Biology subjects

Lu, W.

Publications and source records attributed to Lu, W..

At least 73 records · Page 4Linked to original sources

bioRxiv2024

Risk factors, prognostic factors, and nomograms for distant metastasis in patients with triple-negative breast cancer: a population-based study

BackgroundTriple-negative breast cancer (TNBC) is associated with a poor prognosis, higher invasiveness compared to other breast cancer variants, and limited treatment options. Due to the lack of conventional treatment methods, TNBC patients are prone to distant metastasis. The purpose of this study is to evaluate the clinical and pathological characteristics of TNBC patients, predict the impact of risk factors on their prognosis, and establish a predictive model to assist doctors in treating TNBC patients and improving their outcomes. MethodsWe conducted a retrospective study on patients with TNBC from 2010 to 2015 in the Surveillance, Epidemiology, and End Results (SEER) database. We gathered tumor characteristics and treatment data from these patients. Univariate and multivariate logistic regression analyses were employed to identify independent risk factors for distant metastasis in TNBC patients, while univariate and multivariate Cox proportional hazards regression analyses were utilized to determine independent prognostic factors for TNBC patients with distant metastasis. Based on these factors, a new nomogram was constructed, and patients were categorized into high and low-risk groups using the nomogram scores. The prognosis of the patients was analyzed using Kaplan-Meier (K-M) survival analysis. ResultsThis study enrolled a total of 16,959 patients with TNBC, among whom 730 (4.3%) had distant metastasis at the time of diagnosis. Independent risk factors for distant metastasis in TNBC patients encompass being married, having an invasive lobular carcinoma (ILC) pathology, advanced T stage, advanced N stage, no radiotherapy, no surgery, and tumor size exceeding 50mm. The area under the curve (AUCs) for the training set and validation set were 0.892 and 0.907, respectively. The independent prognostic factors for patients with distant metastasis were being unmarried (P < 0.001), not undergoing surgery (P < 0.001), and not receiving chemotherapy (P < 0.001). The AUCs for the training set at 12, 36, and 60 months were 0.751, 0.709, and 0.691, respectively, while the AUCs for the validation set at 12, 36, and 60 months were 0.690, 0.690, and 0.732, respectively. The AUCs for the extended test set at 12, 36, and 60 months were 0.741, 0.727, and 0.718 respectively. The results of the calibration curve, decision curve analysis (DCA), and K-M survival curve confirm that both nomograms can accurately predict the occurrence of distant metastasis and prognosis in TNBC patients. ConclusionWe constructed two nomograms to predict distant metastasis risk in TNBC patients and prognosis in patients with distant metastasis. Our findings suggest that they could serve as effective predictive tools and may assist in clinical decision-making.

cancer biology↗

bioRxiv2024

Coordinated translational control of multiple immune checkpoints by the integrated stress response pathway in lung cancer

The integrated stress response (ISR) is an adaptive pathway hijacked by cancer cells to survive cellular stresses in the tumor microenvironment. ISR activation potently induces Programmed Death Ligand 1 (PD-L1), leading to suppression of anti-tumor immunity. Here we sought to uncover additional immune checkpoint proteins regulated by the ISR to elucidate mechanisms of tumor immune escape. We show that CD155 and PD-L1 are coordinately induced by the ISR, enhancing translation of both immune checkpoint proteins through bypass of inhibitory upstream open reading frames (uORFs) in their 5' UTRs. Analysis of primary human lung tumors identifies a significant correlation between PD-L1 and CD155 expression. ISR activation accelerates tumorigenesis and inhibits T cell function, effects that can be overcome by combining PD-1 blockade with the ISR inhibitor ISRIB. These studies uncover a novel mechanism by which two immune checkpoint proteins are coordinately regulated and suggest a new therapeutic strategy for lung cancer patients. Statement of SignificanceThis study uncovers a novel mechanism for the coordinated translational regulation of the PD- L1/PD1 and CD155/TIGIT immune checkpoint pathways and highlights the ISR as a therapeutic vulnerability for lung cancer. Inhibition of the ISR pathway bolsters PD-1 blockade, potentially unveiling a new therapeutic strategy for lung cancer patients.

cancer biology↗

bioRxiv2024

Effect heterogeneity reveals complex pleiotropic effects of rare coding variants

Recent expansion of large-scale biobank resources has enabled rare-variant association studies (RVAS) and systematic investigation of rare variant pleiotropy across thousands of phenotypes simultaneously. However, existing statistical frameworks for dissecting pleiotropy were largely developed for common variants and are not well suited to gene-based rare variant signals, limiting the interpretation of cross-phenotype associations. Here, we develop ALLSPICE, a likelihood-based method that tests whether rare variant effects in genes exhibiting cross-phenotype associations are proportional or heterogeneous across continuous traits while accounting for phenotypic correlation using summary statistics. ALLSPICE is well-calibrated in simulations and is implemented as an R package for scalable analysis of gene-level rare-variant burden associations. We applied ALLSPICE to RVAS of 359 continuous traits in the UK Biobank and identified 124 significant heterogeneous events among 11,810 pairs of gene-trait associations. By identifying effect heterogeneity within genes associated with multiple phenotypes, ALLSPICE clarifies shared and heterogeneous rare variant architectures underlying cross-phenotype associations and provides insight into rare variant pleiotropy.

genetics↗

bioRxiv2024

'Mitotic' kinesin-5 is a dynamic brake for axonal growth

During neuronal development, neurons undergo significant microtubule reorganization to shape axons and dendrites, establishing the framework for efficient wiring of the nervous system. Previous studies from our laboratory demonstrated the key role of kinesin-1 in driving microtubule-microtubule sliding, which provides the mechanical forces necessary for early axon outgrowth and regeneration in Drosophila melanogaster. In this study, we reveal the critical role of kinesin-5, a mitotic motor, in modulating the development of postmitotic neurons. Kinesin-5, a conserved homotetrameric motor, typically functions in mitosis by sliding antiparallel microtubules apart in the spindle. Here, we demonstrate that the Drosophila kinesin-5 homolog, Klp61F, is expressed in larval brain neurons, with high levels in ventral nerve cord (VNC) neurons. Knockdown of Klp61F using a pan-neuronal driver leads to severe locomotion defects and complete lethality in adult flies, mainly due to the absence of kinesin-5 in VNC motor neurons during early larval development. Klp61F depletion results in significant axon growth defects, both in cultured and in vivo neurons. By imaging individual microtubules, we observe a significant increase in microtubule motility, and excessive penetration of microtubules into the axon growth cone in Klp61F-depleted neurons. Adult lethality and axon growth defects are fully rescued by a chimeric human-Drosophila kinesin-5 motor, which accumulates at the axon tips, suggesting a conserved role of kinesin-5 in neuronal development. Altogether, our findings show that at the growth cone, kinesin-5 acts as a brake on kinesin-1-driven microtubule sliding, preventing premature microtubule entry into the growth cone. This regulatory role of kinesin-5 is essential for precise axon pathfinding during nervous system development.

cell biology↗

bioRxiv2024

EB-SUN, a New Microtubule Plus-End Tracking Protein in Drosophila.

Microtubule (MT) regulation is essential for oocyte development. In Drosophila, MT stability, polarity, abundance, and orientation undergo dynamic changes across developmental stages. In our effort to identify novel microtubule-associated proteins (MAPs) that regulate MTs in the Drosophila ovary, we identified a previously uncharacterized gene, CG18190, encoding a novel MT end-binding (EB) protein, which we propose to name EB-SUN. We show that EB-SUN colocalizes with EB1 at growing microtubule plus-ends in Drosophila S2 cells. Tissue-specific and developmental expression profiles from Paralog Explorer reveal that EB-SUN is predominantly expressed in the ovary and early embryos, while EB1 is ubiquitously expressed. Furthermore, as early as oocyte determination, EB-SUN comets are highly concentrated in oocytes during oogenesis. EB-SUN knockout (KO) results in a decrease in MT density at the onset of mid-oogenesis (Stage 7) and delays oocyte growth during late mid-oogenesis (Stage 9). Combining EB-SUN KO with EB1 knockdown (KD) in germ cells significantly further reduced MT density at Stage 7. Notably, all eggs from EB-SUN KO/EB1 KD females fail to hatch, unlike single gene depletion, suggesting a functional redundancy between these two EB proteins during embryogenesis. Our findings indicate that EB-SUN and EB1 play distinct roles during early embryogenesis. Significance StatementO_LIOur study identified a novel microtubule end-binding (EB) protein, which is highly expressed in the Drosophila ovary, and we propose to name it EB-SUN. C_LIO_LIWe demonstrate the functional redundancy of EB-SUN and EB1 during oocyte development, while highlighting their distinct roles in early embryogenesis. C_LIO_LIGiven the high conservation of MT-dependent mechanisms in oogenesis and EB proteins across species, these findings provide insights into the differential regulation and tissue-specific functions of the EB family, using Drosophila as a model system, potentially benefiting future research on MT regulation in higher organisms. C_LI

cell biology↗

bioRxiv2024

Spatiotemporally-resolved mapping of extracellular proteomes via in vivo-compatible TyroID

Extracellular proteins play pivotal roles in both intracellular signaling and intercellular communications in health and disease. While recent advancements in proximity labeling (PL) methods, such as peroxidase- and photocatalyst-based approaches, have facilitated the resolution of extracellular proteomes, their in vivo compatibility remains limited. Here, we report TyroID, an in vivo-compatible PL method for unbiased mapping of extracellular proteins with high spatiotemporal resolution. TyroID employs plant- and bacteria-derived tyrosinases to produce reactive o-quinone intermediates, enabling the labeling of multiple residues on endogenous proteins with bioorthogonal handles, thereby allowing for their identification via chemical proteomics. We validate TyroIDs specificity by mapping extracellular proteomes and HER2-neighboring proteins using nanobody-directed recombinant tyrosinases. Demonstrating its superiority over other PL methods, TyroID enables in vivo mapping of extracellular proteomes, including mapping HER2-proximal proteins in tumor xenografts, quantifying the turnover of plasma proteins and labeling hippocampal-specific proteomes in live mouse brains. TyroID emerges as a potent tool for investigating protein localization and molecular interactions within living organisms.

biochemistry↗

bioRxiv2024

Unsilenced inhibitory cortical ensemble gates remote memory retrieval

Acquired information can be consolidated to remote memory for storage but persists in a dormant state until its retrieval. However, it remains unknown how dormant memory is reactivated. Using a combination of simultaneous two-photon calcium imaging and holographic optogenetics in the anterior cingulate cortex (ACC) in vivo, we discover a subset of GABAergic neurons that are specifically associated with dormant memory retrieval. These interneurons display persistent activity and inter-neuronal synchronization at the remote memory stage. In the absence of natural contextual cues, directly activating these interneurons reliably recalls cortical ensembles relevant to remote memory retrieval with context specificity. Conversely, targeted volumetric inactivation of these interneurons suppresses context-induced memory retrieval. Our results reveal an unexpected role of unsilenced inhibitory cortical ensembles in causally gating the retrievability of dormant remote memory.

neuroscience↗

bioRxiv2024

Sulfur Amino Acid Restriction Enhances Exercise Capacity in Mice by Boosting Fat Oxidation

Dietary restriction of the sulfur-containing amino acids methionine and cysteine (SAAR) improves body composition, enhances insulin sensitivity, and extends lifespan; benefits seen also with endurance exercise. Yet, the impact of SAAR on skeletal muscle remains largely unexplored. Here we demonstrate that one week of SAAR in sedentary, young, male mice increases endurance exercise capacity. Indirect calorimetry showed that SAAR increased lipid oxidation at rest and delayed the onset of carbohydrate utilization during exercise. Transcriptomic analysis revealed increased expression of genes involved in fatty acid catabolism especially in glycolytic muscle following SAAR. These findings were functionally supported by increased fatty acid circulatory turnover flux and muscle {beta}-oxidation. Reducing lipid uptake from circulation through endothelial cell (EC)-specific CD36 deletion attenuated the running phenotype. Mechanistically, VEGF-signaling inhibition prevented exercise increases following SAAR, without affecting angiogenesis, implicating noncanonical VEGF signaling and EC CD36-dependent fatty acid transport in regulating exercise capacity by influencing muscle substrate availability.

physiology↗

bioRxiv2024

Retinal microglial cells increase expression and release of IL-1β when exposed to ATP

Cytokine IL-1{beta} is an early component of inflammatory cascades, with both priming and activation steps required before IL-1{beta} release. Here, the P2X7 receptor (P2X7R) for ATP was shown to both prime and release IL-1{beta} from retinal microglial cells. Isolated retinal microglial cells increased expression of Il1b when stimulated with endogenous receptor agonist extracellular ATP; ATP also rapidly downregulated expression of microglial markers Tmem119 and Cd206. Changes to all three genes were reduced by specific P2X7R antagonist A839977, implicating the P2X7R. Microglial cells expressed the P2X7R on ramifications and responded to receptor agonist BzATP with robust and rapid rises in intracellular Ca2+. BzATP increased expression of IL-1{beta} protein colocalizing with CX3CR1-GFP in retinal wholemounts consistent with microglial cells. ATP also triggered release of IL-1{beta} from isolated retinal microglia into the bath; release was inhibited by A839977 and induced by BzATP, supporting a role for the P2X7R in release as well as priming. The IL-1{beta} release triggered by ATP was substantially greater from microglial cells compared to astrocytes from the optic nerve head region. Il1b expression was increased by a transient rise in intraocular pressure and Il1b levels remained elevated 10 days after a single IOP elevation. In summary, this study suggests the P2X7 receptor can both prime IL-1{beta} levels in microglial cells and trigger its release. The P2Y12R was previously identified as a chemoattractant for retinal microglia, suggesting the recruitment of the cells towards the source of released extracellular ATP could position microglia for P2X7R receptor, enabling both priming and release of IL-1{beta}.

neuroscience↗

bioRxiv2024

Fluorescent identification of axons, dendrites and soma of neuronal retinal ganglion cells with a genetic marker as a tool for facilitating the study of neurodegeneration

This study characterizes a fluorescent Slc17a6-tdTomato neuronal reporter mouse line offering strong labeling in axons throughout the optic nerve, dendrites and soma in 99% of retinal ganglion cells (RGCs). The model facilitates neuronal assessment ex vivo with wholemounts quantified to show neurodegeneration following optic nerve crush or elevated IOP as related to glaucoma, in vitro with robust Ca2+ responses to P2X7 receptor stimulation in neuronal cultures, and in vivo using a confocal scanning laser ophthalmoscope (cSLO). While the tdTomato signal showed strong overlap with RGC markers, BRN3A and RBPMS, there was no cross-labeling of displaced amacrine cells in the ganglion cell layer. Controls indicated no impact of Slc17a6-tdTomato expression on light-dependent neuronal function, as determined with a microelectrode array (MEA), or on structure, as measured with optical coherence tomography (OCT). In summary, this novel neuronal reporter mouse model offers an effective means to increase the efficiency for real-time, specific visualization of retinal ganglion cells. It holds substantial promise for enhancing our understanding of RGC pathology in glaucoma and other diseases of the optic nerve, and could facilitate the screening of targeted therapeutic interventions for neurodegeneration. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=140 SRC="FIGDIR/small/599589v1_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@13db8dforg.highwire.dtl.DTLVardef@1345f10org.highwire.dtl.DTLVardef@14d34e1org.highwire.dtl.DTLVardef@1cacb87_HPS_FORMAT_FIGEXP M_FIG Graphical abstract. Fluorescent identification of axons, dendrites and soma of neuronal retinal ganglion cells with a genetic marker as a tool for facilitating the study of neurodegeneration. Puttipong Sripinun, Wennan Lu, Sergei Nikonov, Suhani Patel, Sarah Hennessy, Claire H. Mitchell*. This study delves into a new mouse model, featuring a fluorescent Slc17a6-tdTomato neuronal reporter. This model effectively labels axons in the optic nerve, as well as dendrites and soma in 99% of retinal ganglion cells (RGCs). This allows for both in vitro and in vivo assessment of neurodegeneration, offering a practical tool for real-time, precise visualization of RGCs, with potential applications in various fields of neuroscience and neurology. Created by Biorender.com. C_FIG

neuroscience↗

bioRxiv2024

Piezo1 and Piezo2 channels in retinal ganglion cells and the impact of Piezo1 stimulation on light-dependent neural activity

Piezo channels are associated with neuropathology in diseases like traumatic brain injury and glaucoma, but pathways linking tissue stretch to aberrant neural signaling remain unclear. The present study demonstrates that Piezo1 activation increases action potential frequency in response to light and the spontaneous dark signal from mouse retinal explants. Piezo1 stimulation was sufficient to increase cytoplasmic Ca2+ in soma and neurites, while stretch increased spiking activity in current clamp recordings from of isolated retinal ganglion cells (RGCs). Axon-marker beta-tubulin III colocalized with both Piezo1 and Piezo2 protein in the mouse optic nerve head, while RGC nuclear marker BRN3A colocalized with Piezo channels in the soma. Piezo1 was also present on GFAP-positive regions in the optic nerve head and colocalized with glutamine synthetase in the nerve fiber layer, suggesting expression in optic nerve head astrocytes and Muller glia end feet, respectively. Human RGCs from induced pluripotent stem cells also expressed Piezo1 and Piezo2 in soma and axons, while staining patterns in rats resembled those in mice. mRNA message for Piezo1 was greatest in the RPE/choroid tissue, while Piezo2 levels were highest in the optic nerve, with both channels also expressed in the retina. Increased expression of Piezo1 and Piezo2 occurred both 1 and 10 days after a single stretch in vivo; this increase suggests a potential role in rising sensitivity to repeated nerve stretch. In summary, Piezo1 and Piezo2 were detected in the soma and axons of RGCs, and stimulation affected the light-dependent output of RGCs. The rise in RGCs excitability induced by Piezo stimulation may have parallels to the early disease progression in models of glaucoma and other retinal degenerations. HighlightsO_LIActivation of Piezo1 excites retinal ganglion cells, paralleling the early neurodegenerative progression in glaucoma mouse models and retinal degeneration. C_LIO_LIPiezo1 and Piezo2 were expressed in axons and soma of retinal ganglion cells in mice, rats, and human iPSC-RGCs. C_LIO_LIFunctional assays confirmed Piezo1 in soma and neurites of neurons. C_LIO_LISustained elevation of Piezo1 and Piezo2 occurred after a single transient stretch may enhance damage from repeated traumatic nerve injury. C_LI O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=180 SRC="FIGDIR/small/599602v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@7bb1a0org.highwire.dtl.DTLVardef@cab0dcorg.highwire.dtl.DTLVardef@121690eorg.highwire.dtl.DTLVardef@7848fa_HPS_FORMAT_FIGEXP M_FIG Graphical abstract Piezo1 and Piezo2 channels in retinal ganglion cells and the impact of Piezo1 stimulation on light-dependent neural activity. Puttipong Sripinun, Lily P. See, Sergei Nikonov, Venkata Ramana Murthy Chavali, Vrathasha Vrathasha, Jie He, Joan M. OBrien, Jingsheng Xia, Wennan Lu, Claire H. Mitchell*. Activation of Piezo channels through mechanical or pharmacological stimulation leads to an influx of Ca2+ and other cations into RGCs, depolarizing the membrane and increasing the action potential frequency to modulate the visual signal. Created with Biorender.com C_FIG

neuroscience↗

bioRxiv2024

Digital pathology assessment of kidney glomerular filtration barrier ultrastructure in an animal model of podocytopathy

BackgroundTransmission electron microscopy (TEM) images can visualize kidney glomerular filtration barrier ultrastructure, including the glomerular basement membrane (GBM) and podocyte foot processes (PFP). Podocytopathy is associated with glomerular filtration barrier morphological changes observed experimentally and clinically by measuring GBM or PFP width. However, these measurements are currently performed manually. This limits research on podocytopathy disease mechanisms and therapeutics due to labor intensiveness and inter-operator variability. MethodsWe developed a deep learning-based digital pathology computational method to measure GBM and PFP width in TEM images from the kidneys of Integrin-Linked Kinase (ILK) podocyte-specific conditional knockout (cKO) mouse, an animal model of podocytopathy, compared to wild-type (WT) control mouse. We obtained TEM images from WT and ILK cKO littermate mice at 4 weeks old. Our automated method was composed of two stages: a U-Net model for GBM segmentation, followed by an image processing algorithm for GBM and PFP width measurement. We evaluated its performance with a 4-fold cross-validation study on WT and ILK cKO mouse kidney pairs. ResultsMean (95% confidence interval) GBM segmentation accuracy, calculated as Jaccard index, was 0.73 (0.70-0.76) for WT and 0.85 (0.83-0.87) for ILK cKO TEM images. Automated and manual GBM width measurements were similar for both WT (p=0.49) and ILK cKO (p=0.06) specimens. While automated and manual PFP width measurements were similar for WT (p=0.89), they differed for ILK cKO (p<0.05) specimens. WT and ILK cKO specimens were morphologically distinguishable by manual GBM (p<0.05) and PFP (p<0.05) width measurements. This phenotypic difference was reflected in the automated GBM (p<0.05) more than PFP (p=0.06) widths. ConclusionsThese results suggest that certain automated measurements enabled via deep learning-based digital pathology tools could distinguish healthy kidneys from those with podocytopathy. Our proposed method provides high-throughput, objective morphological analysis and could facilitate podocytopathy research and translate into clinical diagnosis. Key pointsO_LIWe leveraged U-Net architecture in an algorithm to measure the widths of glomerular basement membrane and podocyte foot processes. C_LIO_LIDeep learning-based automated measurement of glomerular filtration barrier morphology has promise in podocytopathy research and diagnosis. C_LI

pathology↗

bioRxiv2024

Short heat shock factor A2 confers heat sensitivity in Arabidopsis: Insights into heat resistance and growth balance

Cells prevent heat damage through the highly conserved canonical heat stress response (HSR), where heat shock factors (HSFs) bind heat shock elements (HSEs) to activate heat shock proteins (HSPs). Plants generate short HSFs (S-HSFs) derived from HSF splicing variants, yet their functions remain poorly understood. While an enhanced canonical HSR confers thermotolerance, its hyperactivation inhibits plant growth. How plants prevent this hyperactivation to ensure proper growth remains unknown. Here, we report that Arabidopsis S-HsfA2, S-HsfA4c, and S-HsfB1 confer sensitivity to extreme heat (45 {degrees}C) and constitute new HSF types featuring a unique truncated DNA-binding domain (tDBD). This tDBD binds a new heat-regulated element (HRE), which confers minimal promoter heat-responsiveness and exhibits heat stress sensing and transmission patterns. Using S-HsfA2, we investigated whether and how S-HSFs prevent canonical HSR hyperactivation. HSP17.6B, a common direct target of HsfA2 and S-HsfA2, confers thermotolerance; however, its overexpression partially causes HSR hyperactivation. Moreover, HRE-HRE-like and HSE elements mediate the HSP17.6B promoters heat response. We further show S-HsfA2 alleviates hyperactivation via two mechanisms: 1) S-HsfA2 negatively regulates HSP17.6B via the HRE-HRE-like element, establishing a noncanonical HSR (S-HsfA2-HRE-HSP17.6B) that antagonistically represses HsfA2-activated HSP17.6B expression. 2) S-HsfA2 binds the HsfA2 DBD, preventing HsfA2 from binding HSEs and thereby attenuating HsfA2-activated HSP17.6B promoter activity. Overall, our findings highlight the essential role of S-HsfA2 in preventing hyperactivation of plant heat tolerance to maintain proper growth.

plant biology↗

bioRxiv2024

Discovery of Potent Degraders of the Dengue Virus Envelope Protein

Targeted protein degradation has been widely adopted as a new approach to eliminate both established and previously recalcitrant therapeutic targets. Here we report the development of small molecule degraders of the envelope (E) protein of dengue virus. We developed two classes of bivalent E-degraders, linking two previously reported E-binding small molecules, GNF-2 and CVM-2-12-2, to a glutarimide-based recruiter of the CRL4CRBN ligase to effect proteosome-mediated degradation of the E protein. ZXH-2-107 (based on GNF-2) is an E degrader with ABL inhibition while ZXH-8-004 (based on CVM-2-12-2) is a selective and potent E-degrader. These two compounds provide proof-of-concept that difficult-to-drug targets such as a viral envelope protein can be effectively eliminated using a bivalent degrader and provide starting points for the future development of a new class antiviral drugs.

biochemistry↗

bioRxiv2024

AlphaFold3, a secret sauce for predicting mutational effects on protein-protein interactions

AlphaFold3 has set the new state-of-the-art in predicting protein-protein complex structures. However, the complete picture of biomolecular interactions cannot be fully captured by static structures alone. In the field of protein engineering and antibody discovery, the connection from structure to function is often mediated by binding energy. This work benchmarks AlphaFold3 against SKEMPI, a commonly used binding energy dataset. We demonstrate that AlphaFold3 learns unique information and synergizes with force field, profile-based, and other deep learning methods in predicting the mutational effects on protein-protein interactions. We hypothesize that AlphaFold3 captures a more global effect of mutations by learning a smoother energy landscape, but it lacks the modeling of full atomic details that are better addressed by force field methods, which possess a more rugged energy landscape. Integrating both approaches could be an interesting future direction. All of our benchmark results are openly available at https://github.com/luwei0917/AlphaFold3_PPI.

biophysics↗

bioRxiv2024

Engineered lysins to modulate human gut microbiome communities

The ability to precisely control the human gut microbiomes composition, metabolic networks, and host interactions would revolutionize medicine and how we treat human disease. Microbiome remodeling approaches such as probiotics, prebiotics, antibiotics, and fecal microbiota transplantation are global perturbations with consequences that are difficult to predict. Lysins are enzymes produced by bacteriophages and bacteria that have exceptional specificity to lyse bacterial cells and could be applied for targeted microbiome remodeling. In this work we demonstrate the ability of lysins to target diverse human gut commensal bacteria, how domain recombination can give rise to chimeric lysins with altered species preferences, and how engineered lysins can directly remodel the structure and function of synthetic human gut microbiomes. Lysins represent a general platform that can be rapidly tailored through protein engineering to control complex microbial ecosystems.

synthetic biology↗

bioRxiv2024

Genome-Wide Identification and Expression Pattern of the N-acetylserotonin deacetylase (ASDAC) Gene Family in Orchidaceae

Orchids are a kind of horticultural plant with highly ornamental and medical value. N-acetylserotonin deacetylase (ASDAC) is the only reverse enzyme of the melatonin biosynthesis pathway, and plays an important role in regulating the balance of melatonin. Melatonin as a multifunctional molecule, is typically involved in plant growth and development regulation, as well as abiotic stress tolerance. Here, we aimed at identifying ASDAC genes from the orchid genome to provide valuable information for further study of the role of melatonin in orchids. In this study, a total of 7 ASDAC genes were identified from the 7 orchid genome with one member in each species. The 7 orchid ASDACs have an HDAC functional domain, and cluster together with functionally confirmed OsHDAC10 and AtHDAC14, it shows that these members may have function of N-acetylserotonin deacetylase. Furthermore, based on the phylogenetic, motif, and gene structure analysis, the same clusters orchid ASDAC or ASDAC-like genes generally contained similar introns and motifs, suggesting the distribution pattern of exons/introns and motifs were strongly related to phylogeny on an evolutionary basis. Interestingly, homologous genes of OsHDAC10 and AtHDAC14 in Gastrodia elata have low homology and not cluster together with rice and A.thaliana ASDACs, showing that ASDAC gene family may lost in the holomycoheterotrophic orchids. The Ka/Ks ratios of ASDAC gene pairs from lower plant to higher plant were less than one, suggested that ASDAC genes have undergone purifying selection during the evolution process. Cis-acting element analysis results showed that the promoter regions of orchid ASDAC genes contained plant growth and development, phytohormone, and stress-responsive elements. Moreover, most orchid ASDACs were expressed in the roots, stems, leaves, flowers, and seeds. Combined cis-acting element and tissue expression analysis, indicating orchid ASDAC genes are involved in melatonin regulation of growth and development, as well as melatonin responding to various stresses in orchids. These findings of orchid ASDAC genes may provide valuable information for further study of the role of melatonin in orchids.

bioinformatics↗

bioRxiv2024

Covalent DNA-Encoded Library Workflow Drives Discovery of SARS-CoV-2 Non-structural Proteins Inhibitors

The global coronavirus disease 2019 (COVID-19) pandemic persists, with the ongoing mutation of the virus. Consequently, the development of inhibitors with diverse binding modes and mechanisms of action, along with the elucidation of novel binding sites is of paramount importance. The 3-chymotrypsin-like protease (3CLpro) and papain-like protease (PLpro) are two validated cysteine proteases that cleave the viral polyprotein and are essential for viral replication. In this study, we utilized covalent DNA-Encoded libraries (CoDELs) workflow to identify two series of triazine-based covalent inhibitors targeting 3CLpro and PLpro. Molecular docking facilitated the identification of optimization pathways, further refined through medicinal chemistry efforts, leading to the development of the non-peptide 3CLpro inhibitor LU9, which exhibited an IC50 value of 0.34 M, and crystal structure of LU10 revealed a unique binding mode within the active site. Additionally, the X-ray cocrystal structure of SARS-CoV-2 PLpro with XD5 uncovered a previously unexplored binding site, adjacent to the catalytic pocket, providing an opportunity for further development of PLpro inhibitors. Overall, these novel compounds serve as valuable chemical probes for target validation and represent promising drug candidates for the continued development of SARS-CoV-2 antivirals.

biochemistry↗