Kuo, N. | Search bioRxiv

bioRxiv2023

A Novel Semi-automated Proofreading and Mesh Error Detection Pipeline for Neuron Extension

AO_SCPLOWBSTRACTC_SCPLOWThe immense scale and complexity of neuronal electron microscopy (EM) datasets pose significant challenges in data processing, validation, and interpretation, necessitating the development of efficient, automated, and scalable error-detection methodologies. This paper proposes a novel approach that employs mesh processing techniques to identify potential error locations near neuronal tips. Error detection at tips is a particularly important challenge since these errors usually indicate that many synapses are falsely split from their parent neuron, injuring the integrity of the connectomic reconstruction. Additionally, we draw implications and results from an implementation of this error detection in a semi-automated proofreading pipeline. Manual proofreading is a laborious, costly, and currently necessary method for identifying the errors in the machine learning based segmentation of neural tissue. This approach streamlines the process of proofreading by systematically highlighting areas likely to contain inaccuracies and guiding proofreaders towards potential continuations, accelerating the rate at which errors are corrected.

neuroscience↗

bioRxiv2021

Acitretin mitigates uroporphyrin-induced bone defects in congenital erythropoietic porphyria models

Congenital erythropoietic porphyria (CEP) is a rare genetic disorder leading to accumulation of uro/coproporphyrin-I in tissues due to inhibition of uroporphyrinogen-III synthase. Clinical manifestations of CEP include bone fragility, severe photosensitivity and photomutilation. Currently there is no specific treatment for CEP, except bone marrow transplantation, and there is an unmet need for treating this orphan disease. Fluorescent porphyrins cause protein aggregation, which led us to hypothesize that uroporphyrin-I accumulation leads to protein aggregation and CEP-related bone phenotype. We developed a zebrafish model that phenocopies features of CEP. As in human patients, uroporphyrin-I accumulated in the bones of zebrafish, leading to impaired bone development. Furthermore, in an osteoblast-like cell line, uroporphyrin-I decreased mineralization, aggregated bone matrix proteins, activated endoplasmic reticulum stress and disrupted autophagy. Using high-throughput drug screening, we identified acitretin, a second-generation retinoid, and showed that it reduced uroporphyrin-I accumulation and its deleterious effects on bones. Our findings provide a new CEP experimental model and a potential repurposed therapeutic.

pathology↗

bioRxiv2018

Resolving the hypotheticome: annotating M. tuberculosis gene function through bibliomic reconciliation and structural modeling

Accurate and timely functional genome annotation is essential for translating basic pathogen research into clinically impactful advances. Here, through literature curation and structure-function inference, we systematically update the functional genome annotation of Mycobacterium tuberculosis virulent type strain H37Rv. First, we systematically curated annotations for 589 genes from 662 publications, including 282 gene products absent from leading databases. Second, we modeled 1,711 under-annotated proteins and developed a semi-automated pipeline that captured shared function between 400 protein models and structural matches of known function on protein data bank, including drug efflux proteins, metabolic enzymes, and virulence factors. In aggregate, these structure- and literature-derived annotations update 940/1,725 under-annotated H37Rv genes and generate hundreds of functional hypotheses. Retrospectively applying the annotation to a recent whole-genome transposon mutant screen provided missing function for 48% (13/27) of under-annotated genes altering antibiotic efficacy and 33% (23/69) required for persistence during mouse TB infection. Prospective application of the protein models enabled us to functionally interpret novel laboratory generated Pyrazinamide-resistant (PZA) mutants of unknown function, which implicated the emerging Coenzyme A depletion model of PZA action in the mutants PZA resistance. Our findings demonstrate the functional insight gained by integrating structural modeling and systematic literature curation, even for widely studied microorganisms. Functional annotations and protein structure models are available at https://tuberculosis.sdsu.edu/H37Rv in human- and machine-readable formats. IMPORTANCEMycobacterium tuberculosis, the primary causative agent of tuberculosis, kills more humans than any other infectious bacteria. Yet 40% of its genome is functionally uncharacterized, leaving much about the genetic basis of its resistance to antibiotics, capacity to withstand host immunity, and basic metabolism yet undiscovered. Irregular literature curation for functional annotation contributes to this gap. We systematically curated functions from literature and structural similarity for over half of poorly characterized genes, expanding the functionally annotated Mycobacterium tuberculosis proteome. Applying this updated annotation to recent in vivo functional screens added functional information to dozens of clinically pertinent proteins described as having unknown function. Integrating the annotations with a prospective functional screen identified new mutants resistant to a first-line TB drug supporting an emerging hypothesis for its mode of action. These improvements in functional interpretation of clinically informative studies underscores the translational value of this functional knowledge. Structure-derived annotations identify hundreds of high-confidence candidates for mechanisms of antibiotic resistance, virulence factors, and basic metabolism; other functions key in clinical and basic tuberculosis research. More broadly, it provides a systematic framework for improving prokaryotic reference annotations.

genomics↗