Biology subjects

Chang, Y.-C.

Publications and source records attributed to Chang, Y.-C..

At least 37 records · Page 2Linked to original sources

bioRxiv2024

Discovery and Engineering of Retrons for Precise GenomeEditing

Retrons are promising gene editing tools because they can produce multi-copy single-stranded DNA in cells via self-primed reverse transcription. However, their potential for inserting genetic cargos in eukaryotes remains largely unexplored. Here we report the discovery and engineering of highly efficient retron-based gene editors for mammalian cells and vertebrates. Through bioinformatic analysis of metagenomic data and functional screening, we identified novel retron reverse transcriptases (RTs) that are highly active in mammalian cells. Rational design further improved the editing efficiency to levels comparable with conventional single-stranded oligodeoxynucleotide donors but from a genetically encoded cassette. Small molecule inhibitors of non-homologous end joining factors and Cas9-DNA repair protein fusions further increase homology-directed repair. Retron editors also exhibited robust activity with Cas12a nuclease and Cas9 nickase, expanding the genomic target scope and bypassing the need for a DNA double-stranded break. Using a rationally engineered retron editor, we incorporate a split GFP epitope tag for live cell imaging. Finally, we develop an all-RNA delivery strategy to enable DNA-free gene editing in cells and vertebrate embryos. This work establishes retron editors as a versatile and efficient tool for precise gene editing, offering new opportunities for biotechnology and biomedical research.

molecular biology↗

bioRxiv2024

Peripheral glia and neurons jointly regulate activity-induced synaptic remodeling at the Drosophila neuromuscular junction

In the nervous system, reliable communication depends on the ability of neurons to adaptively remodel their synaptic structure and function in response to changes in neuronal activity. While neurons are the main drivers of synaptic plasticity, glial cells are increasingly recognized for their roles as active modulators. However, the underlying molecular mechanisms remain unclear. Here, using Drosophila neuromuscular junction as a model system for a tripartite synapse, we show that peripheral glial cells collaborate with neurons at the NMJ to regulate activity-induced synaptic remodeling, in part through a protein called shriveled (Shv). Shv is an activator of integrin signaling previously shown to be released by neurons during intense stimulation at the fly NMJ to regulate activity-induced synaptic remodeling. We demonstrate that Shv is also present in peripheral glia, and glial Shv is both necessary and sufficient for synaptic remodeling. However, unlike neuronal Shv, glial Shv does not activate integrin signaling at the NMJ. Instead, it regulates synaptic plasticity in two ways: 1) maintaining the extracellular balance of neuronal Shv proteins to regulate integrin signaling, and 2) controlling ambient extracellular glutamate concentration to regulate postsynaptic glutamate receptor abundance. Loss of glial cells showed the same phenotype as loss of Shv in glia. Together, these results reveal that neurons and glial cells homeostatically regulate extracellular Shv protein levels to control activity-induced synaptic remodeling. Additionally, peripheral glia maintains postsynaptic glutamate receptor abundance and contribute to activity-induced synaptic remodeling by regulating ambient glutamate concentration at the fly NMJ.

neuroscience↗

bioRxiv2024

Massively Parallel Polyribosome Profiling Reveals Translation Defects of Human Disease-Relevant UTR Mutations

The untranslated regions (UTRs) of mRNAs harbor regulatory elements influencing translation efficiency. Although 3.7% of disease-relevant human mutations occur in UTRs, their exact role in pathogenesis remains unclear. Through metagene analysis, we mapped pathogenic UTR mutations to regions near coding sequences, with a focus on the upstream open reading frame (uORF) initiation site. Subsequently, we utilized massively parallel poly(ribo)some profiling to compare the ribosome associations of 6,555 pairs of wildtype and mutant UTR fragments. We identified 46 UTR variants that altered polysome profiles, with enrichment in pathogenic mutations. Both univariate analysis and the elastic net regression model highlighted the significance of motifs of short repeated sequences, including SRSF2 binding sites, as mutation hotspots that lead to aberrant translation. Furthermore, these polysome-shifting mutations exhibited considerable impact on RNA secondary structures, particularly for upstream AUG-containing 5 UTRs. Integrating these features, our model achieved high accuracy (AUROC > 0.8) in predicting polysome-shifting mutations in the test dataset. Additionally, several lines of evidence indicate that changes in uORF usage underlie the translation deficiency arising from these mutations. Illustrating this, we demonstrate that a pathogenic mutation in the IRF6 5 UTR suppresses translation of the primary open reading frame by creating a uORF. Remarkably, site- directed ADAR editing of the mutant mRNA rescued this translation deficiency. Overall, our study provides insights into the molecular mechanisms of UTR mutations and their links to clinical impacts through translation defects. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=187 SRC="FIGDIR/small/589132v2_ufig1.gif" ALT="Figure 1000"> View larger version (50K): org.highwire.dtl.DTLVardef@57217corg.highwire.dtl.DTLVardef@575410org.highwire.dtl.DTLVardef@15013bforg.highwire.dtl.DTLVardef@408840_HPS_FORMAT_FIGEXP M_FIG C_FIG

genetics↗

bioRxiv2024

Multiplexed Assays of Human Disease-relevant Mutations Reveal UTR Dinucleotide Composition as a Major Determinant of RNA Stability

UTRs contain crucial regulatory elements for RNA stability, translation and localization, so their integrity is indispensable for gene expression. Approximately 3.7% of genetic variants associated with diseases occur in UTRs, yet a comprehensive understanding of UTR variant functions remains limited due to inefficient experimental and computational assessment methods. To systematically evaluate the effects of UTR variants on RNA stability, we established a massively parallel reporter assay on 6,555 UTR variants reported in human disease databases. We examined the RNA degradation patterns mediated by the UTR library in two cell lines, and then applied LASSO regression to model the influential regulators of RNA stability. We found that UA dinucleotides and UA-rich motifs are the most prominent destabilizing element. Gain of UA dinucleotide outlined mutant UTRs with reduced stability. Studies on endogenous transcripts indicate that high UA-dinucleotide ratios in UTRs promote RNA degradation. Conversely, elevated GC content and protein binding on UA dinucleotides protect high-UA RNA from degradation. Further analysis reveals polarized roles of UA- dinucleotide-binding proteins in RNA protection and degradation. Furthermore, the UA- dinucleotide ratio of both UTRs is a common characteristic of genes in innate immune response pathways, implying a coordinated stability regulation through UTRs at the transcriptomic level. We also demonstrate that stability-altering UTRs are associated with changes in biobank-based health indices, underscoring the importance of precise UTR regulation for wellness. Our study highlights the importance of RNA stability regulation through UTR primary sequences, paving the way for further exploration of their implications in gene networks and precision medicine.

genetics↗

bioRxiv2024

Live imaging of Alu elements reveals non-uniform euchromatin dynamics coupled to transcription

Chromatin structure and dynamics are crucial for eukaryotic nuclear functions. Hi-C and related genomic assays have revealed chromatin conformations, such as A/B compartments, in fixed cells, but the dynamic motion of such structures is not well understood. Moreover, elucidating the relationship between the motion of chromatin and transcriptional activity is hampered by a lack of tools for specifically measuring the mobility of active euchromatin. Here, we introduce a CRISPR-based strategy for live imaging of the gene-rich A compartment by labeling Alu elements -- a retrotransposon family enriched within the transcriptionally active A compartment. Surprisingly, within euchromatin, microscopy analysis reveals that Alu-rich regions do not correlate with lower local H2B density, and form irregular foci of a few hundred nanometers in diameter, underscoring the heterogeneity of euchromatin organization. Alu-rich (gene-rich) chromatin is also more mobile than Alu-poor (gene-poor) chromatin, and transcription inhibition by actinomycin D results in decreased chromatin mobility of Alu-rich regions. These observations highlight the complexity of chromatin organization and dynamics and connect them to transcriptional activity on a genome-wide scale.

biophysics↗

bioRxiv2023

Identification of secretory autophagy as a novel mechanism modulating activity-induced synaptic remodeling

The ability of neurons to rapidly remodel their synaptic structure and strength in response to neuronal activity is highly conserved across species and crucial for complex brain functions. However, mechanisms required to elicit and coordinate the acute, activity-dependent structural changes across synapses are not well understood. Here, using an RNAi screen in Drosophila against genes affecting nervous system functions in humans, we uncouple cellular processes important for synaptic plasticity from synapse development. We find mutations associated with neurodegenerative and mental health disorders are 2-times more likely to affect activity-induced synaptic remodeling than synapse development. We further demonstrate that neuronal activity stimulates autophagy activation but diminishes degradative autophagy, thereby driving the pathway towards autophagy-based secretion. Presynaptic knockdown of Snap29, Sec22, or Rab8, proteins implicated in the secretory autophagy pathway, is sufficient to abolish activity-induced synaptic remodeling. This study uncovers secretory autophagy as a novel trans-synaptic signaling mechanism modulating structural plasticity.

neuroscience↗

bioRxiv2023

Validation of human telomere length trans-ancestry meta-analysis association signals identifies POP5 and KBTBD6 as novel human telomere length regulation genes

Telomere length genome-wide association studies (GWAS) have become well-powered to detect novel genes in telomere length regulation. However, no prior work has validated these putative novel genes to confirm the contribution of GWAS loci to telomere length regulation. We conducted a trans-ancestry meta-analysis of 211,369 individuals. Through enrichment analyses of chromatin state and cell-type heritability we identified blood and immune cells as the most relevant cell type to examine telomere length association signals. We validated specific GWAS associations by overexpressing KBTBD6, a component of an E3 ubiquitin ligase complex, and POP5, a component of the Ribonuclease P/MRP complex, and demonstrating that both lengthened telomeres as predicted by our statistical analyses. CRISPR/Cas9 deletion of the predicted causal regions of these association peaks in K562 immortalized blood cells reduced expression of these genes, demonstrating that these loci are related to transcriptional regulation of KBTBD6 and POP5, respectively. Together our results demonstrate the utility of telomere length GWAS in the identification of novel telomere length regulation mechanisms and highlight the importance of the proteasome-ubiquitin pathway in telomere length regulation.

genetics↗

bioRxiv2023

A scalable platform for efficient CRISPR-Cas9 chemical-genetic screens of DNA damage-inducing compounds

Current approaches to define chemical-genetic interactions (CGIs) in human cell lines are resource-intensive. We designed a scalable chemical-genetic screen platform by generating a DNA damage response (DDR)-focused custom sgRNA library. We performed five proof-of-principle compound screens and found that the compounds known modes-of-action (MoA) were enriched among the compounds CGIs. These scalable screens recapitulated expected CGIs at a comparable signal-to-noise ratio (SNR) relative to genome-wide screens. Furthermore, time-resolved CGIs, captured by sequencing screens at various time points, suggested an unexpected, late time point interstrand-crosslinking (ICL) repair pathway response to camptothecin-induced DNA damage. Our approach can facilitate screening compounds at scale and produce biologically informative CGI profiles.

systems biology↗

bioRxiv2023

PGSbuilder: An end-to-end platform for human genome association analysis and polygenic risk score predictions

Understanding the genetic basis of human complex diseases is increasingly important in the development of precision medicine. Over the last decade, genome-wide association studies (GWAS) have become a key technique for detecting associations between common diseases and single nucleotide polymorphisms (SNPs) present in a cohort of individuals. Alternatively, the polygenic risk score (PRS), which often applies results from GWAS summary statistics, is calculated for the estimation of genetic propensity to a trait at the individual level. Despite many GWAS and PRS tools being available to analyze a large volume of genotype data, most clinicians and medical researchers are often not familiar with the bioinformatics tools and lack access to a high-performance computing cluster resource. To fill this gap, we provide a publicly available web server, PGSbuilder, for the GWAS and PRS analysis of human genomes with variant annotations. The user-friendly and intuitive PGSbuilder web server is developed to facilitate the discovery of the genetic variants associated with complex traits and diseases for medical professionals with limited computational skills. For GWAS analysis, PGSbuilder provides the most renowned analysis tool PLINK 2.0 package. For PRS, PGSbuilder provides six different PRS methods including Clumping and Thresholding, Lassosum, LDPred2, GenEpi, PRS-CS, and PRSice2. Furthermore, PGSbuilder provides an intuitive user interface to examine the annotated functional effects of variants from known biomedical databases and relevant literature using advanced natural language processing approaches. In conclusion, PGSbuilder offers a reliable platform to aid researchers in advancing the public perception of genomic risk and precision medicine for human disease genetics. PGSbuilder is freely accessible at http://pgsb.tw23.org.

bioinformatics↗

bioRxiv2023

Condensate-driven interfacial forces reposition DNA loci and measure chromatin viscoelasticity

Biomolecular condensates assemble in living cells through phase separation and related phase transitions. An underappreciated feature of these dynamic molecular assemblies is that they form interfaces with cellular structures, including membranes, cytoskeleton, DNA and RNA, and other membraneless compartments. These interfaces are expected to give rise to capillary forces, but there are few ways of quantifying and harnessing these forces in living cells. Here, we introduce VECTOR (ViscoElastic Chromatin Tethering and ORganization), which uses light-inducible biomolecular condensates to generate capillary forces at targeted DNA loci. VECTOR can be utilized to programmably reposition genomic loci on a timescale of seconds to minutes, quantitatively revealing local heterogeneity in the viscoelastic material properties of chromatin. These synthetic condensates are built from components that naturally form liquid-like structures in living cells, highlighting the potential role for native condensates to generate forces and do work to reorganize the genome and impact chromatin architecture.

biophysics↗

bioRxiv2022

A di-arginine additive for dissociation of gold nanoparticle aggregates: A matrix-insensitive approach with applications in protease detection

We report the reversible aggregation of gold nanoparticle (AuNPs) assemblies via a diarginine peptide additive and thiolated PEGs (HS-PEGs). The AuNPs were first aggregated by attractive forces between the citrate-capped surface and the arginine side chains. We found that HS-PEG thiol group has higher affinity for the AuNPs surface, thus leading to redispersion and colloidal stability. In turn, there was a robust and obvious color change due to on/off plasmonic coupling. The assemblies dissociation was directly related to the HS-PEG structural properties such as their size or charge. As an example, HS-PEGs with a molecular weight below 1 kDa could dissociate 100% of the assemblies and restore the exact optical properties of the initial AuNPs suspension (prior to the assembly). Surprisingly, the dissociation capacity of HS-PEGs was not affected by the composition of the operating medium and could be performed in complex matrices such as plasma, saliva, bile, urine, cell lysates or even sea water. The high affinity of thiols for the gold surface encompasses by far the one of endogenous molecules and is thus favorized. Moreover, starting with AuNPs already aggregated ensured the absence of background signal as the dissociation of the assemblies was far from spontaneous. Remarkably, it was possible to dry the AuNPs assemblies and to solubilize them back with HS-PEGs, improving the colorimetric signal generation. We used this system for protease sensing in biological fluid. Trypsin was chosen as model enzyme and highly positively charged peptides were conjugated to HS-PEG molecules as cleavage substrate. The increase of positive charge of the HS-PEG-peptide conjugate quenched the dissociation capacity of the HS-PEG molecules which could only be restored by the proteolytic cleavage. Picomolar limit of detection was obtained as well as the detection in saliva or urine. TOC O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=111 SRC="FIGDIR/small/508454v1_ufig1.gif" ALT="Figure 1"> View larger version (44K): org.highwire.dtl.DTLVardef@13a6f3org.highwire.dtl.DTLVardef@35b42dorg.highwire.dtl.DTLVardef@1c8249forg.highwire.dtl.DTLVardef@dc8236_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2022

Rapidly Inferring Personalized Neurostimulation Parameters with Meta-Learning: A Case Study of Individualized Fiber Recruitment in Vagus Nerve Stimulation

ObjectiveNeurostimulation is emerging as treatment for several diseases of the brain and peripheral organs. Due to variability arising from placement of stimulation devices, underlying neuroanatomy and physiological responses to stimulation, it is essential that neurostimulation protocols are personalized to maximize efficacy and safety. Building such personalized protocols would benefit from accumulated information in increasingly large datasets of other individuals responses. ApproachTo address that need, we propose a meta-learning family of algorithms to conduct few-shot optimization of key fitting parameters of physiological and neural responses in new individuals. While our method is agnostic to neurostimulation setting, here we demonstrate its effectiveness on the problem of physiological modeling of fiber recruitment during vagus nerve stimulation (VNS). Using data from acute VNS experiments, the mapping between amplitudes of stimulus-evoked compound action potentials (eCAPs) and physiological responses, such as heart rate and breathing interval modulation, is inferred. Main resultsUsing additional synthetic data sets to complement experimental results, we demonstrate that our meta-learning framework is capable of directly modeling the physiology-eCAP relationship for individual subjects with much fewer individually queried data points than standard methods. SignificanceOur meta-learning framework is general and can be adapted to many input-response neurostimulation mapping problems. Moreover, this method leverages information from growing data sets of past patients, as a treatment is deployed. It can also be combined with several model types, including regression, Gaussian processes with Bayesian optimization, and beyond.

neuroscience↗

bioRxiv2022

UHMK1 is a novel splicing regulatory kinase

The U2AF Homology Motif Kinase 1 (UHMK1) is the only kinase that contains the U2AF homology motif (UHM), a common protein interaction domain among splicing factors. Through this motif, UHMK1 interacts with the splicing factors SF1 and SF3B1, known to participate in the 3 splice site recognition during the early steps of spliceosome assembly. Although UHMK1 phosphorylates these splicing factors in vitro, the involvement of UHMK1 in RNA processing has not previously been demonstrated. Here, we identify novel putative substrates of this kinase and evaluate UHMK1 contribution to overall gene expression and splicing, by integrating global phosphoproteomics, RNA-seq, and bioinformatics approaches. Upon UHMK1 modulation, 163 unique phosphosites were differentially phosphorylated in 117 proteins, of which 106 are novel potential substrates of this kinase. Gene Ontology (GO) analysis showed enrichment of terms previously associated with UHMK1 function, such as mRNA splicing, cell cycle, cell division and microtubule organization. The majority of the annotated RNA-related proteins are components of the spliceosome, but are also involved in several steps of gene expression. Comprehensive analysis of splicing showed that UHMK1 affected over 200 alternative splicing events. Moreover, splicing reporter assay further supported UHMK1 function on splicing. Overall, RNA-seq data demonstrated that UHMK1 knockdown had a minor impact on gene expression and pointed to UHMK1 function in epithelial-mesenchymal transition. Finally, the functional assays demonstrated that UHMK1 modulation affects proliferation, colony formation, and migration of the cells. Taken together, our data implicate UHMK1 as a splicing regulatory kinase, connecting protein regulation through phosphorylation and gene expression in key cellular processes.

molecular biology↗

bioRxiv2022

BANKSY: A Spatial Omics Algorithm that Unifies Cell Type Clustering and Tissue Domain Segmentation

Each cell type in a solid tissue has a characteristic transcriptome and spatial arrangement, both of which are observable using modern spatial omics assays. However, the common practice is still to ignore spatial information when clustering cells to identify cell types. In fact, spatial location is typically considered only when solving the related, but distinct, problem of demarcating tissue domains (which could include multiple cell types). We present BANKSY, an algorithm that unifies cell type clustering and domain segmentation by constructing a product space of cell and neighbourhood transcriptomes, representing cell state and microenvironment, respectively. BANKSYs spatial kernel-based feature augmentation strategy improves per-formance and scalability on both tasks when tested on FISH-based and sequencing-based spatial omics data. Uniquely, BANKSY identified hitherto undetected niche-dependent cell states in two mouse brain regions. Lastly, we show that quality control of spatial omics data can be formulated as a domain identification problem and solved using BANKSY. BANKSY represents a biologically motivated, scalable, and versatile framework for analyzing spatial omics data.

bioinformatics↗

bioRxiv2022

Nr6a1 controls axially-restricted body elongation, segmentation, patterning and lineage allocation

The vertebrate main-body axis is laid down during embryonic stages in an anterior-to-posterior (head-to-tail) direction, driven and supplied by posteriorly located progenitors. For the vertebral column, the process of axial progenitor cell expansion that drives elongation, and the process of segmentation which allocates progenitor-descendants into repeating pre-vertebral units, occurs seemingly uninterrupted from the first to the last vertebra. Nonetheless, there is clear developmental and evolutionary support for two discrete modules controlling processes within different axial regions: a trunk and a tail module. Here, we identify Nuclear receptor subfamily 6 group A member 1 (Nr6a1) as a master regulator of elongation, segmentation, patterning and lineage allocation specifically within the trunk region of the mouse. Both gain- and loss-of-function in vivo analyses revealed that the precise level of Nr6a1 acts as a rheostat, expanding or contracting vertebral number of the trunk region autonomously from other axial regions. Moreover, Nr6a1 was found to be required for segmentation, but only for trunk-forming somites, with the timely clearance of Nr6a1 critical in supporting tail formation. In parallel with these morphological outcomes, we reveal Nr6a1 as a novel regulator of global Hox signatures within axial progenitors, preventing the precocious expression of multiple posterior Hox genes as the trunk is being laid down and thus reinforcing that patterning and elongation are coordinated. Finally, our data supports a crucial role for Nr6a1 in regulating gene regulatory networks that guide cell lineage choice of axial progenitors between neural and mesodermal fate. Collectively, these data reveal an axially-restricted role for Nr6a1 in all major cellular and tissue-level events required for vertebral column formation, supporting the view that modulation of Nr6a1 expression level or function is likely to underpin evolutionary changes in axial formulae that exclusively alter the trunk region.

developmental biology↗

bioRxiv2022

Organ- and function-specific organization and bioelectronic modulation of vagus fascicles and fibers

Vagal fibers travel inside fascicles and form branches to innervate organs and regulate organ functions. Vagus nerve stimulation (VNS) therapies activate fibers non-selectively, often resulting in reduced efficacy and side effects from non-targeted organs. Transverse and longitudinal arrangement of fibers according to functions they mediate and organs they innervate is unknown, however it is crucial for selective VNS. Using microcomputed tomography, we found that, in swine, fascicles are arranged in 2 overlapping axes, with sensory and motor fascicles separated cephalad and merging caudad, and larynx-, heart- and lung-specific fascicles separated caudad and progressively merging cephalad. Using immunohistochemistry, we found that the distribution of single fibers is highly nonuniform: myelinated afferents and efferents occupy separate fascicles, unmyelinated efferents co-localize with myelinated afferents, and small unmyelinated afferents are widely distributed. Using a multi-contact cuff electrode, we delivered fascicular cervical VNS in anesthetized and awake swine. Compound action potentials, from distinct fiber types, and organ responses, including laryngeal muscle, cough, breathing, heart rate and blood pressure responses are elicited in a radially asymmetric manner, with consistent angular separations. These results indicate that vagal fibers are anatomically organized according to functions they mediate and organs they innervate and can be asymmetrically activated by fascicular cervical VNS.

neuroscience↗

bioRxiv2021

Dynamin-2 Phosphorylation as A Critical Regulatory Target of Bin1 and GSK3α for Endocytosis in Muscle

Tight regulation of endocytosis ensures accurate control of cellular signaling and membrane dynamics, which are crucial for tissue morphogenesis and functions. Mutations of Bin1 and dynamin-2 (Dyn2), proteins that generate membrane curvature and sever endocytic invaginations, respectively, cause progressive hereditary myopathy. Here, we show that Bin1 inhibits Dyn2 via direct interaction of its SRC Homology 3 (SH3) domain with the proline-rich domain (PRD) of Dyn2. Phosphorylation of S848 of Dyn2 by GSK3, a kinase downstream of insulin signaling, relieves Dyn2 from the inhibition of Bin1 and promotes endocytosis in muscle. Mutations of Bin1 associated with centronuclear myopathy disrupt its inhibition of Dyn2, thereby exaggerating Dyn2 fission activity and causing excessive fragmentation of T-tubules in the muscle cells. Our work reveals how Bin1-Dyn2 interaction fine-tunes membrane remodeling at the molecular level, and lay the foundation for future exploration of endocytic regulation and hereditary muscle diseases.

cell biology↗

bioRxiv2021

Precise control of microtubule disassembly in living cells

Microtubules (MTs) are components of the evolutionarily conserved cytoskeleton, which tightly regulates various cellular activities. Our understanding of MTs is largely based on MT-targeting agents, which, however, are insufficient to dissect the dynamic mechanisms of specific MT populations due to their slow effects on the entire pool of MTs in cells. To address this limitation, we have used chemogenetics and optogenetics to disassemble specific MT subtypes by rapid recruitment of engineered MT-cleaving enzymes. Acute MT disassembly swiftly halted vesicular trafficking and lysosome dynamics. We also used this approach to disassemble MTs specifically modified by tyrosination and several MT-based structures including primary cilia, mitotic spindles, and intercellular bridges. These effects were rapidly reversed by inhibiting the activity or MT association of the cleaving enzymes. The disassembly of targeted MTs with spatial and temporal accuracy enables to uncover new insights of how MTs precisely regulate cellular architectures and functions.

cell biology↗