Biology subjects

Chang, C.-Y.

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

30 records · Page 2Linked to original sources

bioRxiv2022

Dissolvable microgel-templated macroporous hydrogels for controlled cell assembly

Mesenchymal stem cells (MSCs)-based therapies have been widely used to promote tissue regeneration and to modulate immune/inflammatory response. The therapeutic potential of MSCs can be further improved by forming multi-cellular spheroids. Meanwhile, hydrogels with macroporous structures are advantageous for improving mass transport properties for the cell-laden matrices. Herein, we report the fabrication of MSC-laden macroporous hydrogel scaffolds through incorporating rapidly dissolvable spherical cell-laden microgels. Dissolvable microgels were fabricated by tandem droplet-microfluidics and thiol-norbornene photopolymerization using a novel fast-degrading macromer poly(ethylene glycol)-norbornene-dopamine (PEGNB-Dopa). The cell-laden microgels were subsequently encapsulated within another bulk hydrogel matrix, whose porous structure was generated efficiently by the rapid degradation of the PEGNB-Dopa microgels. The cytocompatibility of this in situ pore-forming approach was demonstrated with multiple cell types. Furthermore, adjusting the stiffness and cell adhesiveness of the bulk hydrogels afforded the formation of solid cell spheroids or hollow spheres. The assembly of solid or hollow MSC spheroids led to differential activation of AKT pathway. Finally, MSCs solid spheroids formed in situ within the macroporous hydrogels exhibited robust secretion of HGF, VEGF-A, IL-6, IL-8, and TIMP-2. In summary, this platform provides an innovative method for forming cell-laden macroporous hydrogels for a variety of future biomedical applications.

bioengineering↗

bioRxiv2021

Confined migration induces heterochromatin formation and alters chromatin accessibility

During migration, cells often squeeze through small constrictions, requiring extensive deformation. We hypothesized that the nuclear deformation associated with such confined migration could alter chromatin organization and function. Studying cells migrating through microfluidic devices that mimic interstitial spaces in vivo, we found that confined migration results in increased H3K9me3 and H3K27me3 heterochromatin marks that persist for several days. This "confined migration-induced heterochromatin" (CMiH) was distinct from heterochromatin formation during migration initiation. Confined migration predominantly decreased chromatin accessibility at intergenic regions near centromeres and telomeres, suggesting heterochromatin spreading from existing heterochromatin sites. Consistent with the overall decrease in chromatin accessibility, global transcription was decreased during confined migration. Intriguingly, we also identified increased accessibility at promoter regions of genes linked to chromatin silencing, tumor invasion, and DNA damage response. Inhibiting CMiH reduced migration speed, suggesting that CMiH promotes confined migration. Together, our findings indicate that confined migration induces chromatin changes that regulate cell migration and other cellular functions.

cell biology↗

bioRxiv2021

Increased ACTL6A Occupancy Within mSWI/SNF Chromatin Remodelers Drives Human Squamous Cell Carcinoma

Mammalian SWI/SNF (BAF) chromatin remodelers play dosage-sensitive roles in many human malignancies and neurologic disorders. The gene encoding the BAF subunit, ACTL6A, is amplified at an early stage in the development of squamous cell carcinomas (SCCs), but its oncogenic role remains unclear. Here we demonstrate that ACTL6A overexpression leads to its stoichiometric assembly into BAF complexes and drives its interaction and engagement with specific regulatory regions in the genome. In normal epithelial cells, ACTL6A was sub-stoichiometric to other BAF subunits. However, increased ACTL6A levels by ectopic expression or in SCC cells led to near-saturation of ACTL6A within BAF complexes. Increased ACTL6A occupancy enhanced polycomb opposition over the genome activating SCC genes and also enhanced the recruitment of transcription factor TEAD with its co-activator YAP, promoting their chromatin binding and enhancer accessibility. Both of these mechanisms appeared to be critical and function as a molecular AND gate for SCC initiation and maintenance, thereby explaining the specificity of the role of ACTL6A amplification in SCCs. HighlightsO_LIACTL6A occupancy within BAF complexes is sub-stoichiometric in normal epithelial cells. C_LIO_LISCC cells upregulate ACTL6A thus increasing ACTL6A assembly with BAF complexes. C_LIO_LIGenome-wide chromatin profiling identifies ACTL6A-dependent regulatory regions. C_LIO_LIIncreasing ACTL6A incorporation enhances TEAD-YAP binding to BAF complexes. C_LIO_LIACTL6A overexpression counteracts polycomb-mediated repression at SCC signature genes. C_LI

molecular biology↗

bioRxiv2020

Top-down engineering of complex communities by directed evolution

Directed evolution has been used for decades to engineer biological systems from the top-down. Generally, it has been applied at or below the organismal level, by iteratively sampling the mutational landscape in a guided search for genetic variants of higher function. Above the organismal level, a small number of studies have attempted to artificially select microbial communities and ecosystems, with uneven and generally modest success. Our theoretical understanding of artificial ecosystem selection is still limited, particularly for large assemblages of asexual organisms, and we know little about designing efficient methods to direct their evolution. To address this issue, we have developed a flexible modeling framework that allows us to systematically probe any arbitrary selection strategy on any arbitrary set of communities and selected functions, in a wide range of ecological conditions. By artificially selecting hundreds of in-silico microbial metacommunities under identical conditions, we examine the fundamental limits of the two main breeding methods used so far, and prescribe modifications that significantly increase their power. We identify a range of directed evolution strategies that, particularly when applied in combination, are better suited for the top-down engineering of large, diverse, and stable microbial consortia. Our results emphasize that directed evolution allows an ecological structure-function landscape to be navigated in search for dynamically stable and ecologically and functionally resilient high-functioning communities.

evolutionary biology↗

bioRxiv2020

Fibroblast growth factor 9 stimulates neurite outgrowth through NF-kB signaling in striatal cell Huntington's disease models

Proper development of neuronal cells is important for brain functions, and impairment of neuronal development may lead to neuronal disorders, implying that improvement in neuronal development may be a therapeutic direction for these diseases. Huntingtons disease (HD) is a neurodegenerative disease characterized by impairment of neuronal structures, ultimately leading to neuronal death and dysfunctions of the central nervous system. Based on previous studies, fibroblast growth factor 9 (FGF9) may provide neuroprotective functions in HD, and FGFs may enhance neuronal development and neurite outgrowth. However, whether FGF9 can provide neuronal protective functions through improvement of neuronal morphology in HD is still unclear. Here, we study the effects of FGF9 on neuronal morphology in HD and attempt to understand the related working mechanisms. Taking advantage of striatal cell lines from HD knock-in mice, we found that FGF9 increases neurite outgrowth and upregulates several structural and synaptic proteins under HD conditions. In addition, activation of nuclear factor kappa B (NF-kB) signaling by FGF9 was observed to be significant in HD cells, and blockage of NF-kB leads to suppression of these structural and synaptic proteins induced by FGF9, suggesting the involvement of NF-kB signaling in these effects of FGF9. Taken these results together, FGF9 may enhance neurite outgrowth through upregulation of NF-kB signaling, and this mechanism could serve as an important mechanism for neuroprotective functions of FGF9 in HD.

neuroscience↗

bioRxiv2020

Artificially selecting microbial communities using propagule strategies

Artificial selection is a promising approach to manipulate the function of microbial communities. Here, we report the outcome of two artificial selection experiments at the microbial community level. Both experiments used "propagule" strategies, in which a set of the best-performing communities are used as the inocula to form a new generation of communities. In both cases, the selected communities are compared to a control treatment where communities are randomly selected. The first experiment used a defined set of strains as the starting inoculum, and the function under selection was the amylolytic activity of the consortia. The second experiment used a diverse set of natural communities as the inoculum, and the function under selection was the cross-feeding potential of the resulting communities towards a reference bacterial strain. In both experiments, the selected communities reached a higher mean and a higher maximum function than the control. In the first experiment this is caused by a decline in function of the control, rather than an improvement of the selected line. In the second experiment, the strong response of the mean is caused by the large initial variance in function across communities, and is the immediate consequence of the spread of the top-performing community in the starting group, whose function does not increase. Our results are in agreement with basic expectations of artificial selection theory, pointing out some of the limitations of community-level selection experiments which can inform the design of future studies.

evolutionary biology↗

bioRxiv2020

Metabolic rules of microbial community assembly

To develop a quantitative theory that can predict how microbiomes assemble, and how they respond to perturbations, we must identify which descriptive features of microbial communities are reproducible and predictable, which are unpredictable, and why. The emergent metagenomic structure of communities is often quantitatively convergent in similar habitats, with highly similar fractions of the metagenome being devoted to the same metabolic pathways. By contrast, the species-level taxonomic composition is often highly variable even in replicate environments. The mechanisms behind these patterns are not yet understood. By studying the self-assembly of hundreds of communities in replicate, synthetic habitats, we show that the reproducibility of microbial community assembly reflects an emergent metabolic structure, which is quantitatively predictable from first-principles, genome-scale metabolic models. Taxonomic variability within functional groups arises through multistability in population dynamics, and the species-level community composition is predictably governed by the mutual competitive exclusion of two sub-dominant strains. Our findings provide a mechanistic bridge between microbial community structure at different levels of organization, and show that the evolutionary conservation of metabolic traits, both in terms of growth responses and niches constructed, can be leveraged to quantitatively predict the taxonomic and metabolic structure of microbial communities.

ecology↗

bioRxiv2020

EXO70D isoforms mediate selective autophagic degradation of Type-A ARR proteins to regulate cytokinin sensitivity

The phytohormone cytokinin influences many aspects of plant growth and development, several of which also involve the cellular process of autophagy, including leaf senescence, nutrient re-mobilization, and developmental transitions. The Arabidopsis type-A Response Regulators (type-A ARR) are negative regulators of cytokinin signaling that are transcriptionally induced in response to cytokinin. Here, we describe a mechanistic link between cytokinin signaling and autophagy, demonstrating that plants modulate cytokinin sensitivity through autophagic regulation of type-A ARR proteins. Type-A ARR proteins were degraded by autophagy in an AUTOPHAGY-RELATED (ATG)5-dependent manner. EXO70D family members interacted with Type-A ARR proteins, likely in a phosphorylation-dependent manner, and recruited them to autophagosomes via interaction with the core autophagy protein, ATG8. Consistently, loss-of-function exo70D1,2,3 mutants compromised targeting of type-A ARRs to autophagic vesicles, have elevated levels of type-A ARR proteins, and are hyposensitive to cytokinin. Disruption of both type-A ARRs and EXO70D1,2,3 compromised survival in carbon-deficient conditions, suggesting interaction between autophagy and cytokinin responsiveness in response to stress. These results indicate that the EXO70D proteins act as selective autophagy receptors to target type-A ARR cargos for autophagic degradation, demonstrating that cytokinin signaling can be modulated by selective autophagy.

plant biology↗

bioRxiv2019

Enhancer regulation for induced WNT3A expression during neuronal regeneration

The treatment of traumatic brain injury (TBI) is limited by a lack of knowledge about the mechanisms underlying neuronal regeneration. WNT family members have been implicated in neurogenesis and aberrant WNT signaling has been associated with neurodegenerative diseases. The current study compared the expression of WNT genes during regeneration of injured cortical neurons. Recombinant WNT3A showed positive effect in promoting neuronal regeneration via in vitro and in vivo TBI models. Intranasal administration of WNT3A protein to TBI mice increased NeuN+ cells compared to control mice as well as retained motor function based on behavior analysis. Since TBI is known to reprogram the epigenome, chromatin immunoprecipitation-sequencing of histone H3K27ac and H3K4me3 was performed to address the transcriptional regulation of WNT3A during neuronal regeneration. We predicted, characterized and proposed that a histone H3K4me1-marked enhancer may undergo topological transformation to regulate the WNT3A gene expression.

molecular biology↗

bioRxiv2019

Dual linker UNC-10/SYD-2 is sufficient to bind kinesin-3 UNC-104 to RAB-3 containing synaptic vesicles in the absence of the motor’s PH domain

Kinesin-3 KIF1A (UNC-104 in C. elegans) is the major axonal transporter of synaptic vesicles and mutations in this molecular motor are linked to KIF1A-associated neurological disorders (KAND) including Charcot-Marie-Tooth disease, amyotrophic lateral sclerosis and hereditary spastic paraplegia. UNC-104 binds via its PH (pleckstrin homology) domain to the lipid bilayers of membranous vesicles which is considered a weak interaction. RT-PCR and Western blot experiments reveal genetic relations between SYD-2, UNC-10 and RAB-3. Co-immunoprecipitation assays reveal functional relations and bimolecular fluorescence complementation (BiFC) assays expose in situ interactions between these proteins. Though both SNB-1 and RAB-3 are actively transported by UNC-104, the movement of RAB-3 is generally enhanced and largely depending on the presence of SYD-2/UNC-10. Deletion of UNC-104s PH domain did not affect UNC-104/RAB-3 colocalization but did affect UNC-104/SNB-1 colocalization. Similarly, motility of RAB-3-labeled vesicles is unaltered in nematodes carrying a point mutation in the PH domain while movement of SNB-1 is significantly reduced in anterograde directions. These findings suggest a dual UNC-10/SYD-2 linker acting as a sufficient buttress to connect the motor to RAB-3-containing vesicles to enhance their transport. This additional linker will also strengthen the rather weak motor-lipid interaction. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=70 SRC="FIGDIR/small/723247v4_ufig1.gif" ALT="Figure 1"> View larger version (18K): org.highwire.dtl.DTLVardef@5e41c0org.highwire.dtl.DTLVardef@2ed9c8org.highwire.dtl.DTLVardef@1dbdb9dorg.highwire.dtl.DTLVardef@12f30c2_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2019

Sonogenetic modulation of cellular activities using an engineered auditory-sensing protein

Biomolecules that respond to different external stimuli enable the remote control of genetically modified cells. Chemogenetics and optogenetics, two tools that can control cellular activities via synthetic chemicals or photons, respectively, have been widely used to elucidate underlying physiological processes. These methods are, however, very invasive, have poor penetrability, or low spatiotemporal precision, attributes that hinder their use in therapeutic applications. We report herein a sonogenetic approach that can manipulate target cell activities by focused ultrasound stimulation. This system requires an ultrasound-responsive protein derived from an engineered auditory-sensing protein prestin. Heterogeneous expression of mouse prestin containing two parallel amino acid substitutions, N7T and N308S, that frequently exist in prestins from echolocating species endowed transfected mammalian cells with the ability to sense ultrasound. An ultrasound pulse of low frequency and low pressure efficiently evoked cellular calcium responses after transfecting with prestin(N7T, N308S). Moreover, pulsed ultrasound can also non-invasively stimulate target neurons expressing prestin(N7T, N308S) in deep regions of mice brains. Our study delineates how an engineered auditory-sensing protein can cause mammalian cells to sense ultrasound stimulation. Moreover, owing to the great penetration of low-frequency ultrasound ([~]400 mm in depth), our sonogenetic tools will serve as new strategies for non-invasive therapy in deep tissues of large animals like primates.

bioengineering↗

bioRxiv2019

Recapitulation and reversal of schizophrenia-related phenotypes in Setd1a-deficient mice

SETD1A, a histone methyltransferase, is a key schizophrenia susceptibility gene. Mutant mice carrying a heterozygous loss-of-function mutation of the orthologous gene exhibit alterations in axonal branching and cortical synaptic dynamics, accompanied by specific deficits in working memory that recapitulates SCZ-related alterations. We show that Setd1a targets mostly enhancers and reveal a striking overlap between Setd1a and Mef2 chromatin targets. Setd1a targets are highly expressed in pyramidal neurons and enriched for genes with postnatally-biased expression involved in synaptic structure and function. Notably, evolutionary conserved Setd1a binding sites and target genes are strongly associated with neuropsychiatric genetic risk burden. Reinstating Setd1a expression in adulthood rescues working memory deficits. We identify LSD1 as a major demethylase counteracting the effects of Setd1a methyl transferase activity and show that LSD1 antagonism in adult Setd1a-deficient mice results in a full rescue of the behavioral abnormalities and axonal branching deficits. Our findings advance our understanding of how SETD1A mutations predispose to SCZ and point to therapeutic interventions.

neuroscience↗