Biology subjects

Park, J.

Publications and source records attributed to Park, J..

At least 37 records · Page 2Linked to original sources

bioRxiv2018

System level analyses of motor-related neural activities in larval Drosophila

The way in which the central nervous system (CNS) governs animal movement is complex and difficult to solve solely by the analyses of muscle movement patterns. We tackle this problem by observing the activity of a large population of neurons in the CNS of larval Drosophila. We focused on two major behaviors of the larvae, forward and backward locomotion, and analyzed the neuronal activity related to these behaviors during fictive locomotion that spontaneously occurs in the isolated CNS. We expressed genetically-encoded calcium indicator, GCaMP, and a nuclear marker in all neurons and used digital scanned light-sheet microscopy to record neural activities in the entire ventral nerve cord at a fast frame rate. We developed image processing tools that automatically detect the cell position based on the nuclear staining and allocate the activity signals to each detected cell. We also applied a machine learning-based method that we developed recently to assign motor status in each time frame. Based on these methods, we find cells whose activity is biased to forward versus backward locomotion and vice versa. In particular, we identified a group of neurons near the boundary of subesophageal zone (SEZ) and thoracic neuromeres, which are strongly active during an early phase of backward but not forward fictive locomotion. Our experimental procedure and computational pipeline enable systematic identification of neurons to show characteristic motor activities in larval Drosophila.

neuroscience↗

bioRxiv2018

Jagged1/Notch2 Controls Kidney Fibrosis via Tfam-mediated Metabolic Reprogramming

While Notch signaling has been proposed to play a key role in fibrosis, the direct molecular pathways targeted by Notch signaling and the precise ligand and receptor pair that are responsible for kidney disease remain poorly defined.\n\nIn this study, we found that JAG1 and NOTCH2 showed the strongest correlation with the degree of interstitial fibrosis in a genome wide expression analysis of a large cohort of human kidney samples. RNA sequencing analysis of kidneys of mice with folic acid nephropathy, unilateral ureteral obstruction, or APOL1-associated kidney disease indicated that Jag1 and Notch2 levels were higher in all analyzed kidney fibrosis models. Mice with tubule-specific deletion of Jag1 or Notch2 (Kspcre/Jag1flox/flox, and Kspcre/Notch2flox/flox) had no kidney-specific alterations at baseline, but showed protection from folic acid induced kidney fibrosis. Tubule-specific genetic deletion of Notch1 and global knock-out of Notch3 had no effect on fibrosis. In vitro chromatin immunoprecipitation experiments and genome-wide expression studies identified the mitochondrial transcription factor A (Tfam) as a direct Notch target. Re-expression of Tfam in tubule cells prevented Notch-induced metabolic and profibrotic reprogramming. Kidney tubule specific deletion of Tfam resulted in perinatal lethality.\n\nIn summary, Jag1/Notch2 plays a key role in kidney fibrosis development by regulating Tfam expression and metabolic reprogramming.

pathology↗

bioRxiv2018

Internal gain modulations, but not changes in stimulus contrast, preserve the neural code

Neurons in primary visual cortex (V1) are strongly modulated both by stimulus contrast and by fluctuations of internal inputs. An important question is whether population codes are preserved under these conditions. Changes in stimulus contrast are thought to leave population codes invariant, whereas the effect of internal gain modulations remains unknown. To address these questions we studied how the direction-of-motion of oriented gratings is encoded in layer 2/3 of mouse V1. Surprisingly, we found that, because contrast gain responses across cells are heterogeneous, a change in contrast alters the information distribution profile across cells leading to the failure of contrast invariance. Remarkably, internal input fluctuations that cause commensurate firing rate modulations at the single-cell level, do respect population code invariance. These observations have important implications for visual information encoding, and argue that the brain strives to maintain the stability of the neural code in the face of fluctuating internal inputs.

neuroscience↗

bioRxiv2018

Data-driven analysis of motor activity implicated 5-HT2A neurons in backward locomotion of larval Drosophila

Rhythmic animal behaviors are regulated in part by neural circuits called the central pattern generators (CPGs). Classifying neural population activities correlated with body movements and identifying the associated component neurons are critical steps in understanding CPGs. Previous methods that classify neural dynamics obtained by dimension reduction algorithms often require manual optimization which could be laborious and preparation-specific. Here, we present a simpler and more flexible method that is based on the pre-trained convolutional neural network model VGG-16 and unsupervised learning, and successfully classifies the fictive motor patterns in Drosophila larvae under various imaging conditions. We also used voxel-wise correlation mapping to identify neurons associated with motor patterns. By applying these methods to neurons targeted by 5-HT2A-GAL4, which we generated by the CRISPR/Cas9-system, we identified two classes of interneurons, termed Seta and Leta, which are specifically active during backward but not forward fictive locomotion. Optogenetic activation of Seta and Leta neurons increased backward locomotion. Conversely, thermogenetic inhibition of 5-HT2A-GAL4 neurons or application of a 5-HT2 antagonist decreased backward locomotion induced by noxious light stimuli. This study establishes an accelerated pipeline for activity profiling and cell identification in larval Drosophila and implicates the serotonergic system in the modulation of backward locomotion.

neuroscience↗

bioRxiv2018

Whole Genome Sequencing and Rare Variant Analysis in Essential Tremor Families

Essential tremor (ET) is one of the most common movement disorders. The etiology of ET remains largely unexplained. Whole genome sequencing (WGS) is likely to be of value in understanding a large proportion of ET with Mendelian and complex disease inheritance patterns. In ET families with Mendelian inheritance patterns, WGS may lead to gene identification where WES analysis failed to identify the causative variant due to incomplete coverage of the entire coding region of the genome. Alternatively, in ET families with complex disease inheritance patterns with gene x gene and gene x environment interactions enrichment of functional rare coding and non-coding variants may explain the heritability of ET. We performed WGS in eight ET families (n=40 individuals) enrolled in the Family Study of Essential Tremor. The analysis included filtering WGS data based on allele frequency in population databases, rare variant classification and association testing using the Mixed-Model Kernel Based Adaptive Cluster (MM-KBAC) test and prioritization of candidate genes identified within families using phenolyzer. WGS analysis identified candidate genes for ET in 5/8 (62.5%) of the families analyzed. WES analysis in a subset of these families in our previously published study failed to identify candidate genes. In one family, we identified a deleterious and damaging variant (c.1367G>A, p.(Arg456Gln)) in the candidate gene, CACNA1G, which encodes the pore forming subunit of T-type Ca(2+) channels, CaV3.1, and is expressed in various motor pathways and has been previously implicated in neuronal autorhythmicity and ET. Other candidate genes identified include SLIT3 (family D), which encodes an axon guidance molecule and in three families, phenolyzer prioritized genes that are associated with hereditary neuropathies (family A, KARS, family B, KIF5A and family F, NTRK1). This work has identified candidate genes and pathways for ET that can now be prioritized for functional studies.

genetics↗

bioRxiv2017

Modeling acute myeloid leukemia in a continuum of differentiation states

Here we present a mathematical model of movement in an abstract space representing states of cellular differentiation. We motivate this work with recent examples that demonstrate a continuum of cellular differentiation using single cell RNA sequencing data to characterize cellular states in a high-dimensional space, which is then mapped into [R]2 or [R]3 with dimension reduction techniques. We represent trajectories in the differentiation space as a graph, and model directed and random movement on the graph with partial differential equations. We hypothesize that flow in this space can be used to model normal differentiation processes as well as predict the evolution of abnormal differentiation processes such as those observed during pathogenesis of acute myeloid leukemia (AML).

cancer biology↗

bioRxiv2017

Metabolic Interactions Between Dynamic Bacterial Subpopulations

Within multi-species microbial communities, individual species are known to occupy distinct metabolic niches. By contrast, it has remained largely unclear whether and how metabolic specialization occurs within clonal bacterial populations. The possibility of such metabolic specialization in clonal populations raises several questions: Does specialization occur, and if it does, which metabolic processes are involved? How is specialization coordinated? How rapidly do cells switch between states? And finally, what functions might metabolic specialization provide? One potential function of metabolic specialization could be to manage overflow metabolites such as acetate, which presents a toxic challenge due to low pH, and protective pH-neutral overflow metabolites. Here we show that exponentially dividing Bacillus subtilis cultures divide into distinct interacting metabolic subpopulations including one population that produces acetate, and another population that differentially expresses metabolic genes for the production of acetoin, a pH-neutral storage molecule. These subpopulations grew at distinct rates, and cells switched dynamically between states, with acetate influencing the relative sizes of the different subpopulations. These results show that clonal populations can use metabolic specialization to control the environment through a process of dynamic, environmentally-sensitive state-switching.

microbiology↗

bioRxiv2017

Comprehensive single cell RNAseq analysis of the kidney reveals novel cell types and unexpected cell plasticity

A key limitations to understand kidney function and disease development has been that specific cell types responsible for specific homeostatic kidney function or disease phenotypes have not been defined at the molecular level.\n\nTo fill this gap, we characterized 57,979 cells from healthy mouse kidneys using unbiased single-cell RNA sequencing. We show that genetic mutations that present with similar phenotypes mostly affect genes that are expressed in a single unique differentiated cell type. On the other hand, we found unexpected cell plasticity of epithelial cells in the final segment of the kidney (collecting duct) that is responsible for final composition of the urine. Using computational cell trajectory analysis and in vivo linage tracing, we found that, intercalated cells (that secrete protons) and principal cells (that maintain salt, water and potassium balance) undergo a Notch mediated interconversion via a newly identified transitional cell type. In disease states this transition is shifted towards the principal cell fate. Loss of intercalated cells likely contributes to metabolic acidosis observed in kidney disease.\n\nIn summary, single cell analysis advanced a mechanistic description of kidney diseases by identifying a defective homeostatic cell lineage.\n\nOne Sentence SummaryA comprehensive single cell atlas of the kidney reveals a transitional cell type and cell plasticity determined by Notch signaling which is defective in chronic kidney disease.

cell biology↗

bioRxiv2017

Deciphering programs of transcriptional regulation by combined deconvolution of multiple omics layers

Metazoans are crucially dependent on multiple layers of gene regulatory mechanisms which allow them to control gene expression across developmental stages, tissues and cell types. Multiple recent research consortia have aimed to generate comprehensive datasets to profile the activity of these cell type- and condition-specific regulatory landscapes across many different cell lines and primary cells. However, extraction of genes or regulatory elements specific to certain entities from these datasets remains challenging. We here propose a novel method based on non-negative matrix factorization for disentangling and associating huge multi-assay datasets including chromatin accessibility and gene expression data. Taking advantage of implementations of NMF algorithms in the GPU CUDA environment full datasets composed of tens of thousands of genes as well as hundreds of samples can be processed without the need for prior feature selection to reduce the input size. Applying this framework to multiple layers of genomic data derived from human blood cells we unravel mechanisms of regulation of cell type-specific expression in T-cells and monocytes.

genomics↗

bioRxiv2017

Molecular function limits divergent protein evolution on planetary timescales

Functional conservation is known to constrain protein evolution. Nevertheless, the long-term divergence patterns of proteins maintaining the same molecular function and the possible limits of this divergence have not been explored in detail. We investigate these fundamental questions by characterizing the divergence between ancient protein orthologs with conserved molecular function. Our results demonstrate that the decline of sequence and structural similarities between such orthologs significantly slows down after ~1-2 billion years of independent evolution. As a result, their sequence and structural similarities have not substantially decreased for the past billion years. The effective divergence limit (>25% sequence identity) is not primarily due to protein sites universally conserved in all linages. Instead, less than four amino acid types are accepted, on average, per site in orthologs strictly conserving their molecular function. Our analysis also reveals different divergence patterns for protein sites with experimentally determined small and large fitness effects of mutations.

evolutionary biology↗

bioRxiv2017

Increased axonal bouton stability during learning in the mouse model of MECP2 duplication syndrome

MECP2-duplication syndrome is an X-linked form of syndromic autism caused by genomic duplication of the region encoding Methyl-CpG-binding protein 2. Mice overexpressing MECP2 demonstrate altered patterns of learning and memory, including enhanced motor learning. Previous work associated this enhanced motor learning to abnormally increased stability of dendritic spine clusters formed in the apical tuft of corticospinal, area M1, neurons during rotarod training. In the current study, we measure the structural plasticity of axonal boutons in Layer 5 (L5) pyramidal neuron projections to layer 1 of area M1 during motor learning. In wild-type mice we find that during rotarod training, bouton formation rate changes minimally, if at all, while bouton elimination rate doubles. Notably, the observed upregulation in bouton elimination with learning is absent in MECP2-duplication mice. This result provides further evidence of imbalance between structural stability and plasticity in this form of syndromic autism. Furthermore, the observation that axonal bouton elimination doubles with motor learning in wild-type animals contrasts with the increase of dendritic spine consolidation observed in corticospinal neurons at the same layer. This dissociation suggests that different area M1 microcircuits may manifest different patterns of structural synaptic plasticity during motor learning.\n\nSIGNIFICANCE STATEMENTAbnormal balance between synaptic stability and plasticity is a feature of several autism spectrum disorders, often corroborated by in vivo studies of dendritic spine turnover. Here we provide the first evidence that abnormally increased stability of axonal boutons, the presynaptic component of excitatory synapses, occurs during motor learning in the MECP2 duplication syndrome mouse model of autism. In contrast, in normal controls, axonal bouton elimination in L5 pyramidal neuron projections to layer 1 of area M1 doubles with motor learning. The fact that axonal projection boutons get eliminated, while corticospinal dendritic spines get consolidated with motor learning in layer 1 of area M1, suggests that structural plasticity manifestations differ across different M1 microcircuits.

neuroscience↗

bioRxiv2017

Structural and functional diversity of a dense sample of retinal ganglion cells

To aid understanding of retinal structure and function, we present as an online resource the dendritic arbors and visual responses of ganglion cells in a single patch of mouse retina. We divide the inner plexiform layer, which contains the dendritic arbors of ganglion cells, into four sublaminae defined by a purely anatomical principle of arbor segregation. The sublaminae serve as the starting point for a hierarchical clustering of our ganglion cells. We propose and apply a quantitative criterion for validating a cluster as a ganglion cell type: the aggregate neurite density of a type should be approximately uniform (\"density conservation\"). Finally, we find that ganglion cells arborizing in the inner marginal sublamina of the inner plexi-form layer exhibit significantly more sustained visual responses on average.

neuroscience↗

bioRxiv2017

Pathway centrality in protein interaction networks identifies functional mediators of pulmonary disease

Identification of functional pathways mediating molecular responses may lead to better understanding of disease processes and suggest new therapeutic approaches. We introduce a method to detect such mediating functions using topological properties of protein-protein interaction networks. We introduce the concept of pathway centrality, a measure of communication between disease genes and differentially expressed genes. We find mediating pathways for three pulmonary diseases (asthma; bronchopulmonary dysplasia (BPD); and chronic obstructive pulmonary disease (COPD)) using pathway centrality. Mediating pathways shared by all three pulmonary disorders heavily favor inflammatory or immune responses and include specific pathways such as cytokine production, NF Kappa B, and JAK/STAT signaling. Disease-specific mediators, such as insulin signaling in BPD or homeostasis in COPD, are also highlighted. We support our findings, some of which suggest new treatment approaches, both with anecdotal evidence from the literature and via systematic evaluation using genetic interactions.

bioinformatics↗

bioRxiv2017

Questioning unexpected CRISPR off-target mutations in vivo

To the Editor To the Editor References Recently, Schaefer et al.1 reported that whole genome sequencing (WGS) of two Cas9-treated, gene-corrected mice and a wild-type control mouse unveiled 1,397 single-nucleotide variations (SNVs) and 117 small insertions and deletions (indels) present commonly in the two Cas9-treated mice \"but absent in the uncorrected control\" and from a database of mouse SNVs and indels. There was essentially no sequence homology between the on-target site and these SNVs and indel sites, most of which lacked a protospacer-adjacent motif (PAM) sequence, suggesting that these variations were both small guide RNA (sgRNA)-independent and Cas9-independent, respectively. Nevertheless, the authors made a bold claim that these variations were caused by CRISPR-Cas9 without validating these unexpected off-target effects even at a single SNV or indel site by ...

genomics↗

bioRxiv2017

Discrete and coordinated encoding of punishment contingent on rewarded actions by prefrontal cortex and VTA

Actions motivated by a rewarding outcome are often associated with a risk of punishment. Little is known about the neural representation of punishment that is contingent on reward-guided behavior. We modeled this circumstance by using a task where actions were consistently rewarded but probabilistically punished. Spike activity and local field potentials were recorded during this task simultaneously from VTA and mPFC, two reciprocally connected regions implicated in both reward-seeking and aversive behavioral states. At the single unit level, we found that ensembles of VTA and mPFC neurons encode the contingency between action and punishment. At the network level, we found that coherent theta oscillations synchronize the VTA and mPFC in a bottom-up direction, effectively phase-modulating the neuronal spike activity in the two regions during punishment-free actions. This synchrony declined as a function of punishment contingency, suggesting that during reward-seeking actions, risk of punishment diminishes VTA-driven neural synchrony between the two regions.

neuroscience↗

bioRxiv2017

TDG Regulates Cell Cycle Progression In Human Neural Progenitors

As cells divide, they must replicate both their DNA and generate a new set of histone proteins. The newly synthesized daughter strands and histones are unmodified and must therefore be covalently modified to allow for transmission of important epigenetic marks to daughter cells. Human pluripotent stem cells (hPSCs) display a unique cell cycle profile, and control of the cell cycle is known to be critical for their proper differentiation and survival. A major unresolved question is how hPSCs regulate their DNA methylation status through the cell cycle, namely how passive and active demethylation work to maintain a stable genome. TDG, an embryonic essential gene, has been recently implicated as a major enzyme involved in demethylation1. Here we present new data showing that TDG regulates cell cycle related gene expression in human neural progenitors (NPCs) derived from hPSCs and controls their capacity for neural differentiation. These observations suggest that TDG and active demethylation play an important role in hPSC cell cycle regulation and differentiation.

molecular biology↗

bioRxiv2017

Resolving Drug Effects In Patient-Derived Cancer Cells Links Organoid Responses To Genome Alterations

Cancer drug screening in patient-derived cells holds great promise for personalized oncology and drug discovery but lacks standardization. Whether cells are cultured as conventional monolayer or advanced organoid cultures influences drug effects and thereby drug selection and clinical success. To precisely compare drug profiles in differently cultured primary cells, we developed DeathPro, an automated microscopy-based assay to resolve drug-induced cell death and proliferation inhibition. Using DeathPro, we screened cells from ovarian cancer patients in monolayer or organoid culture with clinically relevant drugs. Drug-induced growth arrest and efficacy of cytostatic drugs differed between the two culture systems. Interestingly, drug effects in organoids were more diverse and had lower therapeutic potential. Genomic analysis revealed novel links between drug sensitivity and DNA repair deficiency in organoids that were undetectable in monolayers. Thus, our results highlight the dependency of cytostatic drugs and pharmacogenomic associations on culture systems, and guide culture selection for drug tests.

cancer biology↗

bioRxiv2017

Multiple reference genome sequences of hot pepper reveal the massive evolution of plant disease resistance genes by retroduplication

Transposable elements (TEs) provide major evolutionary forces leading to new genome structure and species diversification. However, the role of TEs in the expansion of disease resistance gene families has been unexplored in plants. Here, we report high-quality de novo genomes for two peppers (Capsicum baccatum and C. chinense) and an improved reference genome (C. annuum). Dynamic genome rearrangements involving translocations among chromosome 3, 5 and 9 were detected in comparison between C. baccatum and the two other peppers. The amplification of athila LTR-retrotransposons, members of the gypsy superfamily, led to genome expansion in C. baccatum. In-depth genome-wide comparison of genes and repeats unveiled that the copy numbers of NLRs were greatly increased by LTR-retrotransposon-mediated retroduplication. Moreover, retroduplicated NLRs exhibited great abundance across the angiosperms, with most cases lineage-specific and thus recent events. Our study revealed that retroduplication has played key roles in the emergence of new disease-resistance genes in plants.

plant biology↗