Biology subjects

Phillips, J.

Publications and source records attributed to Phillips, J..

At least 19 recordsLinked to original sources

bioRxiv2026

Development and validation of highly selective monoclonal antibodies for the detection of huntingtin neoepitopes

Huntington's disease is caused by a CAG repeat tract expansion in the huntingtin gene, resulting in production of pathogenic N-terminal huntingtin protein fragments associated with disease pathology. Despite their central role, detection of these fragments has relied on a limited antibody repertoire with reproducibility concerns. Here, we describe the generation and characterization of recombinant rabbit monoclonal antibodies targeting two reciprocal neoepitopes flanking the huntingtin exon 1/exon 2 junction corresponding to amino acids P90 and K91. The P90 antibodies (clones 1B12, 11G2) demonstrate fragment-length-selective recognition of the C-terminal HTTexon1 P90 neoepitope with no detectable binding to full length huntingtin. A side-by-side comparison of the widely used monoclonal antibody MW8 from two different sources revealed measurable lot-to-lot drift in its fragment selectivity, whereas the recombinant P90 antibodies, expressed from a defined, sequenced clone, maintained consistent specificity, addressing this source-dependent variability. Whereas P90-positive fragments can arise through alternative splicing of the HTT1a transcript, generation of the reciprocal K91 N-terminal HTTexon2 neoepitope would require site-specific proteolytic cleavage, a mechanism that has not yet been directly tested for lack of a suitable reagent. The K91 antibody (clone 7G10) binds the N-terminal K91 neoepitope with high affinity and specificity over full length huntingtin and provides, for the first time, a tool capable of directly interrogating whether such cleavage occurs. Neoepitope specificity of these antibodies was orthogonally confirmed by protease digestion (Lys-N and Arg-C) coupled with intact mass spectrometry. As an additional outcome of the immunization and selection strategy, we discovered human-mouse cross-reactive antibodies (clones 27F5, 31C10) targeting the proline-rich domain of huntingtin that will facilitate mouse-human translational studies. All antibodies are recombinant, ensuring long-term reproducibility, and are being made available, along with their sequences, to the research community.

neuroscience↗

bioRxiv2026

Enhancing the detection of HTT1a with neoepitope antibodies in mouse models of Huntington's disease

Huntingtons disease is an inherited neurodegenerative disorder caused by a CAG repeat expansion in exon 1 of the Huntingtin (HTT) gene, encoding an expanded polyglutamine tract in the huntingtin (HTT) protein. The pathogenic CAG repeat of HTT is unstable and undergoes progressive somatic expansion in specific brain cells and peripheral tissues throughout life. Genes involved in DNA mismatch repair pathways, which promote repeat expansion, have been identified as genetic modifiers of the disease. Consequently, the rate of CAG repeat expansion is a key determinant driving the age of onset and disease progression. As the CAG repeat expands, alternative processing of HTT pre-mRNA increasingly favours production of the HTT1a transcript, which encodes the highly pathogenic and aggregation-prone HTT1a protein. This process provides a mechanistic link between CAG repeat expansion and disease pathogenesis, as increased HTT1a production accelerates HTT aggregation and neuronal dysfunction. HTT1a has previously been detected in Huntingtons disease mouse models by using immunoprecipitation coupled with western blotting, homogeneous time-resolved fluorescence (HTRF) and Meso Scale Discovery (MSD) bioassays, and immunohistochemistry. These approaches were developed using MW8, a neoepitope antibody that specifically recognizes the C-terminus of HTT1a. MW8 is a relatively weak antibody with limited detection sensitivity. To generate more robust HTT1a-specific reagents, two novel recombinant antibodies, 1B12 and 11G2, have been developed for evaluation. Using an allelic series of knock-in (HdhQ20, HdhQ50, HdhQ80, HdhQ111, CAG140 and zQ175) mice, alongside transgenic YAC128 and N171-82Q models, we extensively evaluated and compared the performance of MW8, 1B12 and 11G2. We demonstrate that 1B12 and 11G2 function as HTT1a-specific neoepitope antibodies by immunoprecipitation with western blotting, and by immunohistochemistry. To enhance HTT1a detection using HTRF and MSD technology platforms, we further evaluated the performance of 1B12 and 11G2 in HTT bioassays using cortical lysates from zQ175 and YAC128 mice. In zQ175 mice, enhanced detection of aggregated HTT1a by HTRF and MSD revealed that HTT fragments longer than HTT1a can be incorporated into HTT1a-containing aggregates. The most sensitive assays were subsequently applied across the allelic series of knock-in mice to assess the effect of polyglutamine length on bioassay performance. For optimal sensitivity, we recommend the preferential use of 1B12 for HTRF assays and 11G2 for MSD assays. Collectively, these findings establish 1B12 and 11G2 as robust antibodies to reliably detect and track HTT1a pathology in vivo and promotes the replacement of previously used MW8-based experimental approaches. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=191 HEIGHT=200 SRC="FIGDIR/small/708805v1_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@16d10faorg.highwire.dtl.DTLVardef@1759f41org.highwire.dtl.DTLVardef@12a8c21org.highwire.dtl.DTLVardef@55fe6a_HPS_FORMAT_FIGEXP M_FIG C_FIG Osborne et al. used Huntingtons disease mouse models to evaluate and compare the performance of HTT1a-specific neoepitope antibodies by using immunoprecipitation with western blotting, bioassays, and immunohistochemistry. In contrast to MW8, they establish that 1B12 and 11G2 are robust antibodies to reliably detect and track HTT1a pathology in vivo.

neuroscience↗

bioRxiv2026

A MET-Targeted Variable New Antigen Receptor Theranostic for Non-Small Cell Lung Cancer

The MET receptor tyrosine kinase is mutated or amplified in [~]6% of non-small cell lung cancer (NSCLC) and overexpressed in [~]80% of all NSCLC cases. A theranostic agent that can both see and treat MET-altered NSCLC has never been described before in the literature. Here, we report a shark-derived single-domain variable new antigen receptor (VNAR) for MET with theranostic applications. Following the immunization of a juvenile nurse shark (Ginglymostoma cirratum) with the extracellular domain of human MET, we identified a VNAR clone that specifically engaged MET with high affinity. Engineering the lead VNAR into a bivalent human Fc, vMET1-Fc, yielded a construct that selectively targeted and was internalized by MET-positive cells without affecting cell viability or downstream MET signaling. When radiolabeled with the positron emitting isotope Zr-89, [89Zr]Zr-vMET1-Fc enabled longitudinal PET/CT imaging. High tumor uptake with low background was observed in MET-positive NSCLC xenografts administered [89Zr]Zr-vMET1-Fc. As a targeted beta-particle radiotherapy, [{superscript 1}Lu]Lu-vMET1-Fc resulted in marked tumor-growth delay and exhibited a favorable toxicity profile, collectively improving progression-free survival in NSCLC mouse models. Non-human primate PET/CT imaging studies with ([Zr]Zr-vMET1-Fc in healthy rhesus macaques confirmed favorable biodistribution and dosimetry, predictable clearance, and minimal off-target uptake. Additional blood chemistry analysis found no significant immune response or cytotoxicity. Together, these findings establish vMET1-Fc as a theranostic agent for imaging and treating MET-altered NSCLC. Statement of SignificanceA shark-derived antibody selectively targeting MET shows preclinical efficacy as a theranostic agent for MET-altered cancer.

cancer biology↗

bioRxiv2025

Progressive Supranuclear Palsy PERK haplotype B selectively translates DLX1 promoting tau toxicity

The unfolded protein response (UPR) sensor PERK exists in two haplotypes termed A and B. PERK-B uniquely confers increased risk for tauopathies like progressive supranuclear palsy (PSP), but the mechanisms distinguishing its function from PERK-A and contributing to its association with tau pathology are not known. Here, we developed a controlled cellular model for a pair-wise comparison of the two PERK haplotypes, finding their UPR functions nearly indistinguishable. However, a careful examination employing puromycin-based proteomics revealed that a subset of mRNA translation events were permissible under PERK-B, but not PERK-A, dependent UPR. Critically, one of the targets that escaped PERK-B suppression was the transcription factor DLX1, which has been genetically linked to PSP risk. Here, we found a shift in the solubility of DLX1 in human PSP brain tissue, and report that silencing of DLX-1 reduced the aggregation of tau in mammalian cells. Furthermore, silencing of the fly homolog of DLX1 was sufficient to decrease tau-induced toxicity, in vivo. Our results detail the haplotype-specific PERK-B/DLX-1 pathway as a novel driver of tau pathology in cells, flies, and likely human brain, revealing new insights into PSP pathogenesis and potential therapeutic targets.

neuroscience↗

bioRxiv2025

Lowering the HTT1a transcript as an effective therapy for Huntingtons disease

Lowering the levels of HTT transcripts has been a major focus of therapeutic development for Huntingtons disease (HD), but which transcript should be lowered? HD is caused by a CAG repeat expansion in exon 1 of the HTT gene, and the rate of somatic expansion of this CAG repeat throughout life is now known to drive the age of onset and rate of disease progression. As the CAG repeat expands, the extent to which the HTT mRNA is alternatively processed to generate the HTT1a transcript and highly aggregation-prone and pathogenic HTT1a protein increases. Several HTT-lowering modalities have entered clinical trials that either target both HTT and HTT1a together, or full-length HTT alone. We have developed siRNAs that target the Htt1a mouse transcript (634/486) and used these, together with a potent Htt-targeting siRNA (10150) to compare the efficacy of lowering either full-length Htt or Htt1a. zQ175 and wild-type mice were treated with 10150 or 634/486 alongside control groups at 2 months of age with treatment to 6 or 10 months, or at 6 months with treatment to 10 months. The siRNA potency and durability were most effective in the hippocampus. Whilst both strategies showed benefits, despite the greater potency of 10150, targeting Htt1a was more effective at delaying HTT aggregation and transcriptional dysregulation than targeting full-length Htt. These data support HTT-lowering strategies that are designed to target the HTT1a transcript, either alone, or together with lowering full-length HTT. One Sentence SummaryLowering HTT1a transcript levels delays the onset of molecular and neuropathological phenotypes in a knock-in mouse model of Huntingtons disease.

neuroscience↗

bioRxiv2025

A method for efficient, rapid, and minimally invasive implantation of individual non-functional motes with penetrating subcellular-diameter carbon fiber electrodes into rat cortex

ObjectiveDistributed arrays of wireless neural interfacing chips with 1-2 channels each, known as "neural dust", could enhance brain machine interfaces (BMIs) by removing the wired connection through the scalp and increasing biocompatibility with their submillimeter size. Although several approaches for neural dust have emerged, a procedure for implanting them in batches that builds upon the safety and performance of currently used electrodes remains to be demonstrated. ApproachHere, we demonstrate the feasibility of implanting batches of wireless motes that rest on the cortical surface with carbon fiber electrodes of subcellular diameter (6.8-8.4 {micro}m) that penetrate to a target brain depth of 1 mm without insertion aids. To simulate their implantation, we assembled more than 230 mechanically equivalent motes and affixed them to insertion tools with polyethylene glycol (PEG), a quickly dissolvable and biocompatible material. Then, we implanted mote grids of multiple configurations into rat cortex in vivo and evaluated insertion success and their arrangement on the brain surface using photos and videos captured during their implantation. Main ResultsWhen placing motes onto the insertion device, we found that they aggregated in molten PEG such that the array pitch was only 5% wider than the dimensions of the mote bases themselves (240 x 240 {micro}m). Overall, we found that motes with this arrangement could be inserted into rat cortex with a high success rate, as 171/186 (92%) motes in 4x4 (N=4) and 5x5 (N=5) square grid configurations were successfully inserted using the insertion device alone. After implantation, measurements of how much motes tilted (22{+/-}9{degrees}, [X]{+/-}S) and had been displaced relative to their original positions were smaller than those measured for devices implanted inside the brain in the literature. SignificanceCollectively, these data establish the viability of safely implementing motes with ultrasmall electrodes and epicortically-situated chips for use in future BMIs.

bioengineering↗

bioRxiv2024

A Digital Microfluidic Platform for the Microscale Production of Functional Immune Cell Therapies

Genetically engineering human immune cells has been shown to be an effective approach for developing novel cellular therapies to treat a wide range of diseases. To expand the scope of these cellular therapies while solving persistent challenges, extensive research and development is still required. Electroporation has recently emerged as one of the most popular techniques for inserting biological payloads into human immune cells to perform genetic engineering. However, several recent studies have reported that electroporation can negatively impact cell functionality. Additionally, the requirement to use large amounts of cells and expensive payloads to achieve efficient delivery can drive up the costs of development efforts. Here we use a digital microfluidic enabled electroporation system (referred to as triDrop) and compare them against two state-of-the-art commercially available systems for the engineering of human T cells. We describe the ability to use triDrop for highly viable, highly efficient transfection while using substantially fewer cells and payload. Subsequently, we perform transcriptomic analysis on cells engineered with each of the three systems and show that electroporation with triDrop lead to less dysregulation of several functionally relevant pathways. Finally, in a direct comparison of immunotherapeutic functionality, we show that T cells engineered with triDrop have an improved ability to mount an immune response when presented with tumor cells. These results show that the triDrop platform is uniquely suited to produce functionally engineered immune cells while also reducing the costs of cell engineering compared to other commercially available systems.

bioengineering↗

bioRxiv2024

Myocardial infarction injury is exacerbated by nicotine in vape aerosol exposure

RationaleVaping is touted as a safer alternative to traditional cigarette smoking but the full spectrum of harm reduction versus comparable risk remains unresolved. Elevated bioavailability of nicotine in vape aerosol together with known risks of nicotine exposure may result in previously uncharacterized cardiovascular consequences of vaping. ObjectiveAssess the impact of nicotine exposure via vape aerosol inhalation upon myocardial response to infarction injury. Methods and ResultsFlavored vape juice containing nicotine (5 mg / ml) or vehicle alone (0 mg) was delivered using identical 4-week treatment protocols. Mice were subjected to acute myocardial infarction injury and evaluated for outcomes of cardiac structure and function. Findings reveal that nicotine exposure leads to worse outcomes with respect to contractile performance regardless of sex. Non-myocyte interstitial cell accumulation following infarction significantly increased with exposure to vape aerosol alone, but a comparable increase was not present when nicotine was included. ConclusionsMyocardial function after infarction is significantly decreased after exposure to nicotine vape aerosol irrespective of sex. Comparable loss of contractile function was not observed in mice exposed to vape aerosol alone, highlighting the essential role of nicotine in loss of contractile function. Increased vimentin immunoreactivity was observed in the vape alone group compared to control and vape nicotine. The correlation between vaping, interstitial cell responses, and cardiac remodeling leading to impaired contractility warrants further investigation. Public health experts seeking to reduce vaping-related health risks should consider messaging that highlights the increased cardiovascular risk especially with nicotine-containing aerosols.

molecular biology↗

bioRxiv2024

Mutant HTT protein decreases with CAG repeat expansion: implications for therapeutics and bioassays.

Huntingtons disease is an inherited neurodegenerative disorder caused by a CAG repeat expansion that encodes a polyglutamine tract in the HTT protein. The mutant CAG repeat is unstable and expands in specific brain cells and peripheral tissues throughout life. Genes involved in the DNA mismatch repair pathways, known to act on expansion, have been identified as genetics modifiers, therefore, it is the rate of somatic CAG repeat expansion that drives the age of onset and rate of disease progression. In the context of an expanded CAG repeat, the HTT pre-mRNA can be alternatively processed to generate the HTT1a transcript, that encodes the aggregation prone and highly pathogenic HTT1a protein. This may be a mechanism through which somatic CAG repeat expansion exerts its pathogenic effects, as the longer the CAG repeat, the more HTT1a and HTT1a is produced. The allelic series of knock-in mouse models: HdhQ20, HdhQ50, HdhQ80, HdhQ111, CAG140 and zQ175 with polyQ expansions of 20, 50, 80, 111 140 and [~]190 can be used to model the molecular and cellular consequences of CAG repeat expansion within a single neuron. By western blot of cortical lysates, we found that mutant HTT levels decreased with increasing CAG repeat length; mutant HTT was only 23% and 10% of wild-type levels in CAG140 and zQ175 cortices, respectively. To identify the optimal bioassays for detecting the full-length HTT and HTT1a isoforms, we interrogated the pairwise combinations of seven well-characterized antibodies on both the HTRF and MSD platforms. In total we tested 32 HTRF and 32 MSD assays to detect full-length mutant HTT, HTT1a, total mutant HTT (full-length HTT and HTT1a) and total full-length HTT (mutant and wild type). None of these assays recapitulated the full-length mutant HTT levels as measured by western blot. We recommend using isoform- and species-specific assays that detect either full-length mutant HTT, HTT1a or wild-type HTT as opposed to those that detect more than one isoform simultaneously. Our finding that as the CAG repeat expands, full-length mutant HTT levels decrease, whilst HTT1a and HTT1a levels increase has implications for therapeutic strategies. If mutant HTT levels in cells containing (CAG)200 are only 10% of wild-type, HTT-lowering strategies targeting full-length HTT at sequences 3 to intron 1 HTT will predominantly lower wild-type HTT, as mutant HTT levels in these cells are already depleted. These data support a therapeutic strategy that lowers HTT1a and depletes levels of the HTT1a protein.

neuroscience↗

bioRxiv2023

A CAG repeat threshold for therapeutics targeting somatic instability in Huntington's disease

The Huntingtons disease mutation is a CAG repeat expansion in the huntingtin gene that results in an expanded polyglutamine tract in the huntingtin protein. The CAG repeat is unstable, and expansions of hundreds of CAGs have been detected in Huntingtons disease post-mortem brains. The age of disease onset can be predicted partially from the length of the CAG repeat as measured in blood. Onset age is also determined by genetic modifiers, which in six cases involve variation in DNA mismatch repair pathways genes. Knocking-out specific mismatch repair genes in mouse models of Huntingtons disease prevents somatic CAG repeat expansion. Taken together, these results have led to the hypothesis that somatic CAG repeat expansion in Huntingtons disease brains is required for pathogenesis. Therefore, the pathogenic repeat threshold in brain is longer than (CAG)40, as measured in blood, and is currently unknown. The mismatch repair gene MSH3 has become a major focus for therapeutic development, as unlike other mismatch repair genes, nullizygosity for MSH3 does not cause malignancies associated with mismatch repair deficiency. Potential treatments targeting MSH3 currently under development include gene therapy, biologics and small molecules, which will be assessed for efficacy in mouse models of Huntingtons disease. The zQ175 knock-in model carries a mutation of approximately (CAG)185 and develops early molecular and pathological phenotypes that have been extensively characterised. Therefore, we crossed the mutant huntingtin allele onto heterozygous and homozygous Msh3 knock-out backgrounds to determine the maximum benefit of targeting Msh3 in this model. Ablation of Msh3 prevented somatic expansion throughout the brain and periphery, and reduction of Msh3 by 50% decreased the rate of expansion. This had no effect on the deposition of huntingtin aggregation in the nuclei of striatal neurons, nor on the dysregulated striatal transcriptional profile. This contrasts with ablating Msh3 in knock-in models with shorter CAG repeat expansions. Therefore, further expansion of a (CAG)185 repeat in striatal neurons does not accelerate the onset of molecular and neuropathological phenotypes. It is striking that highly expanded CAG repeats of a similar size in humans cause disease onset before 2 years of age, indicating that somatic CAG repeat expansion in the brain is not required for pathogenesis. Given that the trajectory for somatic CAG expansion in the brains of Huntingtons disease mutation carriers is unknown, our study underlines the importance of administering treatments targeting somatic instability as early as possible.

neuroscience↗

bioRxiv2023

Challenges in the discovery of tumor-specific alternative splicing-derived cell-surface antigens in glioma

BackgroundDespite advancements in cancer immunotherapy, solid tumors remain formidable challenges. In glioma, profound inter-and intra-tumoral heterogeneity of antigen landscape hampers therapeutic development. Therefore, it is critical to consider alternative sources to expand the repertoire of targetable (neo-)antigens and improve therapeutic outcomes. Accumulating evidence suggests that tumor-specific alternative splicing (AS) could be an untapped reservoir of neoantigens. ResultsIn this study, we investigated tumor-specific AS events in glioma, focusing on those predicted to generate major histocompatibility complex (MHC)-presentation-independent, cell-surface neoantigens that could be targeted by antibodies and chimeric antigen receptor (CAR)-T cells. We systematically analyzed bulk RNA-sequencing datasets comparing 429 tumor samples (from The Cancer Genome Atlas [TCGA]) and 9,166 normal tissue samples (from the Genotype-Tissue Expression project [GTEx]), and identified 13 AS events in 7 genes predicted to be expressed in more than 10% of the patients, including PTPRZ1 and BCAN, which were corroborated by an external RNA-sequencing dataset. Subsequently, we validated our predictions and elucidated the complexity of the isoforms using full-length transcript amplicon sequencing on patient-derived glioblastoma cells. However, analyses of the RNA-sequencing datasets of spatially mapped and longitudinally collected clinical tumor samples unveiled remarkable spatiotemporal heterogeneity of the candidate AS events. Furthermore, proteomics analysis did not reveal any peptide spectra matching the putative neoantigens. ConclusionsOur investigation illustrated the diverse characteristics of the tumor-specific AS events and the challenges of antigen exploration due to their notable spatiotemporal heterogeneity and elusive nature at the protein levels. Redirecting future efforts toward intracellular, MHC-presented antigens could offer a more viable avenue.

cancer biology↗

bioRxiv2023

Wireless in vivo Recording of Cortical Activity by an Ion-Sensitive Field Effect Transistor

Wireless brain technologies are empowering basic neuroscience and clinical neurology by offering new platforms that minimize invasiveness and refine possibilities during electrophysiological recording and stimulation. Despite their advantages, most systems require on-board power supply and sizeable transmission circuitry, enforcing a lower bound for miniaturization. Designing new minimalistic architectures that can efficiently sense neurophysiological events will open the door to standalone microscale sensors and minimally invasive delivery of multiple sensors. Here we present a circuit for sensing ionic fluctuations in the brain by an ion-sensitive field effect transistor that detunes a single radiofrequency resonator in parallel. We establish sensitivity of the sensor by electromagnetic analysis and quantify response to ionic fluctuations in vitro. We validate this new architecture in vivo during hindpaw stimulation in rodents and verify correlation with local field potential recordings. This new approach can be implemented as an integrated circuit for wireless in situ recording of brain electrophysiology.

neuroscience↗

bioRxiv2023

Acute effects of subanesthetic ketamine on cerebrovascular hemodynamics in humans: A TD-fNIRS neuroimaging study

Quantifying neural activity in natural conditions (i.e. conditions comparable to the standard clinical patient experience) during the administration of psychedelics may further our scientific understanding of the effects and mechanisms of action. This data may facilitate the discovery of novel biomarkers enabling more personalized treatments and improved patient outcomes. In this single-blind, placebo-controlled study with a non-randomized design, we use time-domain functional near-infrared spectroscopy (TD-fNIRS) to measure acute brain dynamics after intramuscular subanesthetic ketamine (0.75 mg/kg) and placebo (saline) administration in healthy participants (n = 15, 8 females, 7 males, age 32.4 {+/-} 7.5 years) in a clinical setting. We found that the ketamine administration caused an altered state of consciousness and changes in systemic physiology (e.g. increase in pulse rate and electrodermal activity). Furthermore, ketamine led to a brain-wide reduction in the fractional amplitude of low frequency fluctuations (fALFF), and a decrease in the global brain connectivity of the prefrontal region. Lastly, we provide preliminary evidence that a combination of neural and physiological metrics may serve as predictors of subjective mystical experiences and reductions in depressive symptomatology. Overall, our studies demonstrated the successful application of fNIRS neuroimaging to study the physiological effects of the psychoactive substance ketamine and can be regarded as an important step toward larger scale clinical fNIRS studies that can quantify the impact of psychedelics on the brain in standard clinical settings.

neuroscience↗

bioRxiv2022

JGI Plant Gene Atlas: An updateable transcriptome resource to improve structural annotations and functional gene descriptions across the plant kingdom

Gene functional descriptions, which are typically derived from sequence similarity to experimentally validated genes in a handful of model species, offer a crucial line of evidence when searching for candidate genes that underlie trait variation. Plant responses to environmental cues, including gene expression regulatory variation, represent important resources for understanding gene function and crucial targets for plant improvement through gene editing and other biotechnologies. However, even after years of effort and numerous large-scale functional characterization studies, biological roles of large proportions of protein coding genes across the plant phylogeny are poorly annotated. Here we describe the Joint Genome Institute (JGI) Plant Gene Atlas, a public and updateable data resource consisting of transcript abundance assays from 2,090 samples derived from 604 tissues or conditions across 18 diverse species. We integrated across these diverse conditions and genotypes by analyzing expression profiles, building gene clusters that exhibited tissue/condition specific expression, and testing for transcriptional modulation in response to environmental queues. For example, we discovered extensive phylogenetically constrained and condition-specific expression profiles across many gene families and genes without any functional annotation. Such conserved expression patterns and other tightly co-expressed gene clusters let us assign expression derived functional descriptions to 64,620 genes with otherwise unknown functions. The ever-expanding Gene Atlas resource is available at JGI Plant Gene Atlas (https://plantgeneatlas.jgi.doe.gov) and Phytozome (https://phytozome-next.jgi.doe.gov), providing bulk access to data and user-specified queries of gene sets. Combined, these web interfaces let users access differentially expressed genes, track orthologs across the Gene Atlas plants, graphically represent co-expressed genes, and visualize gene ontology and pathway enrichments.

plant biology↗

bioRxiv2022

A view of the pan-genome of domesticated cowpea (Vigna unguiculata Walp.)

Cowpea, Vigna unguiculata L. Walp., is a diploid warm-season legume of critical importance as both food and fodder in sub-Saharan Africa. This species is also grown in Northern Africa, Europe, Latin America, North America, and East to Southeast Asia. To capture the genomic diversity of domesticates of this important legume, de novo genome assemblies were produced for representatives of six sub-populations of cultivated cowpea identified previously from genotyping of several hundred diverse accessions. In the most complete assembly (IT97K-499-35), 26,026 core and 4,963 noncore genes were identified, with 35,436 pan genes when considering all seven accessions. GO-terms associated with response to stress and defense response were highly enriched among the noncore genes, while core genes were enriched in terms related to transcription factor activity, and transport and metabolic processes. Over 5 million SNPs relative to each assembly and over 40 structural variants >1 Mb in size were identified by comparing genomes. Vu10 was the chromosome with the highest frequency of SNPs, and Vu04 had the most structural variants. Noncore genes harbor a larger proportion of potentially disruptive variants than core genes, including missense, stop gain, and frameshift mutations; this suggests that noncore genes substantially contribute to diversity within domesticated cowpea. Article SummaryThis study reports annotated genome assemblies of six cowpea accessions. Together with the previously reported annotated genome of IT97K-499-35, these constitute a pan-genome resource representing six subpopulations of domesticated cowpea. Annotations include genes, variant calls for SNPs and short indels, larger presence or absence variants, and inversions. Noncore genes are enriched for loci involved in stress response and harbor many genic variants with potential effects on coding sequence.

genomics↗

bioRxiv2021

#COVIDisAirborne: AI-Enabled Multiscale Computational Microscopy of Delta SARS-CoV-2 in a Respiratory Aerosol

We seek to completely revise current models of airborne transmission of respiratory viruses by providing never-before-seen atomic-level views of the SARS-CoV-2 virus within a respiratory aerosol. Our work dramatically extends the capabilities of multiscale computational microscopy to address the significant gaps that exist in current experimental methods, which are limited in their ability to interrogate aerosols at the atomic/molecular level and thus ob-scure our understanding of airborne transmission. We demonstrate how our integrated data-driven platform provides a new way of exploring the composition, structure, and dynamics of aerosols and aerosolized viruses, while driving simulation method development along several important axes. We present a series of initial scientific discoveries for the SARS-CoV-2 Delta variant, noting that the full scientific impact of this work has yet to be realized. ACM Reference FormatAbigail Dommer1{dagger}, Lorenzo Casalino1{dagger}, Fiona Kearns1{dagger}, Mia Rosenfeld1, Nicholas Wauer1, Surl-Hee Ahn1, John Russo,2 Sofia Oliveira3, Clare Morris1, AnthonyBogetti4, AndaTrifan5,6, Alexander Brace5,7, TerraSztain1,8, Austin Clyde5,7, Heng Ma5, Chakra Chennubhotla4, Hyungro Lee9, Matteo Turilli9, Syma Khalid10, Teresa Tamayo-Mendoza11, Matthew Welborn11, Anders Christensen11, Daniel G. A. Smith11, Zhuoran Qiao12, Sai Krishna Sirumalla11, Michael OConnor11, Frederick Manby11, Anima Anandkumar12,13, David Hardy6, James Phillips6, Abraham Stern13, Josh Romero13, David Clark13, Mitchell Dorrell14, Tom Maiden14, Lei Huang15, John McCalpin15, Christo- pherWoods3, Alan Gray13, MattWilliams3, Bryan Barker16, HarindaRajapaksha16, Richard Pitts16, Tom Gibbs13, John Stone6, Daniel Zuckerman2*, Adrian Mulholland3*, Thomas MillerIII11,12*, ShantenuJha9*, Arvind Ramanathan5*, Lillian Chong4*, Rommie Amaro1*. 2021. #COVIDisAirborne: AI-Enabled Multiscale Computational Microscopy ofDeltaSARS-CoV-2 in a Respiratory Aerosol. In Supercomputing 21: International Conference for High Perfor-mance Computing, Networking, Storage, and Analysis. ACM, New York, NY, USA, 14 pages. https://doi.org/finalDOI

biophysics↗

bioRxiv2021

Deletion of Abi3/Gngt2 influences age-progressive amyloid β and tau pathologies in distinctive ways

The S209F variant of Abelson Interactor Protein 3 (ABI3) increases risk for Alzheimers disease (AD), but little is known about ABI3 function. RNAscope showed Abi3 is expressed in microglial and non-microglial cells, though its increased expression appears to be driven in plaque-associated microglia. Here, we evaluated Abi3-/- mice and document that both Abi3 and its overlapping gene, Gngt2, are disrupted in these mice. Expression of Abi3 and Gngt2 are tightly correlated, and elevated, in rodent models of AD. RNA-seq of the Abi3-Gngt2-/- mice revealed robust induction of an AD-associated neurodegenerative signature, including upregulation of Trem2, Plcg2 and Tyrobp. In APP mice, loss of Abi3-Gngt2 resulted in a gene dose- and age-dependent reduction in A{beta} deposition. Additionally, in Abi3-Gngt2-/- mice, expression of a pro-aggregant form of human tau exacerbated tauopathy and astrocytosis. Further, the AD-associated S209F mutation alters the extent of ABI3 phosphorylation. These data provide an important experimental framework for understanding the role of Abi3-Gngt2 function in AD. Our studies also demonstrate that manipulation of glial function could have opposing effects on amyloid and tau pathology, highlighting the unpredictability of targeting such pathways in AD.

neuroscience↗

bioRxiv2021

Intelligent Resolution: Integrating Cryo-EM with AI-driven Multi-resolution Simulations to Observe the SARS-CoV-2 Replication-Transcription Machinery in Action

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) replication transcription complex (RTC) is a multi-domain protein responsible for replicating and transcribing the viral mRNA inside a human cell. Attacking RTC function with pharmaceutical compounds is a pathway to treating COVID-19. Conventional tools, e.g., cryo-electron microscopy and all-atom molecular dynamics (AAMD), do not provide sufficiently high resolution or timescale to capture important dynamics of this molecular machine. Consequently, we develop an innovative workflow that bridges the gap between these resolutions, using mesoscale fluctuating finite element analysis (FFEA) continuum simulations and a hierarchy of AI-methods that continually learn and infer features for maintaining consistency between AAMD and FFEA simulations. We leverage a multi-site distributed workflow manager to orchestrate AI, FFEA, and AAMD jobs, providing optimal resource utilization across HPC centers. Our study provides unprecedented access to study the SARS-CoV-2 RTC machinery, while providing general capability for AI-enabled multi-resolution simulations at scale.

biophysics↗