Biology subjects

Haas, K.

Publications and source records attributed to Haas, K..

At least 19 recordsLinked to original sources

bioRxiv2026

Dynamo: An open-source Python application for quantifying neuronal structural plasticity over time

Neurons exhibit tremendous structural plasticity during the growth of dendritic and axonal arbors in early brain circuit formation, followed by experience-driven structural plasticity throughout life. Advances in labeling and in vivo time-lapse imaging allow capture of dynamic structural changes within intact and awake animals, and fluorescent biosensors of neural activity offer opportunities to link structural and functional plasticity. However, the resulting large multi-dimensional data sets are challenging to quantify due to time-consuming tracking of minute structural changes throughout complex neuronal morphologies across time. Here, we present Dynamo: an open-source Python application that enables dynamic morphometrics, the quantitative analysis of morphological changes over time, by streamlining arbor reconstruction, registration of structures across time, and quantitative analyses of growth behavior. Dynamo yields rich characterization of neural structural changes necessary for determining how rapid growth events culminate into long-term patterning, linking structural and functional plasticity, and for identifying underlying molecular mechanisms.

neuroscience↗

bioRxiv2026

Single-Cell Analytics for Dose Response (SCADR) discriminates PTEN missense variants by lipid and protein phosphatase dysfunction

The proliferation of sequencing efforts has revealed a vast and expanding catalog of single nucleotide gene variants, many associated to, but with unclear roles in disease. Fully charactering variant impacts and linking specific protein dysfunctions to disease are challenging due to the multi-functional nature of many proteins and varying degree of variant effects on these functions. Lagging are sensitive approaches to empirically assess the impact of missense variant-induced single amino acid changes on a wide range of protein functions. To address these issues, we have developed an open-source computational analysis tool called SCADR (Single-Cell Analytics for Dose Response) for simultaneously measuring and comparing impacts of exogenously-expressed variants on multiple signaling pathways using multiplex phospho-antibody spectral flow cytometry in human cell lines. SCADR retains and correlates single-cell measures of signal protein activity states along with expression levels of exogenously-expressed variants, providing rich characterization of multiple protein functions, signaling protein interactions, and enhanced discrimination of variant impacts on different signaling pathways, highlighting each variants unique dysfunction profile. Here, we apply SCADR for analyses of the impact of 6 variants of the tumor-suppressor protein PTEN (P38H, C124S, G129E, Y138L, D268E, 4A) expressed in HEK293 cells on the phosphorylation states of the canonical and noncanonical downstream signaling proteins Akt, S6, CREB, ERK, and p38 detected with fluorophore-conjugated phospho-antibodies, along with an antibody detecting an N-terminal HA tag on PTEN variants allowing measures of dose-response effects of each variants expression on signaling cascades. Results identify variant-specific impacts on downstream signaling cascades.

genomics↗

bioRxiv2026

Decision analysis in support of proactive planning for chronic wasting disease in Vermont, USA

Chronic wasting disease (CWD), a fatal, transmissible disease in white-tailed deer (Odocoileus virginianus) and related species, is spreading across North America but has not yet been detected in Vermont, United States (U.S.). The Vermont Department of Fish and Wildlife, along with partner agencies, wants to develop a proactive prevention and response plan in anticipation of the eventual detection of the disease. Between September 2023 and September 2025, staff from the U.S. Geological Survey and the University of Vermont facilitated a structured decision-making (SDM) process with seven State and Federal agencies that have jurisdiction over some aspect of CWD management in Vermont. The aim of this process was to generate and evaluate alternative response plans against a range of long-term objectives important to the agencies. To aid in the evaluation of the alternatives, we developed a linked set of models for white-tailed deer population and disease dynamics, hunter participation and health, forest health, economic consequences, and agricultural opportunities related to the actions being contemplated as part of the response plan. We evaluated over 256 different permutations of management actions and used multi-criteria decision analysis, a branch of decision analysis designed to help decision makers navigate tradeoffs among competing objectives, to summarize the performance of those alternative strategies against the desired outcomes. Proactive actions, those designed to slow the arrival of CWD to Vermont, were moderately effective, but the most important proactive action was surveillance to detect the disease early after arrival, which triggered response actions after detection. With the insights generated by the SDM process and the results of the analyses, the participating agencies were able to identify a preferred strategy and outline the elements of a proactive response plan. This report describes the SDM process, the technical details of the modeling work, and the results of the analyses.

ecology↗

bioRxiv2026

DksA-Dependent Stringent Stress Response Drives Virulence and Gastrointestinal Persistence of Klebsiella pneumoniae

Successful gastrointestinal colonization (GI) by bacterial pathogens requires adaptation to nutrient competition and host-derived stresses in the gut, with adaptation via the bacterial stringent stress response playing a critical role. Epidemiological data suggest that the GI tract serves as a reservoir from where K. pneumoniae can spread and cause invasive disease or transmit to another host. DksA is a conserved stringent response transcriptional regulator that was identified in an in vivo transposon mutagenesis screen as an important K. pneumoniae gut determinant. However, its role in K. pneumoniae pathogenesis and gut colonization remains uncharacterized. Here, we demonstrate that DksA is required for survival against membrane-targeting antibiotics, consistent with a role in cell envelope stress tolerance. In addition, DksA positively regulates capsule biosynthesis gene expression and hypermucoviscosity and is essential for robust biofilm formation. Using a murine model, we show that DksA functions as a determinant of GI colonization independently of the resident gut microbiota. Furthermore, we demonstrate that DksA is important for environmental survival and transmission by regulating RpoS, thereby providing a mechanistic link between the stringent stress response, environmental survival, and subsequent transmission. Together, these findings establish DksA as a central integrator of the stringent response, coordinating membrane stress resistance, virulence traits, and gastrointestinal colonization in K. pneumoniae. ImportanceK. pneumoniae, a pathobiont, is responsible for multidrug-resistant infections and poses a major threat in hospital settings as well as community-acquired invasive infections. The bacterium tightly coordinates its virulence-associated traits to adapt to diverse environmental conditions and survive; however, the regulatory mechanisms remain poorly understood. In this study, we demonstrated that the conserved stringent response regulator DksA contributes to bacterial membrane stability, thereby affecting antibiotic resistance, inherent virulence, and persistence traits of K. pneumoniae. Additionally, DksA was identified as required for gut colonization, environmental survival through dysregulation of RpoS, and transmission to a naive host. These results enhance our overall understanding of the K. pneumoniae stringent response and will provide new avenues for controlling K. pneumoniae infections.

microbiology↗

bioRxiv2025

Robust antibiotic sensitization of pathogenic Pseudomonas aeruginosa via negative hysteresis in the cell envelope

Antibiotic combination in time and space is a key strategy to combat antimicrobial resistance. The success of such treatment designs requires their robust efficacy across treatment conditions and a pathogens genomic diversity. This study found that an initial treatment with a {beta}-lactam antibiotic causes robust cellular sensitization towards an aminoglycoside antibiotic across the high-risk human pathogen Pseudomonas aeruginosa, including resistant strains. This phenomenon of cellular sensitization, termed negative hysteresis, is modulated by the Cpx envelope stress response system and linked to membrane stress during growth. The increase in efficacy is achieved through a {beta}-lactam induced elevated cellular uptake of the subsequently administered aminoglycoside. Negative hysteresis and the Cpx system are linked in several cases to the expression of synergistic drug interactions, thus enhancing efficacy of antibiotic combinations. Overall, our study identifies the phenomenon of negative hysteresis as a robustly inducible phenotype and thus a unique focus for optimizing antimicrobial therapy.

evolutionary biology↗

bioRxiv2025

Resistance variation and bacterial interactions shape the adaptation of a genetically diverse bacterial population to antimicrobial treatment

Bacterial infections are often polymicrobial and subject to the evolution of antimicrobial resistance (AMR). Existing knowledge on AMR in such polymicrobial infections usually relies on observational patient data, for which cause-effect relationships are difficult to infer, or on studying interactions between different bacterial species, ignoring the commonly encountered variation within species. Here, we therefore asked how mixed populations with strains from the same species evolve under antibiotic treatment. We used a genetically diverse population of the high-risk human pathogen Pseudomonas aeruginosa, and first identified strain variation in both AMR and pairwise bacteria-bacteria interactions, the latter ranging from beneficial, neutral, to competitive. Using experimental evolution, we subsequently demonstrate that the response to selection by different antibiotic treatments is significantly influenced by AMR strain variation, bacterial interactions, and also spatial population structure. Moreover, de novo AMR evolution was additionally impacted by variation in resistance rates towards the two considered antibiotics. A second evolution experiment emphasized the central role of strain variation and bacterial interactions in determining the evolutionary outcome. We conclude that ecological dynamics in genetically diverse pathogen populations are key for our general understanding of infection characteristics and AMR evolution, and, therefore, deserve particular attention during treatment of polymicrobial infections.

evolutionary biology↗

bioRxiv2024

Drug Metabolism and Transport Capacity of Endothelial Cells, Pericytes, and Astrocytes: Implications for CNS Drug Disposition

Therapeutically targeting the brain requires interactions with endothelial cells, pericytes, and astrocytes at the blood brain barrier (BBB). We evaluated regional and cell-type specific drug metabolism and transport mechanisms using rhesus macaques and in vitro treatment of primary human cells. Here, we report heterogenous distribution of representative drugs, tenofovir (TFV), emtricitabine (FTC), and their active metabolites, which cerebrospinal fluid measures could not reflect. We found that all BBB cell types possessed functional drug metabolizing enzymes and transporters that promoted TFV and FTC uptake and pharmacologic activation. Pericytes and astrocytes emerged as pharmacologically dynamic cells that rivaled hepatocytes and were uniquely susceptible to modulation by disease and treatment. Together, our findings demonstrate the importance of considering the BBB as a unique pharmacologic entity, rather than viewing it as an extension of the liver, as each cell type possesses distinct drug metabolism and transport capacities that contribute to differential brain drug disposition.

pharmacology and toxicology↗

bioRxiv2024

A change in the cell wall status initiates the elimination of the nucellus in Arabidopsis

The evolution of the seed habit coincides with a change in the cell fate of the nucellus, the sporophytic tissue responsible for female meiosis. Seeds arose when the nucellus retained the female spores instead of releasing them in the environment. As a consequence, the nucellus was partially eliminated to accommodate the growth of the female gametophyte inside the sporophyte. With the evolution of angiosperm seeds, the process of nucellus elimination was requisitioned to allow the growth of the endosperm, the second fertilization product devoted to store nutrients. Cell elimination differs from most known cell death programs as it leads to the apparent dismantling of the cell wall. Here, we show that nucellus elimination in Arabidopsis is initiated by the lysis of the pectic polysaccharides in the cell wall. This process exposes other cell wall components to possible further degradation and precedes a cell death program that leads to nuclear DNA fragmentation. Both pathways are regulated by TRANSPARENT TESTA 16, a MADS-domain transcription factor that evolved with seed plants. Finally, the causal effect of cell wall modification on nucellus development is demonstrated by inhibiting pectin degradation, thus suggesting that a change in the cell wall status might have driven seed evolution.

plant biology↗

bioRxiv2021

BrainPalmSeq: A curated RNA-seq database of palmitoylating and de-palmitoylating enzyme expression in the mouse brain

Protein S-palmitoylation is a reversible post-translational lipid modification that plays a critical role in neuronal development and plasticity, while dysregulated S-palmitoylation underlies a number of severe neurological disorders. Dynamic S-palmitoylation is regulated by a large family of ZDHHC palmitoylating enzymes, their accessory proteins, and a small number of known de-palmitoylating enzymes. Here, we curated and analyzed expression data for the proteins that mediate S-palmitoylation from publicly available RNAseq datasets, providing a comprehensive overview of their distribution in the mouse nervous system. We developed a web-tool that enables interactive visualization of the expression patterns for these proteins in the nervous system (http://brainpalmseq.med.ubc.ca/), and explored this resource to find region and cell-type specific expression patterns that give insight into the function of palmitoylating and de-palmitoylating enzymes in the brain and neurological disorders. We found coordinated expression of ZDHHC enzymes with their accessory proteins, de-palmitoylating enzymes and other brain-expressed genes that included an enrichment of S-palmitoylation substrates. Finally, we utilized ZDHHC expression patterns to predict and validate palmitoylating enzyme-substrate interactions.

neuroscience↗

bioRxiv2021

Type 1 interferon supports B cell responses to polysaccharide antigens but is not required for MPL/TDCM adjuvant effects on innate B cells

We previously described monophosphoryl lipid A (MPL) and synthetic cord factor, trehalose-6,6-dicorynomycolate (TDCM) significantly increases antibody (Ab) responses to T cell independent type 2 antigens (TI-2 Ags) in a manner dependent on B cell-intrinsic TLR4 expression as well as MyD88 and TRIF adapter proteins. Given the requirement for TRIF in optimal MPL/TDCM adjuvant effects and the capacity of MPL to drive type I IFN production, we aimed to investigate the extent to which adjuvant effects on TI-2 Ab responses depend on type I IFN receptor (IFNAR) signaling. We found IFNAR-/- mice had impaired early TI-2 Ag-induced B cell activation and expansion and that B cell-intrinsic type I IFN signaling on B cells was essential for normal antibody responses to TI-2 Ags, including haptenated Ficoll and the pneumococcal vaccine, Pneumovax23. However, MPL/TDCM significantly increased TI-2 IgM and IgG responses in IFNAR-/- mice. MPL/TDCM enhanced TI-2 Ab production primarily by activating innate B cells (B-1b and splenic CD23- B cells) as opposed to CD23+ enriched follicular B cells. In summary, our study highlights an important role for type I IFN in supporting early B cell responses to TI-2 Ags through B cell-expressed IFNAR, but nonetheless demonstrates an MPL/TDCM adjuvant significantly increases TI-2 Ab responses independently of type I IFN signaling and does so by predominantly supporting increased polysaccharide-specific Ab production by innate B cell populations. Key pointsO_LIB cell-intrinsic IFNAR expression promotes TI-2 Ab responses. C_LIO_LIMPL/TDCM adjuvant effects are independent of type 1 IFN. C_LIO_LIMPL/TDCM promotes TI-2 Ab responses by innate B cells. C_LI

immunology↗

bioRxiv2021

Comprehensive imaging of synaptic activity reveals dendritic growth rules that cluster inputs

The distribution of synapses across dendritic arbors determines their contribution to neural computations since nonlinear conductances amplify co-active clustered inputs. To determine whether, and how patterned synaptic topography arises during development we developed a random-access microscope capable of full-neuron calcium imaging of activity and structural plasticity of developing neurons in awake Xenopus tadpoles. By imaging growing brain neurons in response to plasticity-inducing visual training, we show coordinated growth and synaptogenesis specific to each neurons spike tuning. High evoked activity in neurons tuned to the trained stimulus induced pruning of non-driven inputs across the dendritic arbor as these neurons strengthened their responses to this stimulus. In stark contrast, initially unresponsive neurons that shifted their spike tuning toward the trained stimulus exhibited localized growth and new responsive synapses near existing active inputs. These information-driven growth rules promote clustering of synapses tuned to a developing neurons emerging receptive field. One-Sentence SummarySensory input directs brain neuronal growth and connectivity promoting clustering of synaptic inputs tuned to a neurons encoding properties.

neuroscience↗

bioRxiv2020

A small-molecule inhibitor of the BRCA2-RAD51 interaction modulates RAD51 assembly and potentiates DNA damage-induced cell death

BRCA2 controls RAD51 recombinase during homologous DNA recombination (HDR) through eight evolutionarily-conserved BRC repeats, which individually engage RAD51 via the motif Phe-x-x-Ala. Using structure-guided molecular design, templated on a monomeric thermostable chimera between human RAD51 and archaeal RadA, we identify CAM833, a 529 Da orthosteric inhibitor of RAD51:BRC with a Kd of 366 nM. The quinoline of CAM833 occupies a hotspot, the Phe-binding pocket on RAD51 and the methyl of the substituted -methylbenzyl group occupies the Ala-binding pocket. In cells, CAM833 diminishes formation of damage-induced RAD51 nuclear foci; inhibits RAD51 molecular clustering, suppressing extended RAD51 filament assembly; potentiates cytotoxicity by ionising radiation, augmenting 4N cell-cycle arrest and apoptotic cell death and works with poly-ADP ribose polymerase (PARP)1 inhibitors to suppress growth in BRCA2-wildtype cells. Thus, chemical inhibition of the protein-protein interaction between BRCA2 and RAD51 disrupts HDR and potentiates DNA damage-induced cell death, with implications for cancer therapy.

cancer biology↗

bioRxiv2020

Multi-parametric analysis of 58 SYNGAP1 variants reveal impacts on GTPase signaling, localization and protein stability

SYNGAP1 is a Ras and Rap GTPase with important roles in regulating excitatory synaptic plasticity. While many SYNGAP1 missense and nonsense mutations have been associated with intellectual disability, epilepsy, schizophrenia and autism spectrum disorder (ASD), there are many variants of unknown significance (VUS). In this report, we characterize 58 variants in nine assays that examine multiple aspects of SYNGAP1 function. Specifically, we used multiplex phospho-flow cytometry to measure the impact of variants on pERK, pGSK3{beta} and pCREB and high-content imaging to examine their subcellular localization. We find variants ranging from complete loss-of-function (LoF) to wildtype (WT)-like in their ability to regulate pERK and pGSK3{beta}, while all variants retain at least partial ability to regulate pCREB. Interestingly, our assays reveal that a high percentage of variants located within the disordered domain of unknown function that makes up the C-terminal half of SYNGAP1 exhibited LoF, compared to the more well studied catalytic domain. Moreover, we find protein instability to be a major contributor to dysfunction only for two missense variants both located within the catalytic domain. Using high-content imaging, we find variants with nuclear enrichment/exclusion and aberrant nuclear speckle localization. These variants are primarily located within the C2 domain known to mediate membrane lipid interactions. We find that mislocalization is distinct from altered catalytic activity, highlighting multiple independent molecular mechanisms underlying variant dysfunction. Our multidimensional dataset allows clustering of variants based on functional phenotypes and provides high-confidence pathogenicity classification.

genetics↗

bioRxiv2020

A SARS-CoV-2-Human Protein-Protein Interaction Map Reveals Drug Targets and Potential Drug-Repurposing

An outbreak of the novel coronavirus SARS-CoV-2, the causative agent of COVID-19 respiratory disease, has infected over 290,000 people since the end of 2019, killed over 12,000, and caused worldwide social and economic disruption1,2. There are currently no antiviral drugs with proven efficacy nor are there vaccines for its prevention. Unfortunately, the scientific community has little knowledge of the molecular details of SARS-CoV-2 infection. To illuminate this, we cloned, tagged and expressed 26 of the 29 viral proteins in human cells and identified the human proteins physically associated with each using affinity-purification mass spectrometry (AP-MS), which identified 332 high confidence SARS-CoV-2-human protein-protein interactions (PPIs). Among these, we identify 66 druggable human proteins or host factors targeted by 69 existing FDA-approved drugs, drugs in clinical trials and/or preclinical compounds, that we are currently evaluating for efficacy in live SARS-CoV-2 infection assays. The identification of host dependency factors mediating virus infection may provide key insights into effective molecular targets for developing broadly acting antiviral therapeutics against SARS-CoV-2 and other deadly coronavirus strains.

systems biology↗

bioRxiv2020

Hotspot KRAS mutations in brain metastases at the first metastatic recurrence of cutaneous melanoma

IMPORTANCEBrain metastases occur in 60% of patients with advanced melanoma and are a major cause of melanoma-related mortality and morbidity. Although our understanding of the molecular alterations associated with melanoma progression is improving, there are currently no validated biomarkers which might help identify those patients at highest risk of developing brain metastases. OBJECTIVETo examine the somatic mutational and copy-number landscape of brain metastases that develop as the isolated first visceral site of recurrence - "early brain-metastasis" compared to extracranial melanoma metastases. DESIGN, SETTING AND PARTICIPANTSWhole-exome sequencing of 50 tumors from patients undergoing surgical resection of one or more brain metastasis occurring as the first site of visceral relapse were identified from prospectively maintained databases in Sydney, Wellington, New York and Cambridge. Whole exome sequencing analyses allowed mutational profiles to be compared to cutaneous melanomas in The Cancer Genome Atlas (SKCM-TCGA; n=358) and the Memorial Sloan Kettering (SKCM-MSK-IMPACT; n=186) datasets. An external dataset comprising a further 18 patients with surgically resected early brain metastasis from two additional academic centers served as an independent validation cohort. MAIN OUTCOMES AND MEASURESTo assess the frequency of driver mutations in early brain metastasis and their influence on survival. RESULTSIn concordance with the landmark melanoma sequencing studies, we identified mutations in BRAF (21/50, 42%), NRAS (14/50, 28%) and NF1 (11/50, 22%) as the most frequently mutated melanoma driver genes. When compared to the mutational landscape of cutaneous melanomas in TCGA (SKCM-TCGA), KRAS was the most significantly enriched driver gene, with 5/50 (10%) of brain metastases harboring non-synonymous mutations, of which 4/5 (80%) were in the hotspot positions of codons 12 and 61. This was significantly higher than the corresponding frequency of KRAS-mutations within the entire SKCM-TCGA (2% (7/358), p=0.009, Fishers Exact Test) as well as the SKCM-MSK-IMPACT cohort (1.6% (3/186), p=0.016). Variants in KRAS were mutually exclusive from BRAFV600, NRAS and HRAS mutations and were associated with a significantly reduced overall survival from resection of brain metastasis (relative to KRAS-wild type brain metastases) in multivariate Cox proportional hazard models (HR 1.80, 95% CI 1.46-24.89, p=0.013). Mutations in KRAS were also clonal and concordant with extracranial disease, which suggests these mutations are present within the primary tumor CONCLUSIONS AND RELEVANCEOur analysis, the largest to date, suggests that early metastases to the brain (presenting as the first site of visceral relapse) are characterized by significant enrichment of hotspot KRAS mutations, potentially implicating constitutive RAS-driven cellular programs in neurotropic metastatic behavior in these cases. Based on these data, we suggest that screening for KRAS mutations might help identify those patients with primary melanoma at higher risk of brain metastases or poor survival, and could help inform future surveillance strategies. Key PointsO_ST_ABSQuestionC_ST_ABSWhat is the frequency of driver mutations in early melanoma brain metastases? FindingsIn this study of 50 patients with melanoma metastasizing first to the brain, KRAS mutations were the most significantly enriched driver gene (n=5, 10% of patients) when compared to landmark cutaneous melanoma studies. The high KRAS mutation frequency was also observed in an external validation cohort of 18 patients with early brain metastases. Mutations in KRAS were mutually exclusive from mutations in the key RAS signaling genes and conferred a worse overall survival from resection of brain metastasis. MeaningHotspot KRAS mutations could help identify those patients with primary melanoma at higher risk of brain metastases that may benefit from more intensive, protracted surveillance as well as earlier use of adjuvant therapy.

genomics↗

bioRxiv2020

Sentinel Interaction Mapping (SIM) - A generic approach for the functional analysis of human disease gene variants using yeast

Advances in sequencing technology have led to an explosion in the number of known genetic variants of human genes. A major challenge is to now determine which of these variants contribute to diseases as a result of their effect on gene function. Here we describe a generic approach using the yeast Saccharomyces cerevisiae to quickly develop gene-specific in vivo assays that can be used to quantify the level of function of a genetic variant. Using Synthetic Dosage Lethality screening, "sentinel" yeast strains are identified that are sensitive to overexpression of a human disease gene. Variants of the gene can then be functionalized in high-throughput fashion through simple growth assays using either solid or liquid media. Sentinel Interaction Mapping (SIM) has the potential to create functional assays for the large majority of human disease genes that do not have a yeast orthologue. Using the tumour suppressor gene PTEN as an example, we show that SIM assays can provide a fast and economical means to screen a large number of genetic variants.

genomics↗

bioRxiv2019

Multi-model functionalization of disease-associated PTEN missense mutations identifies multiple molecular mechanisms underlying protein dysfunction

Functional variomics provides the foundation for personalized medicine by linking genetic variation to disease expression, outcome and treatment, yet its utility is dependent on appropriate assays to evaluate mutation impact on protein function. To fully assess the effects of 106 missense and nonsense variants of PTEN associated with autism spectrum disorder, somatic cancer and PHTS, we take a deep phenotypic profiling approach using 18 assays in 5 model systems spanning diverse cellular environments ranging from molecular function to neuronal morphogenesis and behavior. Variants inducing instability occurred across the protein, resulting in partial to complete loss of function (LoF), which was well correlated across models. However, assays were selectively sensitive to variants located in substrate binding and catalytic domains, which exhibited complete LoF or dominant negativity independent of effects on stability. Our results indicate that full characterization of variant impact requires assays sensitive to instability and a range of protein functions.

genetics↗

bioRxiv2019

Systematic phenomics analysis of ASD-associated genes reveals shared functions and parallel networks underlying reversible impairments in habituation learning

A major challenge facing the genetics of Autism Spectrum Disorders (ASD) is the large and growing number of candidate risk genes and gene variants of unknown functional significance. Here, we used Caenorhabditis elegans to systematically functionally characterize ASD-associated genes in vivo. Using our custom machine vision system we quantified 26 phenotypes spanning morphology, locomotion, tactile sensitivity, and habituation learning in 87 strains each carrying a mutation in an ortholog of an ASD-associated gene. We identified hundreds of novel genotype-phenotype relationships ranging from severe developmental delays and uncoordinated movement to subtle deficits in sensory and learning behaviors. We clustered genes by similarity in phenomic profiles and used epistasis analysis to discover parallel networks centered on CHD8*chd-7 and NLGN3*nlg-1 that underlie mechanosensory hyper-responsivity and impaired habituation learning. We then leveraged our data for in vivo functional assays to gauge missense variant effect. Expression of wild-type NLG-1 in nlg-1 mutant C. elegans rescued their sensory and learning impairments. Testing the rescuing ability of all conserved ASD-associated neuroligin variants revealed varied partial loss-of-function despite proper subcellular localization. Finally, we used CRISPR-Cas9 auxin inducible degradation to determine that phenotypic abnormalities caused by developmental loss of NLG-1 can be reversed by adult expression. This work charts the phenotypic landscape of ASD-associated genes, offers novel in vivo variant functional assays, and potential therapeutic targets for ASD.

neuroscience↗