Biology subjects

Kim, K. P.

Publications and source records attributed to Kim, K. P..

9 recordsLinked to original sources

bioRxiv2024

Fibrotic Extracellular Matrix Preferentially Induces a Partial Epithelial-Mesenchymal Transition Phenotype in a 3-D Agent Based Model of Fibrosis

One of the main drivers of fibrotic diseases is epithelial-mesenchymal transition (EMT): a transdifferentiation process in which cells undergo a phenotypic change from an epithelial state to a pro-migratory state. The cytokine transforming growth factor-{beta}1 (TGF-{beta}1) has been previously shown to regulate EMT. TGF-{beta}1 binds to fibronectin (FN) fibrils, which are the primary extracellular matrix (ECM) component in renal fibrosis. We have previously demonstrated experimentally that inhibition of FN fibrillogenesis and/or TGF-{beta}1 tethering to FN inhibits EMT. However, these studies have only been conducted on 2-D cell monolayers, and the role of TGF-{beta}1-FN tethering in 3-D cellular environments is not clear. As such, we sought to develop a 3-D computational model of epithelial spheroids that captures both EMT signaling dynamics and TGF-{beta}1-FN tethering dynamics. We have incorporated the bi-stable EMT switch model developed by Tian et al. (2013) into a 3D multicellular model, capturing both temporal and spatial TGF-{beta}1 signaling dynamics. We show that the addition of TGF-{beta}1 affects cell proliferation, EMT progression, and cell migration. We then incorporate TGF-{beta}1-FN fibril tethering by locally reducing the TGF-{beta}1 diffusion coefficient as a function of EMT to simulate the reduced movement of TGF-{beta}1 when tethered to FN fibrils during fibrosis. We show that incorporation of TGF-{beta}1 tethering to FN fibrils promotes a partial EMT state, independent of exogenous TGF-{beta}1 concentration, indicating a mechanism by which fibrotic ECM can promote a partial EMT state. Author summaryEpithelial-mesenchymal transition (EMT) is a key cellular process where epithelial cells transform and become mesenchymal. The EMT states are not binary, but instead exhibit a spectrum of partial states where epithelial cells express a combination of epithelial and mesenchymal markers, along with several markers distinct to the partial state. In diseases such as fibrosis and cancer, growing evidence supports the finding that diseased epithelial cells exist primarily in a partial EMT state. However, the mechanisms and signaling factors that drive this partial EMT state in fibrotic diseases is unclear. We use an agent-based model that looks at EMT progression in a population of cells embedded in an ECM environment with controllable fibrotic properties to provide a more systematic approach at studying spatial and temporal changes in the microenvironment that could drive EMT progression and maintain specific EMT phenotypes.

cell biology↗

bioRxiv2024

Meiotic DNA break resection and recombination rely on chromatin remodeler Fun30

DNA double-strand breaks (DSBs) are nucleolytically processed to generate single-stranded DNA tails for homologous recombination. In Saccharomyces cerevisiae meiosis, this 5-to-3 resection involves initial nicking by the Mre11-Rad50-Xrs2 complex (MRX) plus Sae2, then exonucleolytic digestion by Exo1. Chromatin remodeling adjacent to meiotic DSBs is thought to be necessary for resection, but the relevant remodeling activity was unknown. Here we show that the SWI/SNF-like ATPase Fun30 plays a major, non-redundant role in resecting meiotic DSBs. A fun30 null mutation shortened resection tract lengths almost as severely as an exo1-nd (nuclease-dead) mutation, and resection was further shortened in the fun30 exo1-nd double mutant. Fun30 associates with chromatin in response to meiotic DSBs, and the constitutive positioning of nucleosomes governs resection endpoint locations in the absence of Fun30. We infer that Fun30 directly promotes both the MRX- and Exo1-dependent steps in resection, possibly by removing nucleosomes from broken chromatids. Moreover, we found that the extremely short resection in the fun30 exo1-nd double mutant is accompanied by compromised interhomolog recombination bias, leading to defects in recombination and chromosome segregation. Thus, this study also provides insight about the minimal resection lengths needed for robust recombination.

molecular biology↗

bioRxiv2024

Assessing Long-Term Stored Tissues for Multi-Omics Data Quality and Proteogenomics Suitability

As research into the complexities of cancer biology deepens, the integration of multi-omics analyses has emerged as a powerful approach to unravel the complex molecular basis of cancers. However, challenges related to sample availability, including size, collection procedures, and storage duration, hinder the broad application of this methodology. Despite these limitations, there is a growing interest in exploring the potential of archived samples to expand the scope of multi-omics research. Our study aims to investigate the impact of storage duration on the measurment in genomic, transcriptomic, and proteomic profiles of archived samples, demonstrating their viability for advancing our understanding of cancer biology. To comprehensively address these trends and limitations, we systematically examined archived samples collected over a decade, focusing on their transcriptomic, proteomic, and phosphoproteomic attributes. Analysis revealed intricate patterns and dynamic shifts, especially in long-term transcriptomic data, with observed declines in read counts related to protein coding and gene coverage. However, these changes did not compromise the fundamental gene expression landscape. Proteomic result also demonstrated that storage period did not significantly influence proteomic measurement. Comparisons of housekeeping gene (HKG) and housekeeping protein (HKP) expressions unveiled consistent transcriptomic levels across samples, while distinctive proteomic disparities between tumor and normal tissues. In conclusion, the challenges posed by limited sample availability in multi-omics studies can be partially overcome through the strategic integration of archived samples. While technical shifts were evident in certain aspects of transcriptomic data, core gene expression patterns remained robust, and the functionality of essential transcription factors (TFs) and kinases remained unaffected. These findings underscore the potential of archived samples as valuable resources for multi-omics research, providing a broader landscape for investigating cancer biology and paving the way for more comprehensive insights into this intricate field.

cancer biology↗

bioRxiv2023

Unbiased phosphoproteomics analysis unveils modulation of insulin signaling by extramitotic CDK1 kinase activity in human myotubes

Sensitivity and plasticity of insulin signaling and glucose uptake in skeletal muscle depends on determinants such as genetic variation and obesity. We collected muscle biopsies and isolated myoblasts from a multi-ethnic cohort of lean South Asians (N=10), lean Chinese (N=10), and obese Chinese (N=10), and analysed the proteome and phosphoproteome dynamics in terminally differentiated myotubes after a low-dose insulin stimulation (10nM at 0, 5, 30 min). The myotubes initially responded with increased abundance and phosphorylation level changes along the PI3K/AKT/mTOR axis, decreased abundance of translation initiation factors, and increased phosphorylation levels on proteins involved in mRNA processing at 5 min. After the acute response, protein abundance returned to baseline at 30 min, while phosphorylation changes persisted in proteins including AKT, RPS6 and AS160 (TBC1D4). A joint kinase-substrate statistical analysis revealed that protein abundance changes of AKT, PAK1 and CDK1 showed concordant phosphorylation changes in their respective substrates upon insulin stimulation. We also observed increased phosphorylation of some substrates uniquely in each group, particularly the substrates of CDKs showing stronger changes in South Asians than in Chinese. Pharmacological inhibition and siRNA knockdown of CDK1, a non-myogenic kinase, in terminally differentiated myotubes reduced glucose uptake and desensitized several phosphorylation-mediated signaling on protein translation initiation factors, IRS1, and AS160. Our data suggest that basal extramitotic activity of CDK1 is required for PI3K/AKT/mTORC1 signaling cascade and glucose uptake in insulin-stimulated myotubes. The data also provide a rich resource for studying the role of other kinases in the mechanism of insulin resistance in human myotubes.

physiology↗

bioRxiv2022

Vutiglabridin improves neurodegeneration in MPTP-induced Parkinson's disease mice by targeting mitochondrial paraoxonase-2

Parkinsons disease (PD), characterized by degeneration of dopaminergic neurons, share pathogenic features with obesity, including mitochondrial dysfunction and oxidative stress. Paraoxonase 2 (PON2) is an inner mitochondrial membrane protein that is highly expressed in dopaminergic neurons and is involved in the regulation of mitochondrial oxidative stress. However, no drug targeting PON2 has ever been developed for the treatment of PD. Here, we show that vutiglabridin, a clinical phase 2-stage drug for the treatment of obesity, has therapeutic effects in PD models, targeting mitochondrial PON2. Vutiglabridin penetrates into the brain, binds to PON2, and restores 1-methyl-4-phenylpyridinium (MPP+)-induced mitochondrial dysfunction in SH-SY5Y neuroblastoma cells. Knockdown of PON2 by lentiviral shRNA infection abolished the effects of vutiglabridin on mitochondria. In mice, vutiglabridin significantly alleviated motor impairments and damage to dopaminergic neurons in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD model, and these effects were also abolished in PON2-knockdown mice, suggesting that vutiglabridin is neuroprotective via PON2. Extensive in vitro and in vivo assessment of potential neurotoxicity showed vutiglabridin to be safe. Overall, these findings provide support for the clinical development of vutiglabridin as a novel PON2 modulator for the treatment of PD. One Sentence SummaryTargeting paraoxonase-2 by a clinical-stage compound vutiglabridin provides neuroprotective effects in preclinical models of Parkinsons disease.

neuroscience↗

bioRxiv2022

ECPAS/Ecm29-Mediated 26S Proteasome Disassembly Is an Adaptive Response to Glucose Starvation

The 26S proteasome consists of loosely associated 20S catalytic and 19S regulatory complexes. Approximately half of the proteasomes in eukaryotic cells exist as free 20S complexes; however, our mechanistic and physiological understanding of what determines the ratio of 26S to 20S species remains incomplete. Here, we show that glucose starvation in mammalian cells results in the uncoupling of 26S holoenzymes into intact 20S and 19S subcomplexes. Subcomplex affinity-purification and quantitative mass spectrometry revealed that Ecm29 proteasome adaptor and scaffold (ECPAS) is a crucial mediator of this structural remodeling. The loss of ECPAS abrogated 26S dissociation, leading to decreased degradation of 20S proteasome substrates such as puromycylated polypeptides and lysine-less cyclin B. In silico modeling analysis suggested that the conformational changes of ECPAS may commence the disassembly process. ECPAS was also essential for proper endoplasmic reticulum stress response and cell survival during glucose starvation. In addition, we evaluated the role of ECPAS in vivo using the mouse xenograft model and observed that glucose-deprived tumor tissues had significantly elevated 20S proteasome levels. Collectively, our results indicate that the 20S-19S disassembly mediated by ECPAS is a novel mechanism adapting global proteolysis to physiological needs and an effective cellular strategy against proteotoxic stress.

biochemistry↗

bioRxiv2022

P176S polymorphism in Maspin rewires electrostatic interaction that alters Maspin functionality

Maspin has been known to regress tumors by inhibiting angiogenesis, however, its roles have been reported to be context and sequence-dependent. There are various proteins and cofactors that bind with maspin possibly explaining the conflicting roles of maspin. Moreover, maspin polymorphic forms have also been linked to tumor regression or survival, for instance, maspin with Ser at 176 (maspin-S176) promotes tumor while maspin with Pro at 176 (maspin-Pro176) has opposing roles in cancer pathogenesis. With the help of long molecular dynamic simulation, a possible link between polymorphic forms and tumor progression has been established. First, the maspin is dynamically stable with either amino acid at 176 position, secondly, differential contacts have been observed among various regions, thirdly, these contacts have significantly altered the electrostatic energetics of various residues, and finally, these altered electrostatics of maspin-S176 and maspin-P176 rewired the polar contacts that abolished the allosteric control in protein. By combining these factors, the altered electrostatics can substantially affect the localization, and the preference of maspin binding partners, thus, culminating in a different maspin-protein (cofactor)-interaction landscape that could have been manifested with conflicting reports in previous studies. Here, the underlying reason has been highlighted and discussed that could be helpful for better therapeutic manipulations. SignificanceProtein altered functions in response to mutations are well documented, however, a slight perturbation in structure can lead to dramatic effects that are being felt at longer distances are rare. Here, we have reported that the substitution of Pro to Ser at 176th position in Maspin can substantially alter the protein electrostatic interactions that can hamper the allosteric control. This could lead to different binding partners, localization preferences, and altered cytoplasmic retention duration resulting in functions that are not associated with normal protein. Moreover, the electrostatic attraction/repulsion can immensely affect the allosteric cohesion of protein resulting in unexpected outcomes.

biophysics↗

bioRxiv2021

Synthetic glabridin derivatives mitigate steatohepatitis in a diet-induced biopsy-confirmed non-alcoholic steatosis hepatitis mouse model through paraoxonase-2

Limited therapeutic agents have been developed for non-alcoholic steatohepatitis (NASH), a common immunometabolic disease that can progress to hepatic cirrhosis and cancer. Glabridin and its derivatives are potential therapeutics for some metabolic diseases. However, the therapeutic effects of glabridin and its derivatives on NASH and their biological functions are unclear. This study demonstrated the role of synthetic glabridin derivatives (SGDs) in alleviating hepatic steatosis and inflammation in a biopsy-confirmed rodent NASH model. SGDs exerted therapeutic effects by activating autophagy and the antioxidant defense system, which mitigate NASH pathogenesis. The cellular target of HSG4112, an SGD, was paraoxonase 2. These findings will enable the development of novel therapeutics for NASH in the future.

pharmacology and toxicology↗

bioRxiv2021

Meiosis-Specific Cohesin Complexes Display Distinct and Essential Roles in Mitotic ESC Chromosomes

Cohesin is a chromosome-associated SMC-kleisin complex that mediates sister chromatid cohesion, recombination, and most chromosomal processes during mitosis and meiosis. Through high-resolution 3D-structured illumination microscopy and functional analyses, we report multiple biological processes associated with the meiosis-specific cohesin components, REC8 and STAG3, and the distinct loss of function of meiotic cohesin during the cell cycle of embryonic stem cells (ESCs). First, we show that REC8 is translocated into the nucleus in a STAG3-dependent manner. REC8/STAG3-containing cohesin regulates chromosome topological properties and specifically maintains centromeric cohesion. Second, REC8 and mitotic cohesin RAD21 are located at adjacent sites but predominantly at nonoverlapping sites on ESC chromosomes, implying that REC8 can function independent of RAD21 in ESCs. Third, knockdown of REC8-cohesin not only leads to higher rates of premature centromere separation and stalled replication forks, which can cause proliferation and developmental defects, but also enhances compaction of the chromosome structure by hyperloading of retinoblastoma protein-condensin complexes from prophase onward. We propose that the delicate balance between mitotic and meiotic cohesins may regulate ESC- specific chromosomal organization and mitotic program.

molecular biology↗