, while non-phosphorylatable DSB-1 rescues the age-dependent decrease in DSBs in dsb-2 mutants. These results suggest that DSB-2 evolved in part to compensate for the inactivation of DSB-1 through phosphorylation, to maintain levels of DSBs in older animals. Our work shows that PP4PPH-4.1, ATRATL-1, and DSB-2 act in concert with DSB-1 to promote optimal DSB levels throughout the reproductive lifespan.

cell biology↗

bioRxiv2022

Novel ERR pan-agonists ameliorate heart failure through boosting cardiac fatty acid metabolism and mitochondrial function

Cardiac metabolic dysfunction is a hallmark of heart failure. Estrogen related receptors ERR and ERR{gamma} are essential regulators for cardiac metabolism. Therefore, activation of ERR could be a potential therapeutic intervention for heart failure. However, no natural or synthetic ERR agonist is available to demonstrate their pharmacological effect in vivo. Using a structure-based design approach, we designed and synthesized two structurally distinct pan-ERR agonists, SLU-PP-332 (332) and SLU-PP-915 (915), which significantly improved ejection fraction and ameliorated fibrosis against pressure overload-induced heart failure without affecting cardiac hypertrophy. Mechanistically, a broad-spectrum of metabolic genes were transcriptionally activated by ERR agonists, particularly genes involved in fatty acid metabolism and mitochondrial function, which were mainly mediated by ERR{gamma}. Metabolomics analysis showed significant normalization of metabolic profiles in fatty acid/lipid and TCA/OXPHOS metabolites by 915 in the mouse heart with 6-week pressure overload. Autophagy was also induced by ERR agonists in cardiomycoyte. On the other hand, ERR agonism led to downregulation of cell cycle and development pathways, which was partially mediated by E2F1 in cardiomyocyte. In summary, ERR agonists maintain oxidative metabolism, which confers cardiac protection against pressure overload-induced heart failure in vivo. Our results provided direct pharmacological evidence supporting the further development of ERR agonists as novel heart failure therapeutics in vivo.

pharmacology and toxicology↗

bioRxiv2022

Single-cell transcriptomic and proteomic analysis of Parkinson's disease Brains

Parkinsons disease (PD) is a prevalent neurodegenerative disorder where recent evidence suggests pathogenesis may be mediated by inflammatory processes. The molecular architecture of the disease remains to be fully elucidated. We performed single-nucleus transcriptomics and unbiased proteomics using postmortem tissue obtained from the prefrontal cortex of 12 individuals with late-stage PD and age-matched controls. We analyzed [~]80,000 nuclei and identified eight major cell types, including brain-resident T cells, each with distinct transcriptional changes in line with the known genetics of PD. By analyzing Lewy body pathology in the same postmortem tissue, we found that -synuclein pathology is inversely correlated with chaperone expression in excitatory neurons. Examining cell-cell interactions, we found a selective abatement of neuron-astrocyte interactions and enhanced neuroinflammation. Proteomic analyses of the same brains identified synaptic proteins in prefrontal cortex that were preferentially downregulated in PD. Strikingly, comparing this dataset to a regionally similar published analysis for Alzheimers disease (AD), we found no common differentially expressed genes in neurons, but identified many shared differentially expressed genes in glial cells, suggesting that disease etiology in PD and AD are likely distinct. These data are presented as a resource for interrogating the molecular and cellular basis of PD and other neurodegenerative diseases. One Sentence SummaryWe provide an extensive single cell analysis profiling nearly 80,000 brain nuclei from prefrontal cortex of late-stage Parkinsons disease brains, demonstrate that -synuclein pathology is inversely correlated with chaperone expression in excitatory neurons, found a selective abatement of neuron-astrocyte interactions with enhanced neuroinflammation, and augmented the study with proteomic analysis and cross-comparisons with Alzheimers disease datasets, providing valuable insights into the pathways of neurodegeneration and a deep definition of the underlying molecular pathology for Parkinsons disease.

neuroscience↗

bioRxiv2022

Speciation in Nearctic oak gall wasps is frequently correlated with changes in host plant, host organ, or both.

Quantifying the frequency of shifts to new host plants within diverse clades of specialist herbivorous insects is critically important to understand whether and how host shifts contribute to the origin of species. Oak gall wasps (Hymenoptera: Cynipidae: Cynipini) comprise a tribe of ~1000 species of phytophagous insects that induce gall formation on various organs of trees in the family Fagacae, -- primarily the oaks (genus Quercus; ~435 sp). The association of oak gall wasps with oaks is ancient (~50 my), and most oak species are galled by one or more gall wasp species. Despite the diversity of both gall wasp species and their plant associations, previous phylogenetic work has not identified a strong signal of host plant shifting among oak gall wasps. However, most emphasis has been on the Western Palearctic and not the Nearctic where both oaks and oak gall wasps are considerably more species rich and where oaks are more phylogenetically diverse. We collected 86 species of Nearctic oak gall wasps from 10 of the 14 major clades of Nearctic oaks and sequenced >1000 Ultra Conserved Elements (UCEs) and flanking sequences to infer wasp phylogenies. We assessed the relationships of Nearctic gall wasps to one another and, by leveraging previously published UCE data, to the Palearctic fauna. We then used phylogenies to infer historical patterns of shifts among host tree species and tree organs. Our results indicate that oak gall wasps have moved between the Palearctic and Nearctic at least four times, that some Palearctic clades have their proximate origin in the Nearctic, and that gall wasps have shifted within and between oak tree sections, subsections, and organs considerably more often than the analysis of previous data have suggested. Given that host shifts have been demonstrated to drive reproductive isolation between host-associated populations in other phytophagous insects, our analyses of Nearctic gall wasps suggest that host shifts are key drivers of speciation in this clade, especially in hotspots of oak diversity. Though formal assessment of this hypothesis requires further study, two putatively oligophagous gall wasp species in our dataset show signals of host-associated genetic differentiation unconfounded by geographic distance, suggestive of barriers to gene flow associated with the use of alternative host plants.

evolutionary biology↗

bioRxiv2022

Chronic Intermittent Hypobaric Hypoxia Prevents Contrast-Induced Acute Kidney Injury By Modulating The HIF-1α Signaling Pathway

The aim of this study was to explore the role of CIHH in preventing contrast-induced acute kidney injury (CI-AKI) in rats and its mechanism. Rats mean arterial pressure, heart rate, serum creatinine and blood urea nitrogen levels were measured. The kidney tissue pathological changes, superoxide dismutase (SOD) activity, malondialdehyde (MDA) levels, hypoxia inducible factor-1, Bcl-2/adenovirus E1B-19kDa-interacting protein3 (BNIP3) , cysteiny aspartate specific protease3(caspase3) and poly(ADP-ribose) polymerase (PARP) expression levels were testing. The results showed that CIHH prevented CI-AKI group mean arterial pressure, heart rate, serum creatinine and blood urea nitrogen levels were reduced, kidney tissue SOD activity was increased, MDA levels was reduced, HIF-1,BNIP3,caspase3 and PARP levels were increased than the CI-AKI group. This study indicates that CIHH pretreatment may have a protective effect on contrast-induced early kidney injury by activating the HIF-1/BNIP3 signaling pathway to regulate mitochondrial autophagy and enhance cellular anti-apoptotic and renal antioxidant capacity, for the first time.

physiology↗

bioRxiv2022

Fate tracing reveals differences between Reelin+ HSCs and Desmin+ HSCs in activation, migration, and proliferation activities

The activation of hepatic stellate cells (HSCs) which comprise distinct clusters, is the main cause of liver fibrogenesis in response to different etiologies of chronic liver injuries. In this study, we constructed a novel ReelinCreERT2 transgenic mouse in which cells expressing Reelin were fully marked and demonstrated that about 50% HSCs were labeled. These ReelinCreERT2-labeled HSCs displayed distinct characteristics in migration, activation and proliferation compared to Desmin+ HSCs (total HSCs) in cholestatic (bile duct ligation; BDL) or hepatotoxic (carbon tetrachloride; CCl4) liver injuries. In BDL-induced fibrotic livers, Desmin+ HSCs were activated with increased proliferation and accumulation activities around the portal triad, but mGFP+ HSCs did not show proliferation or accumulation activity around the portal triad, and only a small part was activated. In CCl4-induced fibrotic livers, most of Desmin+ and mGFP+ HSCs were activated along with proliferation and accumulation potential around the central vein, however fewer mGFP+ HSCs were activated compared to Desmin+ HSCs. Moreover, in the regression of CCl4-induced fibrosis, mGFP+ HSCs were apoptosed whereas Desmin+ HSCs recovered to normal state. Besides, we didnt find evidence that mGFP+ HSCs transdifferentiated into hepatocytes or cholangiocytes through mesenchymal-epithelial transition (MET).

pathology↗

bioRxiv2022

A potent SARS-CoV-2 neutralizing antibody recognizing a conserved epitope with broad mutant variant and SARS-CoV activity.

COVID-19 is the deadliest respiratory virus pandemic since 1918 and the latest of several coronavirus epidemics and pandemics in recent years. Despite the unprecedented response by both the government and private sectors to develop vaccines and therapies, the evolution of SARS-CoV-2 variants resistant to these interventions reveals a crucial need for therapeutics that maintain their efficacy against current and future mutant variants. Here we describe a SARS-CoV-2 neutralizing antibody, ABP-310, with potent activity against all variants tested including the Omicron variant. ABP-310 also displays potent neutralizing activity against SARS-CoV, highlighting the conserved nature of the ABP-310 epitope. By targeting a conserved epitope, we believe that ABP-310 has therapeutic promise not only against the current SARS-CoV-2 variants but would be expected to maintain efficacy against future variants and possibly even novel coronaviruses.

microbiology↗

bioRxiv2022

Single-nuclei transcriptomics of dog hippocampus reveals the distinct cellular mechanism of domestication

The process of dog domestication leads to dramatic differences in behavioral traits compared to grey wolves. A class of putative positively selected genes is related to learning and memory, for instance, long-term potentiation and long-term depression. In this study, we constructed a single-nuclei transcriptomic atlas of the dog hippocampus to illustrate its cell types, cell lineage, and molecular features. Using the transcriptomes of 105,057 single-nuclei from the hippocampus of a Beagle dog, we identified 26 cell clusters and a putative trajectory of oligodendrocyte development. Comparative analysis revealed a significant convergence between dog differentially expressed genes (DEGs) and putative positively selected genes (PSGs). 40 putative PSGs were DEGs in the glutamatergic neurons, especially in the cluster 14, which is related to the regulation of nervous system development. In summary, this study provided a blueprint to understand the cellular mechanism of dog domestication.

evolutionary biology↗

bioRxiv2022

Transcriptome Analysis of the Nematode Caenorhabditis elegans and Litoditis marina in a Chemically Defined Food Environment

Diets regulate animal development, reproduction, and lifespan. However, the underlying molecular mechanisms remain elusive. A chemically defined CeMM diet attenuates development and promotes longevity of C. elegans, but whether it impact on other nematodes is unknown. Here, we studied the effect of the CeMM diet on the development and longevity of the marine nematode Litoditis marina, which belongs to the same family as C. elegans. We further investigated genome-wide transcriptional responses to CeMM and OP50 diet for both nematodes, respectively. We observed that the CeMM diet attenuated L. marina development but did not extend its lifespan. We found that many of the FOXO DAF-16 target genes, lysosome and xenobiotic metabolism related genes were significantly increased on the CeMM, which might contribute to the lifespan extension of C. elegans. Notably, we found that the expression of lysosome and xenobiotic metabolism pathway genes was significantly down regulated in L. marina on CeMM, which might explain why the CeMM diet could not promote the lifespan of L. marina compared to bacterial feeding. Additionally, down-regulation of several RNA transcription and protein generation and related processes genes might not only be involved in extending longevity but also contribute to attenuating development of C. elegans on CeMM, while down-regulation of unsaturated fatty acids synthesis genes might contribute to slow down the growth of L. marina on CeMM. Further genetic analysis of candidate gene(s) of longevity and development in C. elegans and L. marina will provide the molecular mechanisms underlying how diets regulate animal physiology and health in the context of global climate change with variable nutritional environments.

zoology↗

bioRxiv2022

DNA uptake by cell wall-deficient bacteria reveals a putative ancient macromolecule uptake mechanism

Horizontal gene transfer in bacteria is widely believed to occur via three main mechanisms: conjugation, transduction and transformation. These mechanisms facilitate the passage of DNA across the protective cell wall using sophisticated machinery. We present here a new mechanism of DNA uptake that is independent of canonical DNA uptake machineries and is used by bacteria that live without a cell wall. We show that the cell wall-deficient bacteria engulf extracellular material, whereby intracellular vesicles are formed, and DNA is internalized. This mechanism is not specific to DNA, and allows uptake of other macromolecules and even 125 nm lipid nanoparticles (LNPs). Uptake was prevented by molecules known to inhibit eukaryotic endocytosis, suggesting this to be an energy-dependent process. Given that cell wall-deficient bacteria are considered a model for early life forms, our work provides a possible mechanism for primordial cells to acquire new genetic material or food before invention of the bacterial cell wall.

microbiology↗

bioRxiv2022

Type IV pili is a critical virulence factor in clinical isolates of Paenibacillus thiaminolyticus

Hydrocephalus, the leading indication for childhood neurosurgery worldwide, is particularly prevalent in low-and-middle-income countries (LMICs). Hydrocephalus preceded by an infection, or postinfectious hydrocephalus (PIH), accounts for up to 60% of hydrocephalus in LMICs. Since many children with hydrocephalus suffer poor long-term outcomes despite surgical intervention, prevention of hydrocephalus remains paramount. Our previous studies implicated a novel bacterial pathogen, Paenibacillus thiaminolyticus, as a contributor to PIH in Uganda. Here we report the isolation of three P. thiaminolyticus strains, Mbale, Mbale2, and Mbale3, from patients with PIH and the demonstration that the three clinical isolates exhibit virulence in mice while P. thiaminolyticus type strain, B-4156, does not. We constructed complete genome assemblies of the clinical isolates as well as the reference strain and performed comparative genomics and proteomics analyses to identify potential virulence factors. One candidate virulence factor is a cluster of genes carried on a mobile genetic element that encodes a type IV pilus and is present in all three PIH patient strains but absent in the type strain. Proteomic and transcriptomic data confirmed the expression of this cluster of genes in the Mbale strain, while CRISPR-mediated deletion of the gene cluster substantially reduced the virulence of this strain. Our comparative proteogenomic analysis also identified various antibiotic resistance loci in the virulent strains. These results provide insight into the mechanism of virulence of Paenibacillus thiaminolyticus and suggest avenues for the diagnosis and treatment of this novel bacterial pathogen. Author SummaryPostinfectious hydrocephalus (PIH), a devastating sequela of neonatal infection, is associated with increased childhood mortality and morbidity. Paenibacillus thiaminolyticus was recently identified as the dominant organism highly associated with PIH in an African cohort. Our whole-genome sequencing, RNA sequencing and proteomics of three clinical isolates and a type strain in combination with CRISPR editing has revealed the type IV pili (T4P), encoded in a mobile genetic element, as a critical virulence factor for P. thiaminolyticus infection. Given the widespread presence of T4P in pathogens, the presence of T4P operon could serve as an important diagnostic and therapeutic target in P. thiaminolyticus and related bacteria.

microbiology↗

bioRxiv2022

Found in translation: Microproteins are a new class of potential host cell impurity in mAb drug products

Chinese hamster ovary (CHO) cells are used to produce almost 90% of therapeutic monoclonal antibodies (mAbs). The annotation of non-canonical translation events in these cellular factories remains incomplete, limiting not only our ability to study CHO cell biology but also detect host cell protein (HCP) contaminants in the final mAb drug product. We utilised ribosome footprint profiling (Ribo-seq) to identify novel open reading frames (ORFs) including N-terminal extensions and thousands of short ORFs (sORFs) predicted to encode microproteins. Mass spectrometry-based HCP analysis of four commercial mAb drug products using the extended protein sequence database revealed the presence of microprotein impurities for the first time. We also show that microprotein abundance varies with growth phase and can be affected by the cell culture environment. In addition, our work provides a vital resource to facilitate future studies of non-canonical translation as well as the regulation of protein synthesis in CHO cell lines.

genomics↗

bioRxiv2022

Improving the Identification Coverage of Protein Interactome by Enhancing the Click Chemistry-based Cross-linking Enrichment Efficiency

Chemical cross-linking coupled with mass spectrometry has emerged as a powerful strategy which enables global profiling of protein interactome with direct interaction interfaces in complex biological systems. The alkyne-tagged enrichable cross-linkers are preferred to improve the coverage of low-abundance cross-linked peptides, combined with click chemistry for biotin conjugation to allow the cross-linked peptides enrichment. However, a systematic evaluation on the efficiency of click approaches (protein-based or peptide-based) and diverse cleavable click chemistry ligands (acid, reduction, photo) for cross-linked peptides enrichment and release is lacking. Herein, together with in vivo chemical cross-linking by alkyne-tagged cross-linker, we explored the click chemistry-based enrichment approaches on protein and peptide level with three cleavable click chemistry ligands, respectively. By comparison, the approach of protein-based click chemistry conjugation with acid-cleavable tag was demonstrated to permit the most cross-linked peptides identification. The advancement of this strategy enhanced the proteome-wide cross-linking analysis, permitting a detection of 5,017 protein-protein interactions among 1,909 proteins across all subcellular compartments with wide abundance distribution in cell. Therefore, all these results demonstrated a guideline value of our work for efficient cross-linked peptides enrichment, thus facilitated the in-depth profiling of protein interactome for functional analysis. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=67 SRC="FIGDIR/small/475819v1_ufig1.gif" ALT="Figure 1"> View larger version (17K): org.highwire.dtl.DTLVardef@1a7ae4forg.highwire.dtl.DTLVardef@1a4d6d2org.highwire.dtl.DTLVardef@113a0eborg.highwire.dtl.DTLVardef@1a9c193_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2022

RNF213 reshapes vascular transcriptome and epigenome; A potential role of Alternative splicing in Moyamoya pathology

RationaleMoyamoya disease (MMD) is a rare cerebrovascular occlusive disease that affects Asian population more often. The pathogenesis of MMD is related to mutation in RNF213 gene. However, why, and how RNF213 mutation leads to MMD is still not fully understood. ObjectiveAnalyze the impact of RNF213 loss of function on vascular cells and the observed changes correlate with MMD. Methods and resultsRNF213 KD was conducted in HUVEC and vascular smooth muscle cells (vSMCs). First, HUVEC cells showed alteration of angiogenesis, migration under LPS stimulation, Leukocyte endothelial transmigration, and endothelial-to-vSMCs communication. Transcriptome analysis revealed downregulation of genes regulating cell division and mitosis. Interestingly, Alternative splicing (AS) analysis revealed hundreds of AS events to occur after RNF213 KD in various types of AS. The alternatively spliced genes showed minimal overlap with transcriptome profiling. Pathway analysis revealed many processes and pathways to be regulated by AS events observed. Transcriptome profiling was also performed after LPS treatment and revealed the basis for increased sensitivity to LPS observed in our analysis. AS changes were also observed after LPS treatment in RNF213 KD HUVEC. Different types of AS showed different patterns of convergence or divergence in terms of the regulated pathways after LPS treatment. Finally, transcriptome and AS analysis was performed in vSMCs and showed various processes that impact vSMCs function and phenotype in RNF213 loss of function. ConclusionOur data provide a wealth of information on RNF213 gene function in vascular tissues and shed important light on different processes that contribute to MMD pathogenesis. Our results signify an immune-driven and hemodynamically linked process of MMD initiation and progression.

molecular biology↗

bioRxiv2022

Spatially resolved profiling of protein conformation and interactions by biocompatible chemical cross-linking in living cells

The protein structures and interactions that maintain and regulate cellular processes in different subcellular organelles are heterogeneous and dynamic. However, it remains challenging to characterize the subcellular specificity and translocation of protein complexes in terms of conformation and interactions. Herein, we developed a spatially resolved protein complex profiling approach by biocompatible chemical cross-linking in living cells (SPACX) to monitor the dynamics of protein conformation, interactions and translocation. The advancement of fast capturing protein complexes in the physiological state, coupled with efficient enrichment of the cross-linked peptides, ensured deep-coverage analysis of the protein interactome in living cells. By ensemble structure refinement with cross-linking restraints, subcellular-specific conformation heterogeneity was identified for PTEN. PTEN displayed a broader range of dynamic conformation changes on the dual specificity domains in the nucleus than in the cytoplasm. Moreover, based on conformational differences, different interacting assemblies involving 25 cytoplasm-exclusively and 18 nucleus-exclusively PTEN-interacting proteins were found to account for diverse biological functions. Upon ubiquitin-proteasome system (UPS) stress, the assembly of PTEN and its interacting partners changed obviously during translocation. We newly identified 36 PTEN-interacting proteins, which were found to be highly enriched in functional pathways closely related to cell apoptosis. Inspiringly, the interactions among PTEN isoforms and their interacting proteins were accessible by the determination of sequence-unique cross-linking interfaces for direct interactions. All these results indicate the promise of SPACX to elucidate the functional heterogeneity of proteins in individual subcellular sociology.

biochemistry↗