Biology subjects

Chen, M.

Publications and source records attributed to Chen, M..

33 records · Page 2Linked to original sources

bioRxiv2018

Single-step Enzymatic Glycoengineering for the Construction of Antibody-cell Conjugates

Employing live cells as therapeutics is a direction of future drug discovery. An easy and robust method to modify the surfaces of cells directly to incorporate novel functionalities is highly desirable. However, many current methods for cell-surface engineering interfere with cells endogenous properties. Here we report an enzymatic approach that enables the transfer of biomacromolecules, such as a full length IgG antibody, to the glycocalyx on the surfaces of live cells when the antibody is conjugated to the enzymes natural donor substrate GDP-fucose. This method is fast and biocompatible with little interference to cells endogenous functions. We applied this method to construct two antibody-cell conjugates (ACCs) using different immune cells, and the modified cells exhibited specific tumor targeting and resistance to inhibitory signals produced by tumor cells, respectively. Remarkably, Herceptin-NK-92MI conjugates exhibits enhanced activities to induce the lysis of HER2+ cancer cells both ex vivo and in a murine tumor model, indicating its potential for further development as a clinical candidate.

bioengineering↗

bioRxiv2018

Stimulus-responsive self-assembly of enzymatic fractal structures by computational design

Fractal topologies, which are statistically self-similar over multiple length scales, are pervasive in nature. The recurrence of patterns at increasing length scales in fractal-shaped branched objects, e.g., trees, lungs, and sponges, results in high effective surface areas, and provides key functional advantages, e.g., for molecular trapping and exchange. Mimicking these topologies in designed protein-based assemblies will provide access to novel classes of functional biomaterials for wide ranging applications. Here, we describe a modular, multi-scale computational design method for the reversible self-assembly of proteins into tunable supramolecular fractal-like topologies in response to phosphorylation. Computationally-guided atomic-resolution modeling of fusions of symmetric, oligomeric proteins with Src homology 2 (SH2) binding domain and its phosphorylatable ligand peptide was used to design iterative branching leading to fractal-like assembly formation by enzymes of the atrazine degradation pathway. Structural characterization using various microscopy techniques and Cryo-electron tomography revealed a variety of dendritic, hyperbranched, and sponge-like topologies which are self-similar over three decades ([~]10nm-10m) of length scale, in agreement with models from multi-scale computational simulations. We demonstrate control over mesoscale topology (by linker design), formation dynamics, and functional enhancements due to dynamic multi-component assemblies constructed with three atrazine degradation pathway enzymes. The described design method should enable the construction of a variety of novel, spatiotemporally responsive catalytic biomaterials featuring fractal topologies.

biophysics↗

bioRxiv2018

The Specificity of IQGAP1 Toward the PI3K-Akt Pathway is Dependent on the IQ3 motif

Epidermal growth factor receptor (EGFR) and its downstream phosphatidylinositol 3-kinase (PI3K) pathway are commonly deregulated in many cancers including head and neck cancer (HNC). Recently, we have shown that the IQ motif-containing GTPase-activating protein 1 (IQGAP1) provides a molecular platform to scaffold all the components from the PI3K-Akt pathway and results in the sequential generation of phosphatidylinositol-3,4,5-triphosphate (PI3,4,5P3). This makes the IQGAP1-PI3K scaffold a promising therapeutic target. In addition to the PI3K-Akt pathway, IQGAP1 also scaffolds the Ras-ERK pathway. To identify an IQGAP1 mutant that specifically loses IQGAP1-PI3K signaling but not other functions, we have focused on the IQ3 motif since this region binds with both the PIPK1 and PI3K enzymes and a short peptide derived from this sequence blocks binding and PI3K signaling. An IQ3 deletion mutant ({Delta}IQ3) in IQGAP1 was functionally compared with wild-type (WT) IQGAP1. We found that the IQ3 domain specifically mediates the PI3K-Akt pathway but does not regulate the Ras-ERK pathway. The IQ3 deletion mutant lost interactions with PI3K-Akt components but retained its binding of ERK pathway components and cell surface receptors, EGFR and integrins. In addition, the IQ3 deletion mutant lost regulation of cell migration. Consistently, the IQ3 motif derived peptide blocked Akt activation and led to the blockage of invasion mediated by EGFR organized into an integrin complex by syndecan-4. Taken together, this work has defined the IQGAP1 IQ3 motif as a specific target sequence for the scaffolding of the PI3K-AKT pathway.

cell biology↗

bioRxiv2018

A novel long noncoding RNA LINC00844 regulates prostate cancer cell migration and invasion through androgen receptor signaling

The majority of the human genome is transcribed, yielding a rich repository of non-coding transcripts that are involved in a myriad of biological processes including cancer. However, how non-coding transcripts such as long non-coding RNAs (lncRNAs) function in prostate cancer is still unclear. In this study, we have identified a novel set of clinically relevant androgen-regulated lncRNAs in prostate cancer. Among this group, we showed LINC00844 is a direct androgen-regulated target that is actively transcribed in androgen receptor (AR)-dependent prostate cancer cells. The expression of LINC00844 is higher in normal prostate compared to malignant and metastatic prostate cancer samples and patients with low expression demonstrate poor prognosis and significantly increased biochemical recurrence, suggesting LINC00844 may function in suppressing tumor progression and metastasis. Indeed, in-vitro loss-of-function studies revealed that LINC00844 prevents prostate cancer cell migration and invasion. Moreover, findings from gene expression analysis indicated that LINC00844 functions in trans, affecting global androgen-regulated gene transcription. Mechanistically, we provide evidence to show LINC00844 is important in facilitating AR binding to the chromatin. Finally, we demonstrated LINC00844 mediates its phenotypic effects in part by activating the expression of NDRG1, a crucial cancer metastasis suppressor. Collectively, our findings suggest LINC00844 is a novel coregulator of AR that plays a central role in the androgen transcriptional network and the development and progression of prostate cancer.

cancer biology↗

bioRxiv2018

Aged blood inhibits hippocampal neurogenesis and activates microglia through VCAM1 at the blood-brain barrier

An aged circulatory environment can promote brain dysfunction and we hypothesized that the blood-brain barrier (BBB) mediates at least some of these effects. We observe brain endothelial cells (BECs) in the aged mouse hippocampus express an inflammatory transcriptional profile with focal upregulation of Vascular Cell Adhesion Molecule 1 (VCAM1), a protein that facilitates vascular-immune cell interactions. Concomitantly, the shed, soluble form of VCAM1 is prominently increased in the aged circulation of humans and mice, and aged plasma is sufficient to increase VCAM1 expression in cultured BECs and young mouse hippocampi. Systemic anti-VCAM1 antibody or genetic ablation of VCAM1 in BECs counteracts the detrimental effects of aged plasma on young brains and reverses aging aspects in old mouse brains. Thus, VCAM1 is a negative regulator of adult neurogenesis and inducer of microglial reactivity, establishing VCAM1 and the luminal side of the BBB as possible targets to treat age-related neurodegeneration.

neuroscience↗

bioRxiv2017

Dynamics of a Form-Fitting Protein in a Nanopore: Avidin in ClyA

We probe the molecular dynamics of a protein, avidin, as it is captured and trapped in a nanopore, ClyA, with time-resolved single-molecule electrical conductance measurements, and we present a method for visualizing this process from the data. The case of avidin in ClyA has rich time-dependent conductance spectra of discrete levels that correlate with different configurations of the protein in the pore. One is very long-lasting, stable and noise-free, and portends the use of this system as a platform for more general studies of proteins and other molecules, where avidin acts as a shuttle that ferries analytes into the pore for probing. We demonstrate this by the sensitive detection of a biotin molecule attached to avidin captured by the pore. We also present an approach to determining the nanopore size based on a 3D printed model of the pore.

biophysics↗

bioRxiv2017

Genomic MET amplification occurs early in NF1-related malignant peripheral nerve sheath tumor (MPNST) progression and is a potent therapeutic target

Malignant Peripheral Nerve Sheath Tumors (MPNSTs) are highly resistant sarcomas that occur in up to 13% of individuals with Neurofibromatosis Type 1 (NF1). Genomic analysis of longitudinally collected tumor samples in a case of MPNST disease progression revealed early hemizygous microdeletions in NF1 and TP53, with concomitant amplifications of MET, HGF, and EGFR. To examine the role of MET in MPNST progression, we developed mice with enhanced MET expression and NF1 ablation (NF1fl/KO;lox-stop-loxMETtg/+;Plp-creERTtg/+; referred to as NF1-MET). NF1-MET mice express a robust MPNST phenotype in the absence of additional mutations. A comparison of NF1-MET MPSNTs with MPNSTs derived from NF1KO/+;p53R172H;Plp-creERTtg/+ (NF1-P53) and NF1KO/+;Plp-creERTtg/+ (NF1) mice revealed unique Met, Ras, and PI3K signaling patterns. To investigate the therapeutic potential of MET inhibition among tumorgrafts derived from the respective MPNST models, we tested the highly selective MET inhibitor, capmatinib. NF1-MET MPNSTs were uniformly sensitive to MET inhibition whereas only a small subset of NF1-P53 and NF1 MPNSTs were inhibited. These results confirm that MET activation is sufficient for Schwann cell dedifferentiation into MPNSTs in the context of NF1 deficiency. RAS-MET signal interactions may be an important driver of MPSNT disease progression.

cancer biology↗

bioRxiv2017

Bringing plants and soils to life through a simple role-playing activity

Interactions are at the core of many ecological and evolutionary forces in nature. Plant-soil interactions provide a rich example of the interconnectedness of living systems, but are hidden from everyday view and overshadowed in the classroom by more popular teaching examples involving animals, reptiles, or invertebrates. To highlight the importance and relevance of plant-soil relationships, we devised a simple role-playing activity suitable for college students. Specifically, the activity simulates how feedbacks between plants and soil environments influence plant species abundance and community richness. With this activity, students will gain a better understanding of these prolific, but overlooked, forms of biological interactions that impact the diversity and functioning of ecosystems.

scientific communication and education↗

bioRxiv2017

Sugar metabolism changes in response to the ultraviolet B irradiation of peach (Prunus persica L.)

The protected cultivation of peach (Prunuspersica L.) trees is more economical and efficient than traditional cultivation, resulting in increased farmers incomes, but the peach sugar contents are lower than in open planting. In the greenhouse, a high-sugar variety of peach Lumi 1 was irradiated with 1.44 KJ{middle dot}m-2.d-1 intensity ultraviolet B radiation. The soluble sugar contents in fruit, peel and leaf were quantified using liquid chromatography. Overall, sucrose and sorbitol increased before the second fruit-expansion period. To further understand the mechanisms regulating sucrose and sorbitol accumulation in peach fruit, expression profiles of genes involved in sugar metabolism and transport were measured. The activity and translocation protein contents of these enzymes were measured by enzyme-linked immunosorbent assay. The increased sucrose synthase activity and sucrose transporter level in the pericarp promoted the synthesis of sucrose and intake of sucrose into fruit. Sorbitol transport into fruit was promoted by the increased sorbitol transporter protein levels in leaves. In summary, greenhouse the sucrose and sorbitol contents were increased when supplemented with 1.44 kJ{middle dot}m-2{middle dot}d-1 ultraviolet B radiation before the second fruit-expansion period of peach.

plant biology↗

bioRxiv2017

Polyclonal HIV envelope-specific breast milk antibodies limit founder SHIV acquisition and cell-associated virus loads in infant rhesus monkeys.

Vertical HIV-1 transmission via breastfeeding is the predominant contributor to pediatric infections that are ongoing in this era of highly effective antiretroviral therapy (ART). Remarkably, only ~10% of infants chronically exposed to the virus via breastfeeding from untreated HIV-infected mothers become infected, suggesting the presence of naturally protective factors in breast milk. HIV-specific maternal antibodies are obvious candidates as potential contributors to this protection. This study assessed the protective capacity of common HIV envelope-specific non-broadly neutralizing antibodies isolated from breast milk of HIV-infected women in an infant rhesus monkey (RM), tier 2 SHIV oral challenge model. Prior to oral SHIV challenge, infant RMs were i.v. infused with either a single weakly-neutralizing monoclonal antibody (mAb), a tri-mAb cocktail with neutralizing and ADCC functionalities, or an anti-influenza HA control mAb. Of these groups, the fewest tri-mAb-treated infants developed plasma viremia (2/6, 3/6, and 6/8 animals viremic in tri-mAb, single-mAb, and control mAb groups, respectively). Tri-mAb-treated infants demonstrated significantly fewer transmitted/founder SHIV variants in plasma and decreased peripheral CD4+ T cell proviral loads at 8 week post-challenge compared to control mAb-treated infants. Abortive infection was observed as detectable CD4+ T cell provirus in non-viremic control mAb- and single-mAb-, but not tri-mAb-treated animals. Taken together, these results support the potential viability of maternal or infant vaccine strategies that elicit non-broadly neutralizing antibodies to prevent vertical transmission of HIV through breastfeeding.

immunology↗

bioRxiv2017

Inducing Chondrogenesis In MSC/Chondrocyte Co-Cultures Using Exogenous TGF-β: A Mathematical Model

The differentiation of mesenchymal stem cells (MSCs) into chondrocytes (native cartilage cells), or chondrogenesis, is a key step in the tissue engineering of articular cartilage, where the motility and high proliferation rate of MSCs used as seed cells are exploited. Chondrogenesis is regulated by transforming growth factor-beta (TGF-{beta}), a short-lived cytokine whose effect is prolonged by storage in the extracellular matrix. Tissue engineering applications require the complete differentiation of an initial population of MSCs, and two common strategies used to achieve this in vitro are (1) co-culture the MSCs with chondrocytes, which constitutively produce TGF-{beta}; or (2) add exogenous TGF-{beta}. To investigate these strategies we develop an ordinary differential equation model of the interactions between TGF-{beta}, MSCs and chondrocyte. Here the dynamics of TGF-{beta} are much faster than those of the cell processes; this difference in time-scales is exploited to simplify subsequent model analysis. Using our model we demonstrate that under strategy 1 complete chondrogenesis will be induced if the initial proportion of chondrocytes exceeds a critical value. Similarly, under strategy 2 we find that there is a critical concentration of exogenous TGF-{beta} above which all MSCs will ultimately differentiate. Finally, we use the model to demonstrate the potential advantages of adopting a hybrid strategy where exogenous TGF-{beta} is added to a co-culture of MSCs and chondrocytes, as compared to using either strategy 1 or 2 in isolation.

bioengineering↗

bioRxiv2017

The AWSEM-Amylometer: Predicting Amyloid Propensity And Fibril Topology Using An Optimized Folding Landscape Model

Amyloids are fibrillar protein aggregates with simple repeated structural motifs in their cores, usually {beta}-strands but sometimes -helices. Identifying the amyloid-prone regions within protein sequences is important both for understanding the mechanisms of amyloid-associated diseases and for understanding functional amyloids. Based on the crystal structures of seven cross-{beta} amyloidogenic peptides with different topologies and one recently solved cross- fiber structure, we have developed a computational approach for identifying amyloidogenic segments in protein sequences using the Associative memory, Water mediated, Structure and Energy Model. The AWSEM-Amylometer performs favorably in comparison with other predictors in predicting aggregation-prone sequences in multiple datasets. The method also predicts the specific topologies (the relative arrangement of {beta}-strands in the core) of the amyloid fibrils well. An important advantage of the AWSEM-Amylometer over other existing methods is its direct connection with an efficient, optimized protein folding simulation model, AWSEM. This connection allows one to combine efficient and accurate search of protein sequences for amyloidogenic segments with the detailed study of the thermodynamic and kinetic roles that these segments play in folding and aggregation in the context of the entire protein sequence. We present new simulation results that highlight the free energy landscapes of peptides that can take on multiple fibril topologies. We also demonstrate how the Amylometer methodology can be straightforwardly extended to the study of functional amyloids that have the recently discovered cross- fibril architecture.

biophysics↗

bioRxiv2017

Morphine selectively promotes glutamate release from glutamatergic terminals of projection neurons from medial prefrontal cortex to dopamine neurons of ventral tegmental area

Recently, we found that morphine promoted presynaptic glutamate release of dopamine (DA) neurons in the ventral tegmental area (VTA), which constituted the main mechanism for morphine-induced increase in VTA-DA neuron firing and related behaviors (Chen et al., 2015). However, what source of presynaptic glutamate release of DA neurons in the VTA is promoted by morphine remains unknown. To address this question, we used optogenetic strategy to selectively activate glutamatergic inputs from different projection neurons and then observed the effect of morphine on them. The result shows that morphine promotes glutamate release from glutamatergic terminals of projection neurons from the medial prefrontal cortex (mPFC) to VTA DA neurons, but has no effect on that from the basolateral amygdala (BLA) or the lateral hypothalamus (LH) to VTA DA neurons, and the inhibition of glutamatergic projection neurons from the mPFC to the VTA significantly reduces morphine-induced increase in locomotor activity of mice.

neuroscience↗

bioRxiv2017

Preexisting antibodies can protect against congenital cytomegalovirus infection in monkeys

Human cytomegalovirus (HCMV) is the most common congenital infection and a known cause of microcephaly, sensorineural hearing loss, and cognitive impairment among newborns worldwide. Natural maternal HCMV immunity reduces the incidence of congenital infection, but does not prevent the disease altogether. We employed a nonhuman primate model of congenital CMV infection to investigate the ability of preexisting antibodies to protect against placental CMV transmission. Pregnant, CD4+ T cell-depleted, rhesus CMV (RhCMV)-seronegative rhesus monkeys were treated with either standardly-produced hyperimmune globulin (HIG) from RhCMV-seropositive macaques or dose-optimized, potently RhCMV-neutralizing HIG prior to intravenous challenge with an RhCMV swarm. HIG passive infusion provided complete protection against fetal loss in both groups, and the potently-neutralizing HIG additionally inhibited placental transmission of RhCMV. Our findings suggest that antibody alone at the time of primary infection can prevent congenital CMV and therefore could be a primary target of vaccines to eliminate this neonatal infection.

immunology↗

bioRxiv2016

Genetic footprint of population fragmentation and contemporary collapse in a freshwater cetacean

Understanding demographic trends and patterns of gene flow in an endangered species is crucial for devising conservation strategies. Here, we examined the extent of population structure and recent evolution of the critically endangered Yangtze finless porpoise (Neophocaena asiaeorientalis asiaeorientalis). By analysing genetic variation at the mitochondrial and nuclear microsatellite loci for 148 individuals, we identified three populations along the Yangtze River, each one connected to a group of admixed ancestry. Each population displayed extremely low genetic diversity, consistent with extremely small effective size ([≤]92 individuals). Habitat degradation and distribution gaps correlated with highly asymmetric gene-flow that was inefficient in maintaining connectivity between populations. Genetic inferences of historical demography revealed that the populations in the Yangtze descended from a small number of founders colonizing the river from the sea during the last Ice Age. The colonization was followed by a rapid population split during the last millennium predating the Chinese Modern Economy Development. However, genetic diversity showed a clear footprint of population contraction over the last 50 years leaving only ~2% of the pre-collapsed size, consistent with the population collapses reported from field studies. This genetic perspective provides background information for devising mitigation strategies to prevent this species from extinction.

evolutionary biology↗