Biology subjects

Hwang, J.

Publications and source records attributed to Hwang, J..

53 records · Page 3Linked to original sources

bioRxiv2023

A Simple Bias Reduction Algorithm for RNA Sequencing Datasets

RNA sequencing (RNA-seq) is the conventional genome-scale approach used to capture the expression levels of all detectable genes in a biological sample. This is now regularly used in the clinical diagnostic space for cancer patients. While the information gained is intended to impact treatment decisions, numerous technical and quality issues remain. This includes inaccuracies in the dissemination of gene-gene relationships. For such reasons, clinical decisions are still mostly driven by DNA biomarkers, such as gene mutations or fusions. In this study, we aimed to correct for systemic bias based on RNA-sequencing platforms in order to improve our understanding of the gene-gene relationships. To do so, we examined standard pre-processed RNA-seq datasets obtained from three studies conducted by two consortium efforts including The Cancer Genome Atlas (TCGA) and Stand Up 2 Cancer (SU2C). We particularly examined the TCGA Bladder Cancer (n = 408) and Prostate Cancer (n = 498) studies as well as the SU2C Prostate Cancer study (n = 208). Using various statistical tests, we detected expression-level dependent, per-sample biases in all datasets. Using simulations, we show that these biases corrupt the results of t-tests designed to identify expression level differences between subpopulations. Importantly, these biases introduce large errors into estimates of gene-gene correlations. To mitigate these biases, we introduce Local Leveling as a novel mathematical approach that transforms count level data and corrects these observed biases. Local Leveling specifically corrects for the bias due to the inherent differential detection of transcripts that is driven by differential expression levels. Based on standard forms of count data (Raw counts, transcripts per million, fragments per kilobase of exon per million), we demonstrate that local leveling effectively removes the observed per-sample biases, and improves the accuracy in simulated statistical tests. Importantly, this led to systemic changes of gene-gene relationships when examining the correlation of key oncogenes, such as the Androgen Receptor, with all other detectable genes. Altogether, Local Leveling improves our capacity towards understanding gene-gene relationships, which may lead to novel ways to utilize the information derived from clinical tests.

genomics↗

bioRxiv2023

Micro-Scale Control of Oligodendrocyte Morphology and Myelination by the Intellectual Disability-Linked Protein Acyltransferase ZDHHC9

Mutations in the X-linked ZDHHC9 gene cause cognitive deficits in humans, with a subset of patients suffering from epilepsy. X-linked intellectual disability (XLID) is often ascribed to neuronal deficits, but here we report that expression of human and mouse ZDHHC9 orthologs is far higher in myelinating oligodendrocytes (OLs) than in other CNS cell types. ZDHHC9 codes for a protein acyltransferase (PAT), and we found that ZDHHC9 is the most highly expressed PAT in OLs. Wild type ZDHHC9 localizes to Golgi outposts in OL processes, but other PATs and XLID mutant forms of ZDHHC9 are restricted to OL cell bodies. Using genetic tools for OL progenitor fate tracing and sparse cell labeling, we show that mice lacking Zdhhc9 have grossly normal OL development but display extensive morphological and structural myelin abnormalities. Consistent with the hypothesis that these deficits are OL-autonomous, they are broadly phenocopied by acute Zdhhc9 knockdown in cultured conditions. Finally, we found that ZDHHC9 palmitoylates Myelin Basic Protein (MBP) in heterologous cells, and that palmitoylation of MBP is impaired in the Zdhhc9 knockout brain. Our findings provide critical insights into the mechanisms of ZDHHC9-associated XLID and shed new light on the palmitoylation-dependent control of myelination.

neuroscience↗

bioRxiv2023

Electrical Brain Activity during Human Walking with Parametric Variations in Terrain Unevenness and Walking Speed

Mobile brain imaging with high-density electroencephalography (EEG) can provide insight into the cortical processes involved in complex human walking tasks. While uneven terrain is common in the natural environment and poses challenges to human balance control, there is limited understanding of the supraspinal processes involved with traversing uneven terrain. The primary objective of this study was to quantify electrocortical activity related to parametric variations in terrain unevenness for neurotypical young adults. We used high-density EEG to measure brain activity when thirty-two young adults walked on a novel custom-made uneven terrain treadmill surface with four levels of difficulty at a walking speed tailored to each participant. We identified multiple brain regions associated with uneven terrain walking. Alpha (8 - 13 Hz) and beta (13 - 30 Hz) spectral power decreased in the sensorimotor and posterior parietal areas with increasing terrain unevenness while theta (4 - 8 Hz) power increased in the mid/posterior cingulate area with terrain unevenness. We also found that within stride spectral power fluctuations increased with terrain unevenness. Our secondary goal was to investigate the effect of parametric changes in walking speed (0.25 m/s, 0.5m/s, 0.75 m/s, 1.0 m/s) to differentiate the effects of walking speed from uneven terrain. Our results revealed that electrocortical activities only changed substantially with speed within the sensorimotor area but not in other brain areas. Together, these results indicate there are distinct cortical processes contributing to the control of walking over uneven terrain versus modulation of walking speed on smooth, flat terrain. Our findings increase our understanding of cortical involvement in an ecologically valid walking task and could serve as a benchmark for identifying deficits in cortical dynamics that occur in people with mobility deficits.

neuroscience↗

bioRxiv2023

Identification of ERAD-dependent degrons for the endoplasmic reticulum lumen.

Degrons are minimal protein features that are sufficient to target proteins for degradation. In most cases, degrons allow recognition by components of the cytosolic ubiquitin proteasome system. Currently, all of the identified degrons only function within the cytosol. Using Saccharomyces cerevisiae, we identified the first short linear sequences that function as degrons from the endoplasmic reticulum (ER) lumen. We show that when these degrons are transferred to proteins, they facilitate proteasomal degradation through the ERAD system. These degrons enable degradation of both luminal and integral membrane ER proteins, expanding the types of proteins that can be targeted for degradation in budding yeast and mammalian tissue culture. This discovery provides a framework to target proteins for degradation from the previously unreachable ER lumen and builds toward therapeutic approaches that exploit the highly-conserved ERAD system.

cell biology↗

bioRxiv2023

Saracatinib synergizes with enzalutamide to downregulate androgen receptor activity in castration resistant prostate cancer

Prostate cancer (PCa) remains the most diagnosed non-skin cancer amongst the American male population. Treatment for localized prostate cancer consists of androgen deprivation therapies (ADTs), which typically inhibit androgen production and the androgen receptor (AR). Though initially effective, a subset of patients will develop resistance to ADTs and the tumors will transition to castration-resistant prostate cancer (CRPC). Second generation hormonal therapies such as abiraterone acetate and enzalutamide are typically given to men with CRPC. However, these treatments are not curative and typically prolong survival only by a few months. Several resistance mechanisms contribute to this lack of efficacy such as the emergence of AR mutations, AR amplification, lineage plasticity, AR splice variants (AR-Vs) and increased kinase signaling. Having identified SRC kinase as a key tyrosine kinase enriched in CRPC patient tumors from our previous work, we evaluated whether inhibition of SRC kinase synergizes with enzalutamide or chemotherapy in several prostate cancer cell lines expressing variable AR isoforms. We observed robust synergy between the SRC kinase inhibitor, saracatinib, and enzalutamide, in the AR-FL+/AR-V+ CRPC cell lines, LNCaP95 and 22Rv1. We also observed that saracatinib significantly decreases AR Y534 phosphorylation, a key SRC kinase substrate residue, on AR-FL and AR-Vs, along with the AR regulome, supporting key mechanisms of synergy with enzalutamide. Lastly, we also found that the saracatinib-enzalutamide combination reduced DNA replication compared to the saracatinib-docetaxel combination, resulting in marked increased apoptosis. By elucidating this combination strategy, we provide pre-clinical data that suggests combining SRC kinase inhibitors with enzalutamide in select patients that express both AR-FL and AR-Vs.

cancer biology↗

bioRxiv2023

Deep mutational scanning highlights a new role for cytosolic regions in Hrd1 function

Misfolded endoplasmic reticulum proteins are degraded through a process called endoplasmic reticulum associated degradation (ERAD). Soluble, lumenal ERAD targets are recognized, retrotranslocated across the ER membrane, ubiquitinated, extracted from the membrane, and degraded by the proteasome using an ERAD pathway containing a ubiquitin ligase called Hrd1. To determine how Hrd1 mediates these processes, we developed a deep mutational scanning approach to identify residues involved in Hrd1 function, including those exclusively required for lumenal degradation. We identified several regions required for different Hrd1 functions. Most surprisingly, we found two cytosolic regions of Hrd1 required for lumenal ERAD substrate degradation. Using in vivo and in vitro approaches, we defined roles for disordered regions between structural elements that were required for Hrd1s ability to autoubiquitinate and interact with substrate. Our results demonstrate that disordered cytosolic regions promote substrate retrotranslocation by controlling Hrd1 activation and establishing directionality of retrotranslocation for lumenal substrate across the endoplasmic reticulum membrane.

biochemistry↗

bioRxiv2022

SARS-CoV-2 variants show temperature-dependent enhancedpolymerase activity in the upper respiratory tract and high transmissibility

With the convergent global emergence of SARS-CoV-2 variants of concern (VOC), a precise comparison study of viral fitness and transmission characteristics is necessary for the prediction of dominant VOCs and the development of suitable countermeasures. While airway temperature plays important roles in the fitness and transmissibility of respiratory tract viruses, it has not been well studied with SARS-CoV-2. Here we demonstrate that natural temperature differences between the upper (33{degrees}C) and lower (37{degrees}C) respiratory tract have profound effects on SARS-CoV-2 replication and transmission. Specifically, SARS-COV-2 variants containing the P323L or P323L/G671S mutation in the NSP12 RNA-dependent RNA polymerase (RdRp) exhibited enhanced RdRp enzymatic activity at 33{degrees}C compared to 37{degrees}C and high transmissibility in ferrets. MicroScale Thermophoresis demonstrated that the NSP12 P323L or P323L/G671S mutation stabilized the NSP12-NSP7-NSP8 complex interaction. Furthermore, reverse genetics-derived SARS-CoV-2 variants containing the NSP12 P323L or P323L/G671S mutation displayed enhanced replication at 33{degrees}C, and high transmission in ferrets. This suggests that the evolutionarily forced NSP12 P323L and P323L/G671S mutations of recent SARS-CoV-2 VOC strains are associated with increases of the RdRp complex stability and enzymatic activity, promoting the high transmissibility.

microbiology↗

bioRxiv2022

ST6GAL1 sialyltransferase promotes acinar to ductal metaplasia and pancreatic cancer progression

The role of aberrant glycosylation in pancreatic ductal adenocarcinoma (PDAC) remains an under-investigated area of research. In this study, we determined that the ST6GAL1 sialyltransferase, which adds 2,6-linked sialic acids to N-glycosylated proteins, is upregulated in patients with early-stage PDAC, and further increased in advanced disease. A tumor-promoting function for ST6GAL1 was elucidated using tumor xenograft models with human PDAC cells. Additionally, we developed a genetically-engineered mouse (GEM) with transgenic expression of ST6GAL1 in the pancreas, and found that mice with dual expression of ST6GAL1 and oncogenic KRASG12D have greatly accelerated PDAC progression and mortality compared with mice expressing KRASG12D alone. As ST6GAL1 imparts progenitor-like characteristics, we interrogated ST6GAL1s role in acinar to ductal metaplasia (ADM), a process that fosters neoplasia by reprogramming acinar cells into ductal, progenitor-like cells. We confirmed that ST6GAL1 promotes ADM using multiple models including the 266-6 cell line, GEM-derived organoids and tissues, and an in vivo model of inflammation-induced ADM. EGFR is a key driver of ADM and is known to be activated by ST6GAL1-mediated sialylation. Importantly, EGFR activation was dramatically increased in acinar cells and organoids from mice with transgenic ST6GAL1 expression. These collective results highlight a novel glycosylation-dependent mechanism involved in early stages of pancreatic neoplasia.

cancer biology↗

bioRxiv2022

Uneven terrain treadmill walking in younger and older adults

We developed a method for altering terrain unevenness on a treadmill to study gait kinematics. Terrain consisted of rigid polyurethane disks (12.7 cm diameter, 1.3-3.8 cm tall) which attached to the treadmill belt using hook-and-loop fasteners. Here, we tested four terrain unevenness conditions: Flat, Low, Medium, and High. The main objective was to test the hypothesis that increasing the unevenness of the terrain would result in greater gait kinematic variability. Seventeen younger adults (age 20-40 years), 25 higher-functioning older adults (age 65+ years), and 29 lower-functioning older adults (age 65+ years, Short Physical Performance Battery score < 10) participated. We customized the treadmill speed to each participants walking ability, keeping the speed constant across all four terrain conditions. Participants completed two 3-minute walking trials per condition. Using an inertial measurement unit placed over the sacrum and pressure sensors in the shoes, we calculated the stride-to-stride variability in step duration and sacral excursion (coefficient of variation; standard deviation expressed as percentage of the mean). Participants also self-reported their perceived stability for each condition. Terrain was a significant predictor of step duration variability, which roughly doubled from Flat to High terrain for all participant groups: younger adults (Flat 4.0%, High 8.2%), higher-functioning older adults (Flat 5.0%, High 8.9%), lower-functioning older adults (Flat 7.0%, High 14.1%). Similarly, all groups exhibited significant increases in sacral excursion variability for all three uneven terrain conditions compared to Flat. Participants were also significantly more likely to report feeling less stable walking over all three uneven terrain conditions compared to Flat. These findings support the hypothesis that altering terrain unevenness on a treadmill will increase gait kinematic variability and reduce perceived stability in younger and older adults.

bioengineering↗

bioRxiv2022

Single Cell Spatial Analysis Reveals the Topology of Immunomodulatory Purinergic Signaling in Glioblastoma

Glioblastoma develops an immunosuppressive microenvironment that fosters tumorigenesis and resistance to current therapeutic strategies. Here we use multiplexed tissue imaging and single-cell RNA-sequencing to characterize the composition, spatial organization, and clinical significance of extracellular purinergic signaling in glioblastoma. We show that glioblastoma exhibit strong expression of CD39 and CD73 ectoenzymes, correlating with increased adenosine levels. Microglia are the predominant source of CD39, while CD73 is principally expressed by tumor cells, particularly in tumors with amplification of EGFR and astrocyte-like differentiation. Spatially-resolved single-cell analyses demonstrate strong spatial correlation between tumor CD73 and microglial CD39, and that their spatial proximity is associated with poor clinical outcomes. Together, this data reveals that tumor CD73 expression correlates with tumor genotype, lineage differentiation, and functional states, and that core purine regulatory enzymes expressed by neoplastic and tumor-associated myeloid cells interact to promote a distinctive adenosine-rich signaling niche and immunosuppressive microenvironment potentially amenable to therapeutic targeting.

cancer biology↗

bioRxiv2021

The Lyme Disease agent co-opts adiponectin receptor-mediated signaling in its arthropod vector

Adiponectin-mediated pathways contribute to mammalian homeostasis; however, little is known about adiponectin and adiponectin receptor signaling in arthropods. In this study, we demonstrate that Ixodes scapularis ticks have an adiponectin receptor-like protein (ISARL) but lack adiponectin - suggesting activation by alternative pathways. ISARL expression is significantly upregulated in the tick gut after Borrelia burgdorferi infection suggesting that ISARL-signaling may be co-opted by the Lyme disease agent. Consistent with this, RNA interference (RNAi)-mediated silencing of ISARL significantly reduced the B. burgdorferi burden in the tick. RNA-seq-based transcriptomics and RNAi assays demonstrate that ISARL-mediated phospholipid metabolism by phosphatidylserine synthase I is associated with B. burgdorferi survival. Furthermore, the tick complement C1q-like protein 3 interacts with ISARL, and B. burgdorferi facilitates this process. This study identifies a new tick metabolic pathway that is connected to the life cycle of the Lyme disease spirochete. Significance StatementAdiponectin binds to adiponectin receptors and participates in glucose and lipid metabolism in mammals. In this study, we found that ticks have an adiponectin receptor-like protein but lack adiponectin. Importantly, we demonstrated that the Lyme disease agent, Borrelia burgdorferi, takes advantage of the adiponectin receptor signaling pathway to establish infection in its arthropod vector, Ixodes scapularis. Our study sheds light on the understanding of Borrelia-tick interactions and provides insights into how a human infectious disease agent may evolve to manipulate host metabolism for its own benefits. Understanding this pathway may lead to new ways to interfere with the Borrelia life cycle, and this mechanism may be applicable to additional microbes that are transmitted by ticks, mosquitoes or other arthropods.

microbiology↗

bioRxiv2020

Biologically informed deep neural network for prostate cancer classification and discovery

Determination of molecular features that mediate clinically aggressive phenotypes in prostate cancer (PrCa) remains a major biological and clinical challenge. Here, we developed a biologically informed deep learning model (P-NET) to stratify PrCa patients by treatment resistance state and evaluate molecular drivers of treatment resistance for therapeutic targeting through complete model interpretability. Using a molecular cohort of 1,238 prostate cancers, we demonstrated that P-NET can predict cancer state using molecular data that is superior to other modeling approaches. Moreover, the biological interpretability within P-NET revealed established and novel molecularly altered candidates, such as MDM4 and FGFR1, that were implicated in predicting advanced disease and validated in vitro. Broadly, biologically informed fully interpretable neural networks enable preclinical discovery and clinical prediction in prostate cancer and may have general applicability across cancer types.

bioinformatics↗

bioRxiv2020

Reprogramming of the FOXA1 cistrome in treatment-emergent neuroendocrine prostate cancer

Lineage plasticity, the ability of a cell to alter its identity, is an increasingly common mechanism of adaptive resistance to targeted therapy in cancer1,2. An archetypal example is the development of neuroendocrine prostate cancer (NEPC) after treatment of prostate adenocarcinoma (PRAD) with inhibitors of androgen signaling. NEPC is an aggressive variant of prostate cancer that aberrantly expresses genes characteristic of neuroendocrine (NE) tissues and no longer depends on androgens. To investigate the epigenomic basis of this resistance mechanism, we profiled histone modifications in NEPC and PRAD patient-derived xenografts (PDXs) using chromatin immunoprecipitation and sequencing (ChIP-seq). We identified a vast network of cis-regulatory elements (N~15,000) that are recurrently activated in NEPC. The FOXA1 transcription factor (TF), which pioneers androgen receptor (AR) chromatin binding in the prostate epithelium3,4, is reprogrammed to NE-specific regulatory elements in NEPC. Despite loss of dependence upon AR, NEPC maintains FOXA1 expression and requires FOXA1 for proliferation and expression of NE lineage-defining genes. Ectopic expression of the NE lineage TFs ASCL1 and NKX2-1 in PRAD cells reprograms FOXA1 to bind to NE regulatory elements and induces enhancer activity as evidenced by histone modifications at these sites. Our data establish the importance of FOXA1 in NEPC and provide a principled approach to identifying novel cancer dependencies through epigenomic profiling.

cancer biology↗

bioRxiv2020

Macrophage scavenger receptor 1 controls Chikungunya virus infection through autophagy

Macrophage scavenger receptor 1 (MSR1) mediates the endocytosis of modified low-density lipoproteins and plays an important antiviral role. However, the molecular mechanism underlying MSR1 antiviral actions remains elusive. Herein, we report that MSR1 activates autophagy to restrict infection of Chikungunya virus (CHIKV), an arthritogenic alphavirus that causes acute and chronic crippling arthralgia. Msr1 expression was rapidly upregulated after CHIKV infection in mice. Msr1 knockout mice had elevated viral loads and increased susceptibility to CHIKV arthritis along with a normal type I IFN response. Induction of LC3 lipidation by CHIKV, a marker of autophagy, was reduced in Msr1-/- cells. Mechanistically, MSR1 interacted with ATG12 through its cytoplasmic tail and this interaction was enhanced by CHIKV nsP1 protein. MSR1 repressed CHIKV replication through ATG5-ATG12-ATG16L1 and this was dependent on the FIP200-and-WIPI2-binding domain, but not the WD40 domain of ATG16L1. Our results elucidate an antiviral role for MSR1 involving the autophagic function of ATG5-ATG12-ATG16L1.

microbiology↗

bioRxiv2018

Real-time fine aerosol exposures in taconite mining operations

Respiratory health effects such as mesothelioma, silicosis, and lung cancer have been shown to be associated with working in the taconite mining industry. Taconite workers may also have elevated risks from cardiovascular disease (CVD), although the relationship of CVD to dust exposures at these mines has not been well-studied. Motivated by evidence from environmental epidemiological studies and occupational cohorts that have implicated the effects of fine particulates with increased risk of cardiovascular diseases, we conducted an air monitoring campaign to characterize fine aerosol concentrations at 91 locations across six taconite mines using an array of direct-reading instruments to obtain measurements of mass concentrations (PM2.5 or particles with aerodynamic diameter less than 2.5 m, and respirable particulate matter or RPM), alveolar-deposited surface area concentrations (ADSA), particle number concentrations (PN), and particle size distributions. To analyze these data, we fit a Bayesian hierarchical model with an AR(1) correlation structure to estimate exposure while accounting for temporal correlation. The highest estimated geometric means (GMs) were observed in the pelletizing and concentrating departments (pelletizing maintenance, balling drum operator, and concentrator operator) for PM2.5 and RPM. ADSA and PN generally had highest GMs in the pelletizing department, which processed large amounts of powder-like particles into iron pellets. The within-location variability (GSD_WL) generally ranged from 1 to 3 for all exposure metrics, except for a few locations which indicated changes of activities that caused the exposures to change. Between-location variability (GSD_BL) estimates were generally higher than GSD_WL, indicating larger differences in exposure levels at different locations between mines than at individual locations over the course of several hours. Ranking between PM2.5 and RPM generally agree with each other, whereas ADSA and PN were more consistent with each other, with some overlap with PM2.5 and RPM. Differences in ranking these groups may have potential implication for occupational epidemiological studies that rely on exposure information to detect an exposure-response relationship for various job groups. Future epidemiological studies investigating fine aerosol exposures and health risks in occupational settings are encouraged to use multiple metrics to see how they influence health outcomes risk.

epidemiology↗

bioRxiv2017

DEET as a feeding deterrent

The insect repellent N,N-diethyl-3-methylbenzamide (DEET), is a multimodal compound that acts as a spatial repellent as well as an irritant (contact repellent), thus being perceived by the insects olfactory and gustatory systems as an odorant and a tastant, respectively. Soon after DEET was developed, almost 6 decades ago, it was reported that it reduced mosquito feeding on blood mixed with this repellent. It is now known that the mosquito proboscis senses contact repellents with the tips (labella) of the labium, which remain in direct contact with the outer layers of the skin, while the stylets, including the feeding deterrent sensor (labrum), penetrate the skin. We designed a behavioral assay that allowed us to tease apart contact repellency from feeding deterrence. First, we demonstrate here that when DEET was mixed with blood and covered by Parafilm(R) layers, it did not leak to the outer surface. In our assays, the mean number of landings and duration of contacts with surfaces covering blood mixed with DEET or blood plus solvent (dimethyl sulfoxide) did not differ significantly. The feeding times, however, were significantly different. When blood was mixed either with 0.1 or 1% DEET, female southern house mosquitoes spent significantly less time feeding than the time spent feeding on blood mixed only with the solvent. By contrast, there were no significant differences in the mean times of feeding on blood containing 1% picaridin and blood plus solvent. Like DEET, the contact repellent and insecticide, permethrin, caused a significant reduction in feeding time. We, therefore, concluded, that in this context, DEET and permethrin act as feeding deterrents.

animal behavior and cognition↗

bioRxiv2017

HiGlass: Web-based Visual Comparison And Exploration Of Genome Interaction Maps

We present HiGlass, an open source visualization tool built on web technologies that provides a rich interface for rapid, multiplex, and multiscale navigation of 2D genomic maps alongside 1D genomic tracks, allowing users to combine various data types, synchronize multiple visualization modalities, and share fully customizable views with others. We demonstrate its utility in exploring different experimental conditions, comparing the results of analyses, and creating interactive snapshots to share with collaborators and the broader public. HiGlass is accessible online at http://higlass.io and is also available as a containerized application that can be run on any platform.

bioinformatics↗