Biology subjects

Galaz-Montoya, J. G.

Publications and source records attributed to Galaz-Montoya, J. G..

6 recordsLinked to original sources

bioRxiv2024

CryoViT: Efficient Segmentation of Cryogenic Electron Tomograms with Vision Foundation Models

Cryogenic electron tomography (cryoET) directly visualizes subcellular structures in 3D at the nanometer scale. Quantitative analyses of cryoET data can reveal structural biomarkers of diseases, provide novel mechanistic insights, and inform the effects of treatments on phenotype. However, existing automated annotation approaches primarily focus on localizing molecular features with few methods accurately quantifying complex structures such as organelles. We address this challenge with CryoViT, a paradigm shift from traditional convolutional neural networks that leverages vision transformers to enhance the segmentation of large pleomorphic structures that can occupy almost the entire field of view in high-magnification images, such as mitochondria. CryoViT is powered by a large-scale vision foundation model and overcomes limitations of popular U-Net based methods, particularly when training data are scarce. We demonstrate the efficacy of CryoViT on a large cryoET dataset of neurons differentiated from iPSCs derived from Huntington disease (HD) patients and cultured HD mouse model neurons.

bioinformatics↗

bioRxiv2024

Missing Wedge Completion via Unsupervised Learning with Coordinate Networks

Cryogenic electron tomography (cryoET) is a powerful tool in structural biology, enabling detailed 3D imaging of biological specimens at a resolution of nanometers. Despite its potential, cryoET faces challenges such as the missing wedge problem, which limits reconstruction quality due to incomplete data collection angles. Recently, supervised deep learning methods leveraging convolutional neural networks (CNNs) have considerably addressed this issue; however, their pretraining requirements render them susceptible to inaccuracies and artifacts, particularly when representative training data is scarce. To overcome these limitations, we introduce a proof-of-concept unsupervised learning approach using coordinate networks (CNs) that optimizes network weights directly against input projections. This eliminates the need for pretraining, reducing reconstruction runtime by 3 - 20x compared to supervised methods. Our in silico results show improved shape completion and reduction of missing wedge artifacts, assessed through several voxel-based image quality metrics in real space and a novel directional Fourier Shell Correlation (FSC) metric. Our study illuminates benefits and considerations of both supervised and unsupervised approaches, guiding the development of improved reconstruction strategies.

molecular biology↗

bioRxiv2022

CryoET Reveals Organelle Phenotypes in Huntington Disease Patient iPSC-Derived and Mouse Primary Neurons

Huntingtons Disease (HD) is caused by an expanded CAG repeat in the huntingtin gene, yielding a Huntingtin protein with an expanded polyglutamine tract. Patient-derived induced pluripotent stem cells (iPSCs) can help understand disease; however, defining pathological biomarkers is challenging. Here, we used cryogenic electron tomography to visualize neurites in HD patient iPSC-derived neurons with varying CAG repeats, and primary cortical neurons from BACHD, deltaN17-BACHD, and wild-type mice. In HD models, we discovered mitochondria with enlarged granules and distorted cristae, and thin sheet aggregates in double membrane-bound organelles. We used artificial intelligence to quantify mitochondrial granules, and proteomics to show differential protein content in HD mitochondria. Knockdown of Protein Inhibitor of Activated STAT1 ameliorated aberrant phenotypes in iPSC-neurons and reduced phenotypes in BACHD neurons. We show that integrated ultrastructural and proteomic approaches may uncover early HD phenotypes to accelerate diagnostics and the development of targeted therapeutics for HD.

neuroscience↗

bioRxiv2021

Previously uncharacterized rectangular bacteria in the dolphin mouth

Much remains to be explored regarding the diversity of uncultured, host-associated microbes. Here, we report the discovery of unusual rectangular bacterial structures (RBSs) in the mouths of bottlenose dolphins. DNA staining revealed multiple paired bands within RBSs that suggested cells dividing along the longitudinal axis. Cryogenic transmission electron microscopy and tomography revealed parallel membrane-bound segments, suspected to be cells, encapsulated by an S-layer-like periodic surface covering. RBSs displayed novel pilus-like appendages with bundles of threads splayed at the tips. Multiple lines of evidence suggested that RBSs are bacterial and distinct from the Neisseriaceae genera Simonsiella and Conchiformibius, with which they share similar morphology and division patterning, including genomic DNA sequencing of micromanipulated RBSs, 16S rRNA gene sequencing, and fluorescence in situ hybridization. Our findings highlight the diversity of novel microbial forms and lifestyles that await discovery and characterization using tools complementary to genomics such as microscopy.

microbiology↗

bioRxiv2020

Topological Insights into Mutant Huntingtin Exon 1 and PolyQ Aggregates by Cryo-Electron Tomography

Huntington disease (HD) is a neurodegenerative trinucleotide repeat disorder caused by an expanded poly-glutamine (polyQ) tract in the mutant huntingtin (mHTT) protein. The formation and topology of filamentous mHTT inclusions in the brain (hallmarks of HD implicated in neurotoxicity) remain elusive. Using cryo-electron tomography and subtomogram averaging, here we show that mHTT exon 1 and polyQ-only aggregates in vitro are structurally heterogenous and filamentous, similar to prior observations with other methods. Yet, we observed some filaments in both types of aggregates under [~]2 nm in width, thinner than previously reported, while other regions form large sheets. In addition, our data show a prevalent subpopulation of filaments exhibiting a lumpy, slab-shaped morphology in both aggregates, supportive of the "polyQ core" model. This provides a basis for future cryoET studies of various aggregated mHTT and polyQ constructs to improve their structure-based modeling and their identification in cells without fusion tags.

molecular biology↗

bioRxiv2020

Multi-Scale 3D Cryo-Correlative Microscopy for Vitrified Cells

Three-dimensional (3D) visualization of vitrified cells can uncover structures of subcellular complexes without chemical fixation or staining. Here, we present a pipeline integrating three imaging modalities to visualize the same specimen at cryogenic temperature at different scales: cryo-fluorescence confocal microscopy, volume cryo-focused ion beam scanning electron microscopy, and transmission cryo-electron tomography. Our proof-of-concept benchmark revealed the 3D distribution of organelles and subcellular structures in whole heat-shocked yeast cells, including the ultrastructure of protein inclusions that recruit fluorescently-labelled chaperone Hsp104. Since our workflow efficiently integrates imaging at three different scales and can be applied to other types of cells, it could be used for large-scale phenotypic studies of frozen-hydrated specimens in a variety of healthy and diseased conditions with and without treatments.

cell biology↗