Biology subjects

An, S.

Publications and source records attributed to An, S..

50 records · Page 3Linked to original sources

bioRxiv2022

Functional regulation of 4D metabolic network between multienzyme glucosome condensates and mitochondria

Glucose metabolism is biochemically intertwined between energy metabolism and building block biosynthesis in living cells. However, it has not been investigated yet how its metabolic network is orchestrated to govern glucose flux in space and time. Since we reported that human enzymes in glucose metabolism are spatially organized into metabolically active membraneless compartments (i.e., glucosomes), we have employed lattice light sheet microscopic imaging and other biophysical and biochemical techniques to understand their functional significance in cellular metabolism. Now, we demonstrated that glucosome assemblies behave like liquid droplets in human cells and thus reversibly respond to environmental changes. In addition, we characterized a molecular architecture of the glucosome, which appears to be constructed from higher-ordered oligomeric structures of its scaffolder enzyme along with transient enzyme-enzyme interactions. Importantly, we found that enzymatic compositions of glucosomes are altered when they are spatially in proximity to mitochondria to functionally couple glycolysis with mitochondrial metabolism in human cells. Collectively, we envision that the subcellular localization-function relationship between glucosomes and mitochondria may represent one of fundamental principles by which 4-dimensional metabolic networks are not only dynamically but also efficiently regulated in living human cells. One Sentence SummaryInvestigation of a 4D functional network of glucose metabolism uncovers a fundamental principal of subcellular metabolic regulation in human cells.

biochemistry↗

bioRxiv2022

Regulation of EGF-stimulated activation of the PI-3K/AKT pathway by exocyst-mediated exocytosis

The phosphoinositide-3 kinase (PI-3K)/AKT cell survival pathway is an important pathway activated by EGFR signaling. Here we show, that in addition to previously described critical components of this pathway, i.e., the docking protein Gab1, the PI-3K/AKT pathway in epithelial cells is regulated by the exocyst complex, which is a vesicle tether that is essential for exocytosis. Using live-cell imaging, we demonstrate that PI(3,4,5)P3 levels fluctuate at the membrane on a minutes time scale and that these fluctuations are associated with local PI(3,4,5)P3 increases at sites where recycling vesicles undergo exocytic fusion. Supporting a role for exocytosis in PI(3,4,5)P3 generation, acute promotion of exocytosis by optogenetically driving exocyst-mediated vesicle tethering upregulates PI(3,4,5)P3 production and AKT activation. Conversely, acute inhibition of exocytosis using Endosidin2, a small-molecule inhibitor of the exocyst subunit Exo70, impairs PI(3,4,5)P3 production and AKT activation induced by EGF stimulation of epithelial cells. Moreover, prolonged inhibition of EGF signaling by EGFR tyrosine kinase inhibitors results in spontaneous reactivation of AKT without a concomitant relief of EGFR inhibition. However, this reactivation can be negated by acutely inhibiting the exocyst. These experiments demonstrate that exocyst-mediated exocytosis - by regulating PI(3,4,5)P3 levels at the plasma membrane - subserves activation of the PI-3K/AKT pathway by EGFR in epithelial cells.

cell biology↗

bioRxiv2022

Mosquito odorant receptor sensitive to natural spatial repellents and inhibitory compounds

Previously, we have identified an odorant receptor (OR) from the southern house mosquito Culex quinquefasciatus, CquiOR32, which responded to both odorants (agonists) and inhibitory compounds (antagonists). CquiOR32/CquiOrco-expressing oocytes responded to methyl salicylate and other odorants with inward (regular) currents but gave currents in the reverse direction when challenged with eucalyptol and other inhibitors. To determine whether hitherto unknown ORs show this intrareceptor inhibition, we have now examined two other receptors in the same cluster, CquiOR27 and CquiOR28. We cloned and tested four variants of CquiOR28, but none of the 250 compounds in our panel of odorants, including an Orco ligand candidate (OCL12), elicited inward or outward currents. By contrast, CquiOR27/CquiOrco-expressing oocytes gave robust, dose-dependent inward currents when challenged with {gamma}-octalactone and other odorants. On the other hand, octylamine and other phenolic compounds elicited dose-dependent currents in the reverse direction. When stimulatory and inhibitory compounds were presented in binary mixtures, {gamma}-octalactone-elicited inward currents were attenuated in a dose-dependent manner according to the concentration of octylamine. As part of our chemical ecology approach, we tested the repellency activity of the best ligands in the surface landing and feeding assay and a newly reported hand-in cage assay. Protection elicited by {gamma}-octalactone did not differ significantly from that of DEET at the same dose. In the hand-in cage assay, a cream formulation of {gamma}-octalactone showed 97.0 {+/-} 1.3% protection, with 47.6 {+/-} 8.3% and 1.4 {+/-} 0.7% landings per trial in the hands covered with a control and {gamma}-octalactone cream, respectively (N=8, p=0.0078, Wilcoxon matched-pairs signed-rank test).

neuroscience↗

bioRxiv2022

Functional oscillation of a multienzyme glucosome assembly during cell cycle progression

Glucose metabolism has been studied extensively to understand functional interplays between metabolism and a cell cycle. However, our understanding of cell cycle-dependent metabolic adaptation particularly in human cells remains largely elusive. Meanwhile, human enzymes in glucose metabolism are shown to functionally organize into three different sizes of a multienzyme metabolic assembly, the glucosome, to regulate glucose flux in a size-dependent manner. Here, using fluorescence single-cell imaging techniques, we discover that glucosomes spatiotemporally oscillate during a cell cycle in an assembly size-dependent manner. Importantly, their oscillation at single-cell levels is in accordance with functional contributions of glucose metabolism to cell cycle progression at a population level. Collectively, we demonstrate functional oscillation of glucosomes during cell cycle progression and thus their biological significance to human cell biology.

cell biology↗

bioRxiv2021

Serine/arginine-rich splicing factor 7 plays oncogenic roles through specific regulation of m6A RNA modification

Serine/Arginine-Rich Splicing Factor 7 (SRSF7), which is previously recognized as a splicing factor, has been revealed to play oncogenic roles in multiple cancers. However, the mechanisms underlying its oncogenic roles have not been well addressed. Here, based on N6-methyladenosine (m6A) co-methylation network analysis across diverse cell lines, we found SRSF7 positively correlated with glioblastoma cell-specific m6A methylation. We then proved SRSF7 is a novel m6A regulator that specifically facilitates the m6A methylation near its binding sites on the mRNAs involved in cell proliferation and migration through recruiting methyltransferase complex. Moreover, SRSF7 promotes the proliferation and migration of glioblastoma cells largely dependent on the m6A methyltransferase. The two single-nucleotide m6A sites on PBK are regulated by SRSF7 and partially mediate the effects of SRSF7 on glioblastoma cells through recognition by IGF2BP2. Together, our discovery revealed a novel role of SRSF7 in regulating m6A and timely confirmed the existence and functional importance of RNA binding protein (RBP) mediated specific regulation of m6A.

molecular biology↗

bioRxiv2020

Aird: A computation-oriented mass spectrometry data format enables higher compression ratio and less decoding time

With the precision of mass spectrometer going higher and the emergence of data independence acquisition (DIA), the file size is increasing rapidly. Beyond the widely-used open format mzML (Deutsch 2008), near-lossless or lossless compression algorithms and formats have emerged. The data precision is often related to the instrument and subsequent processing algorithms. Unlike storage-oriented formats, which focusing more on lossless compression and compression rate, computation-oriented formats focus as much on decoding speed and disk read strategy as compression rate. Here we describe "Aird", an opensource and computation-oriented format with controllable precision, flexible indexing strategies and high compression rate. Aird uses JavaScript Object Notation (JSON) for metadata storage, multiple indexing, and reordered storage strategies for higher speed of data randomly reading. Aird also provides a novel compressor called Zlib-Diff-PforDelta (ZDPD) for m/z data compression. Compared with Zlib only, m/z data size is about 65% lower in Aird, and merely takes 33% decoding time. AvailabilityAird SDK is written in Java, which allow scholars to access mass spectrometry data efficiently. It is available at https://github.com/Propro-Studio/Aird-SDK AirdPro can convert vendor files into Aird files, which is available at https://github.com/Propro-Studio/AirdPro

bioinformatics↗

bioRxiv2020

Up-Regulation of αCaMKII Impairs Cued Fear Extinction and NMDAR-Dependent LTD in the Lateral Amygdala

Impaired fear extinction is one of the hallmark symptoms of post-traumatic stress disorder (PTSD). The roles of CaMKII have been not extensively studied in fear extinction and LTD. Here, we found PTSD susceptible mice exhibited significant up-regulation of CaMKII in the lateral amygdala (LA). Consistently, increasing CaMKII in LA profoundly not only caused PTSD-like symptoms such as impaired fear extinction and anxiety-like behaviors, but also attenuated NMDAR-dependent LTD at thalamo-LA synapses, reduced GluA1-Ser845/Ser831 dephosphorylation and AMPAR internalization. Suppressing the elevated CaMKII to normal level could completely reverse both PTSD-like symptoms and the impairments in LTD, GluA1-Ser845/Ser831 dephosphorylation and AMPAR internalization. Intriguingly, deficits in AMPAR internalization and GluA1-Ser845/Ser831 dephosphorylation were detected not only after impaired fear extinction, but also after attenuated LTD Our results demonstrate for the first time GluA1-Ser845/Ser831 dephosphorylation and AMPAR internalization are molecular links between LTD and fear extinction, and suggest CaMKII may be a potential molecular determinant of PTSD.

neuroscience↗

bioRxiv2019

Cryo-EM structure of the human Mixed Lineage Leukemia-1 complex bound to the nucleosome

Mixed Lineage Leukemia (MLL) family histone methyltransferases are the key enzymes that deposit histone H3 Lys4 (K4) mono-/di-/tri-methylation and regulate gene expression in mammals. Despite extensive structural and biochemical studies, the molecular mechanism by which the MLL complexes recognize histone H3K4 within the nucleosome core particle (NCP) remains unclear. Here, we report the single-particle cryo-electron microscopy (cryo-EM) structure of the human MLL1 core complex bound to the NCP. The MLL1 core complex anchors on the NCP through RbBP5 and ASH2L, which interacts extensively with nucleosomal DNA as well as the surface close to histone H4 N-terminal tail. Concurrent interactions of RbBP5 and ASH2L with the NCP uniquely align the catalytic MLL1SET domain at the nucleosome dyad, allowing symmetrical access to both H3K4 substrates within the NCP. Our study sheds light on how the MLL1 complex engages chromatin and how chromatin binding promotes MLL1 tri-methylation activity.

biochemistry↗

bioRxiv2019

A non-enzymatic function associated with a putative histone demethylase licenses epigenetic inheritance

H3K9 methylation (H3K9me) specifies the establishment and maintenance of transcriptionally silent epigenetic states or heterochromatin. The enzymatic erasure of histone modifications is widely assumed to be the primary mechanism that resets epigenetic states during and after DNA replication. Here, we demonstrate that a putative histone-demethylase Epe1 in fission yeast, regulates epigenetic inheritance through a non-enzymatic process. Mutations that map to its putative catalytic domain disrupt its interaction with Swi6HP1 and heterochromatin specific pattern of localization without any requirement for enzymatic activity. Epe1 and Swi6HP1 form an inhibitory complex which displaces histone deacetylases from sites of heterochromatin formation. Sequence-specific recruitment of a histone deacetylase, Clr3 renders heterochromatin refractory to the anti-silencing functions of Epe1 and licenses the inheritance of epigenetic states in cis. Epigenetic inheritance solely depends on the read-write activity of the H3K9 methyltransferase Clr4 which competes with genome-wide nucleosome turnover processes in the absence of enzymatic erasure.

biochemistry↗

bioRxiv2019

YAP/TAZ as a Novel Regulator of cell volume

How mammalian cells regulate their physical size is currently poorly understood, in part due to the difficulty of accurately quantifying cell volume in a high throughput manner. Here, using the fluorescence exclusion method, we demonstrate that the mechanosensitive transcriptional regulators YAP (Yes-associated protein) and TAZ (transcriptional coactivator with PDZ-binding motif) are novel regulators of single cell volume. We report that the role of YAP/TAZ in cell volume regulation must go beyond its influence on total cell cycle duration or the cell shape to explain the observed changes in volume. Moreover, for our experimental conditions, volume regulation by YAP/TAZ is independent of mTOR. Instead, we find YAP/TAZ directly impacts the cell division volume. Based on the idea that YAP/TAZ is a mechanosensor, we find that inhibiting the assembly of myosin and cell tension slows cell cycle progression from G1 to S. These results suggest that YAP/TAZ and the Hippo pathway may be modulating cell volume in combination with cytoskeletal tension during cell cycle progression.

bioengineering↗

bioRxiv2018

Microbial Cells Harboring a Mitochondrial Gene Are Capable of CO2 Capture

Global warming is escalating with increased temperatures reported worldwide. Given the enormous land mass on the planet, biological capture of CO2 remains a viable approach to mitigate the crisis as it is economical and easy to implement. In this study, a gene capable of CO2 capture was identified via selection in minimal media. This mitochondrial gene named as OG1 encodes the OK/SW-CL.16 protein and shares homology with cytochrome oxidase subunit III of various species and PII uridylyl-transferase from Loktanella vestfoldensis SKA53. CO2 capture experiments indicate that {delta}13C was substantially higher in the cells harboring the gene OG1 than the control in the nutrition-poor media. This study suggests that CO2 capture using engineered microorganisms in barren land can be exploited to address the soaring CO2 level in the atmosphere, opening up vast land resources to cope with global warming.\n\nIMPORTANCEGlobal warming crisis is deteriorating with increased CO2 levels in the atmosphere each year. Action must be taken before catastrophic consequences occur in the not-so-distant future. Biological capture of CO2 is a feasible approach to alleviate the current crisis. We have identified a mitochondrial gene which demonstrated CO2 utilization capability. Data presented in this study suggest that CO2 capture using engineered microorganisms can be harnessed to address the ever-rising CO2 level in the atmosphere.

bioengineering↗

bioRxiv2018

MMOD-induced structural changes of hydroxylase in soluble methane monooxygenase

Soluble methane monooxygenase in methanotrophs converts methane to methanol under ambient conditions1-3. The maximum catalytic activity of hydroxylase (MMOH) is achieved via interplay of its regulatory protein (MMOB) and reductase4-6. An additional auxiliary protein, MMOD, is believed to function as an inhibitor of the catalytic activity of MMOH; however, the mechanism of its action remains unknown7,8. Herein, we report the crystal structure of MMOH-MMOD complex from Methylosinus sporium strain 5 (2.6 [A]), which illustrates that two molecules of MMOD associate symmetrically with the canyon region of MMOH in a manner similar to MMOB, indicating that MMOD competes with MMOB for MMOH recognition. Further, MMOD binding disrupts the geometry of the di-iron centre and opens the substrate access channel. Notably, the electron density of 1,6-hexanediol at the substrate access channel mimics products of sMMO in hydrocarbon oxidation. The crystal structure of MMOH-MMOD unravels the inhibitory mechanism by which MMOD suppresses the MMOH catalytic activity, and reveals how hydrocarbon substrates/products access to the di-iron centre.

biochemistry↗

bioRxiv2018

DensityPath: a level-set algorithm to visualize and reconstruct cell developmental trajectories for large-scale single-cell RNAseq data

Cell fates are determined by transition-states which occur during complex biological pro-cesses such as proliferation and differentiation. The advance in single-cell RNA sequencing (scRNAseq) provides the snapshots of single cell transcriptomes, thus offering an essential opportunity to study such complex biological processes. Here, we introduce a novel algorithm, DensityPath, which visualizes and reconstructs the underlying cell developmental trajectories for large-scale scRNAseq data. DensityPath has three merits. Firstly, by adopting the nonlinear dimension reduction algorithm elastic embedding, DensityPath reveals the intrinsic structures of the data. Secondly, by applying the powerful level set clustering method, DensityPath extracts the separate high density clusters of representative cell states (RCSs) from the single cell multimodal density landscape of gene expression space, enabling it to handle the heterogeneous scRNAseq data elegantly and accurately. Thirdly, DensityPath constructs cell state-transition path by finding the geodesic minimum spanning tree of the RCSs on the surface of the density landscape, making it more computationally efficient and accurate for large-scale dataset. The cell state-transition path constructed by DensityPath has the physical interpretation as the minimum-transition-energy (least-cost) path. We demonstrate that DensityPath is capable of identifying complex cell development trajectories with bifurcating and trifurcating branches on the human preimplantation embryos. We demonstrate that DensityPath is robust and has high accuracy of pseudotime calculation and branch assignment on the real scRNAseq as well as simulated datasets.

systems biology↗

bioRxiv2017

Structural insights into the architecture of human Importin4_histone H3/H4_Asf1a complex and its histone H3 tail binding

Importin4 transports histone H3/H4 in complex with Asf1a to the nucleus for chromatin assembly. Importin4 recognizes the nuclear localization sequence located at the N-terminal tail of histones. Here, we analyzed the structures and interactions of human Importin4, histones and Asf1a by cross-linking mass spectrometry, X-ray crystallography, negative-stain electron microscopy, small-angle X-ray scattering and integrative modeling. The XL-MS data showed that the C-terminal region of Importin4 interacts extensively with the histone H3 tail. We determined the crystal structure of the C-terminal region of Importin4 bound to the histone H3 peptide, thus revealing that the acidic path in Importin4 accommodates the histone H3 tail and that histone H3 Lys14 is the primary residue interacting with Importin4. Furthermore, the molecular architecture of the Importin4_histone H3/H4_Asf1a complex was produced through an integrative modeling approach. Overall, this work provides structural insights into how Importin4 recognizes histones and their chaperone complex.

biochemistry↗