Biology subjects

Lin, S.-H.

Publications and source records attributed to Lin, S.-H..

18 recordsLinked to original sources

bioRxiv2026

PHENOMS: Open-Source Software for Rust-Accelerated Hydrogen-Bond Analysis in Molecular Dynamics Simulations

Hydrogen-deuterium exchange mass spectrometry (HDX-MS) provides powerful insight into protein structure and dynamics, but interpretation of protection patterns at atomic resolution often requires complementary computational modeling. We present PHENOMS, an open-source Python package that integrates molecular dynamics trajectories with HDX-MS data to analyze backbone hydrogen-bond networks by tracking how individual hydrogen bonds and residue-residue contacts form, break, and reorganize throughout the simulations. This residue-level analysis can reveal local structural changes that may be missed by conventional global trajectory metrics. PHENOMS employs a multithreaded, Rust-accelerated computational engine that achieved an average 16-fold speedup (up to 100-fold) over widely used Python-based MD analysis packages in our benchmarks, while retaining individual hydrogen-bond resolution in every trajectory frame. We demonstrate PHENOMS through multiple retrospective case studies using published and in-house datasets, showing that it can recapitulate HDX-MS protection patterns, identify key residues that drive coordinated structural changes, and benchmark the computational efficiency of the approach. With automated quality-control workflows, PHENOMS provides a framework for integrating hydrogen-bond analysis with HDX-MS data or serves as a standalone, high-throughput tool for analyzing MD trajectories for broader applications in protein engineering, drug discovery, and structural biology.

biophysics↗

bioRxiv2026

Mechanism of molecular recognition revealed through dynamic drug binding pathways to SARS-CoV-2 main protease

Characterization of drug-binding pathways remains experimentally limited by transient intermediates and computationally challenging due to long timescales intractable for conventional molecular dynamics. To address these challenges, we combined solution NMR titrations with weighted ensemble (WE) enhanced sampling simulations to resolve atomistic pathways of nirmatrelvir binding to the SARS-CoV-2 main protease. NMR titration revealed residue-dependent heterogeneity spanning fast, intermediate, and slow exchange regimes. WE simulations complement the NMR by providing insights into unassigned residues and adding time-resolved and three-dimensional structural context. We map key interactions along two distinct binding pathways, provide dynamic explanations for residues involved in resistance, and capture unique backbone conformations compared to those sampled in unbound or bound states. Our comprehensive binding model is consistent with a combined conformational selection and induced fit mechanism in which early transient contacts are made with residues E47 and L50 and allosteric motions are centered around residue V204 of the distal domain. This synergistic application of WE and titration NMR enables a more comprehensive characterization of drug binding than either method alone, providing an integrated framework that may have broader applicability to defining structure-kinetic relationships and guiding design of next-generation inhibitors.

biophysics↗

bioRxiv2026

Assessing the Generalizability of Machine Learning and Physics Methods for DNA-Encoded Libraries

Predicting protein-ligand binding is a central challenge in computational drug discovery, and while machine learning (ML) and co-folding methods have advanced rapidly, their ability to generalize beyond training or parameterization regimes remains insufficiently understood. DNA-encoded libraries (DELs) enable ultra-large screening of billions of molecules simultaneously, providing a useful testbed for evaluating these approaches at scale. A recent NeurIPS competition revealed that even top performing ML models trained on DEL data failed at generalizing to out-of-distribution (OOD) chemical space. We investigated whether integrating structural modeling could bridge this generalization gap. We systematically assessed state-of-the-art ML, docking, and co-folding methods including Schrodinger Glide, Rosetta GALigandDock, and Boltz-2 with three biologically diverse protein targets screened against libraries containing multiple DEL synthesis formats. While ML excels in-distribution, OOD hit discrimination is dependent on both the target and ligand context, with no single method consistently dominating. These findings demonstrate that benchmark performance alone is insufficient to predict OOD performance, highlighting the need for system-dependent evaluation of binding prediction methods. We provide an open-source package for assessing protein-ligand prediction methods and analyzing high-throughput screening data: DEL-iver. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=118 SRC="FIGDIR/small/719394v2_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@cd8a17org.highwire.dtl.DTLVardef@252d0aorg.highwire.dtl.DTLVardef@b020f0org.highwire.dtl.DTLVardef@1429e27_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2025

Astrocytic FKBP5 Regulates Neuroinflammation and Cognitive Outcomes in Excitotoxic Brain Injury

FK506-binding protein 51 (FKBP51, encoded by FKBP5) is a multisignaling cochaperone that regulates cellular responses to stress. FKBP51 is upregulated in reactive astrocytes; however, the role of FKBP51 in excitotoxic brain injury remains unknown. Here, we investigated how both global and astrocyte-specific Fkbp5 deletion influence seizure susceptibility, astrogliosis, neuroinflammation, and cognition in male mice subjected to a kainic acid (KA)-induced epilepsy mouse model. Global Fkbp5 knockout (Fkbp5-KO) presented lower seizure activity along with decreased neuronal loss and astrogliosis in the hippocampus compared with the wild-type mice. Astrocyte-specific Fkbp5 conditional knockout (aFkbp5-cKO) mice similarly attenuated seizure severity, decreased astrogliosis, improved novel object recognition, and preserved glutamate transporter 1 (GLT-1) expression in hippocampal CA3. Glia-neuron mixed cultures derived from Fkbp5-KO brains showed reduction of NMDA-induced neurotoxicity, astrogliosis, accompanied by decreased NF-{kappa}B p65 phosphorylation. Notably, overexpression of an Fkbp5 quadruple mutant that disrupts the FKBP51-NF-{kappa}B interaction inhibited proinflammatory lipopolysaccharide (LPS)-induced astrogliosis and NF-{kappa}B activation. The hippocampal transcriptome of the LPS-treated Fkbp5-KO mice revealed suppression of NF-{kappa}B signaling. In summary, this study highlights FKBP51 as a key mediator of excitotoxin-induced neuroinflammation and GLT-1 dysfunction and underlines NF-{kappa}B-mediated inflammatory astrogliosis as a potential intervention target for excitotoxic brain injury.

neuroscience↗

bioRxiv2025

Wntless interacts with Notch signaling to balance the generation of neurons and gliocytes in vertebrate dorsal diencephalon

Wnt chaperon Wntless (Wls) mediates the intracellular transport of Wnts and plays important roles in early vertebrate brain development. Spatially restricted induction of Wls denotes the earliest differentiation of non-telencephalic cells in human brain organoids. In zebrafish developing diencephalon, loss-of-Wls reduces the formation of habenula (HA) neurons but how Wls influences HA neurogenesis is unclear and whether Wls regulates gliocyte development is unknown. Here we report that the formations of cholinergic, substance P-ergic or glutamatergic neurons in HA are reduced differentially but the generation of gliocyte-derived choroid plexus (ChP) epithelia is increased in wls null mutants. At earlier stage, three-dimensional gene expression analyses revealed that while neurog1 expressions in HA progenitor zones are reduced, the expressions of Notch downstream effector her6 are increased and expanded into HA progenitor zones in wls mutants. Over-expressing Her6 in neurog1-positive cells reduced neurog1 expressions in HA progenitor zones. These results indicate that Wls restricts the expressions of Notch effector her6 to promote the specification of neurog1 proneurons and demotes the generation of ChP epithelia in zebrafish embryonic dorsal diencephalon.

developmental biology↗

bioRxiv2025

Activating RET Mutations Promotes Osteoblastic Bone Metastases in Medullary Thyroid Cancer

Development of bone metastases increases mortality in patients with medullary thyroid cancer (MTC), with [~]50% survival at 5 years after diagnosis, but the underlying mechanisms are unknown. We show that patient-derived MTC cells (RETC634W mutant TT cells and RETM918T mutant MZCRC1 cells) promote an osteoblastic phenotype due to reduced bone resorption. Mechanistically, activated RET increases the expression of osteoprotegerin (OPG), an inhibitor of bone resorption, leading to decreased osteoclast differentiation. Furthermore, RET knockdown or pharmacological RET inhibition attenuates tumor burden and osteoblastic lesions in MTC-bearing femurs. Circulating levels of OPG were increased in the plasma of MTC patients who developed bone metastases and this was associated with poor overall survival. Patients who were treated with multi-kinase inhibitors have lower circulating levels of OPG. These novel findings identify a link between the RET signaling pathway and abnormal osteoblastic bone formation and suggest OPG as a potential biomarker of MTC bone metastases. HighlightsO_LIPatient-derived MTC cells promote osteoblastic lesions in a mouse model. C_LIO_LIActivating the RET mutation promotes osteoprotegerin. C_LIO_LIBlocking RET kinase activity inhibits tumor growth and the osteoblastic lesion phenotype. C_LIO_LIHigh levels of circulating osteoprotegerin are associated with poor overall survival. C_LI

cancer biology↗

bioRxiv2025

Unraveling forensic timelines using molecular markers in Phormia regina maggots

In forensic entomology, estimating the time of death is critical and traditionally relies on changes in observable traits of carrion feeding insect larvae. Traits such as size, weight, and morphology can be used to predict the insect specimen age and help estimate the time since death. The blowfly Phormia regina is a key forensic insect, yet age estimation for older maggots in this species is particularly challenging due to the limited morphological changes in the late-stage larvae. To enhance precision, we employed transcriptomic profiling on blowfly maggots, aiming to identify genes as markers for time of death estimation. Our study characterized maggot development, reinforcing that weight and behavior cannot precisely determine age between 100 and 130 hours. We built a chromosomal scale annotated genome, establishing a reliable database for uncovering transcriptomic signatures during larval development. Applying differential gene expression analyses, weighted gene co-expression network analysis, and the generalized linear model, we identified nine candidate genes (Y5078, Y5076, agt2, ech1, dhb4, asm, gabd, acohc, Ivd) that delineate the age of otherwise indeterminate maggots. This research introduces a molecular approach to address a longstanding problem in forensic entomology and promises to increase precision in determining the time of death at a crime scene. AUTHOR SUMMARYIn forensic entomology, estimating the time of death often depends on insect morphological changes. The blowfly Phormia regina is a key species, but aging older maggots in the third instar larval phase is difficult due to minimal changes in observable traits. To improve accuracy, we used transcriptomic profiling to identify genetic markers for time of death estimation. Our study showed that weight and behavior are not reliable indicators of age in maggots 100 to 130 hours old. We developed an annotated genome to identify transcriptomic signatures during larval development. Through various analyses, we identified nine genes (Y5078, Y5076, agt2, ech1, dhb4, asm, gabd, acohc, Ivd) that can determine the age of late third instar maggots. This molecular approach aims to enhance precision in forensic entomology.

developmental biology↗

bioRxiv2024

Leveraging Machine Learning and AlphaFold2 Steering to Discover State-Specific Inhibitors Across the Kinome

Protein kinases are structurally dynamic proteins that control downstream signaling cascades by phosphorylating their substrates. Protein kinases regulate their function by adopting several conformational states in their active site determined by the movements of several motifs such as the C-Helix, DFG residues and the activation loop. Each conformational state represents a distinct physicochemical environment that accepts or precludes ligand binding. However, most of the kinome have not been crystalized across these possible conformational states. It has been shown that shallow Multiple Sequence Alignments (MSA) can enable AlphaFold2 (AF2) to model kinases in alternative conformations. However, it is unclear if these models can be leveraged for structure-based drug discovery. Additionally, there are several machine learning tools to predict protein-ligand interactions based on ligand chemotype and binding pocket properties, but these models cannot be used to identify ligands with clear state specificity. Here, we first present an approach called AlphaFold2 Steering (AF2-Steering), a systematic methodology to direct AF2 to sample kinases in the active and inactive conformations. We use our approach to model the protein kinome in precise conformational states. We demonstrate the utility of these AF2-steered kinase models by employing them in a prospective virtual screening study that integrates machine learning with docking to find state specific inhibitors for well-studied and dark kinases that lack structures in the active conformational state. We then experimentally validate the hits, an essential step often overlooked, and later experimentally confirm the conformation-specificity of the ligands identified for FLT3, a protein kinase that currently lacks an active state crystal structure. Against a strict binding criterion of at least 1M Kd, our modelled structures achieved an overall hit rate of 53%. We also confirm the conformation-specificity of 4/7 FLT3 ligands, thus demonstrating the value of MSA-steered AF2 modelled kinase structures combined with machine learning and docking to guide conformation-specific kinase drug discovery.

bioinformatics↗

bioRxiv2023

Prostate cancer-induced endothelial-to-osteoblast transition generates an immunosuppressive bone tumor microenvironment

Immune checkpoint therapy has limited efficacy for patients with bone metastatic castrate-resistant prostate cancer (bmCRPC). In this study, we revealed a novel mechanism that may account for the relative resistance of bmCRPC to immune checkpoint therapy. We found that prostate cancer (PCa)-induced bone via endothelial-to-osteoblast (EC-to-OSB) transition causes an ingress of M2-like macrophages, leading to an immunosuppressive bone tumor microenvironment (bone-TME). Analysis of a bmCRPC RNA-seq dataset revealed shorter overall survival in patients with an M2-high versus M2-low signature. Immunohistochemical (IHC) analysis showed CD206+ M2-like macrophages were enriched in bmCRPC specimens compared with primary tumors or lymph node metastasis. In osteogenic PCa xenografts, CD206+ macrophages were enriched adjacent to tumor-induced bone. FACS analysis showed an increase in CD206+ cells in osteogenic tumors compared to non-osteogenic tumors. Genetic or pharmacological inhibition of the EC-to-OSB transition reduced aberrant bone and M2-like macrophages in osteogenic tumors. RNAseq analysis of tumor-associated macrophages from osteogenic (bone-TAMs) versus non-osteogenic (ctrl-TAMs) tumors showed high expression of an M2-like gene signature, canonical and non-canonical Wnt pathways, and a decrease in an M1-like gene signature. Isolated bone-TAMs suppressed T-cell proliferation while ctrl-TAMs did not. Mechanistically, EC-OSB hybrid cells produced paracrine factors, including Wnts, CXCL14 and LOX, which induced M2 polarization and recruited M2-like TAMs to bone-TME. Our study thus links the unique EC-to-OSB transition as an "upstream" event that drives "downstream" immunosuppression in the bone-TME. These studies suggest that therapeutic strategies that inhibit PCa-induced EC-to-OSB transition may reverse immunosuppression to promote immunotherapeutic outcomes in bmCRPC. SignificanceThe insight that prostate cancer-induced bone generates an immunosuppressive bone tumor microenvironment offers a strategy to improve responses to immunotherapy approaches in patients with bone metastatic castrate-resistant prostate cancer.

cancer biology↗

bioRxiv2022

Alendronate Conjugate for Targeted Delivery to Bone-Forming Prostate Cancer

Bone is the primary metastasis site for lethal prostate cancer, often resulting in poor prognosis, crippling pain, and diminished functioning that drastically reduce both quality of life and survivability. Uniquely, prostate cancer bone metastasis induces aberrant bone overgrowth, due to an increase of osteoblasts induced by tumor-secreted bone morphogenetic protein 4 (BMP4). Conjugating drugs to substances that target the tumor-induced bone area within the metastatic tumor foci would be a promising strategy for drug delivery. To develop such a strategy, we conjugated a near infrared (NIR) fluorescent probe, the dye Cy5.5, to serve as a surrogate for drugs, with alendronate, which targets bone. Characterization, such as infrared spectroscopy, confirmed the synthesis of the Cy5.5-ALN conjugate. The maximum absorbance of free Cy5.5, which was at 675 nm, did not change upon conjugation. Alendronate targeted the bone component hydroxyapatite in a dose-dependent manner up to 2.5 M, with a maximum of 85% of Cy5.5-ALN bound to hydroxyapatite, while free Cy5.5 alone had 6% binding. In in vitro cell binding studies, Cy5.5-ALN bound specifically with mineralized bone matrix of differentiated MC3T3-E1 cells or 2H11 endothelial cells that were induced to become osteoblasts through endothelial-to-osteoblast transition, the underlying mechanism of prostate-cancer-induced bone formation. Neither Cy5.5-ALN nor free Cy5.5 bound to undifferentiated MC3T3-E1 or 2H11 cells. Bone-targeting efficiency studies in non-tumor-bearing mice revealed accumulation over time in the spine, jaw, knees, and paws injected with Cy5.5-ALN, and quantification showed higher accumulation in femurs than in muscle at up to 28 days, while the free Cy5.5 dye was observed circulating without preferential accumulation and decreased over time. There was a linear relationship with fluorescence when the injected concentration of Cy5.5-ALN was between 0.313 and 1.25 nmol/27 g of mouse, as quantified in mouse femurs both in vivo and ex vivo. Ex vivo evaluation of bone-targeting efficiency in nude mice was 3 times higher for bone-forming C4-2b-BMP4 tumors compared to non-bone-forming C4-2b tumors (p-value < 0.001). Fluorescence microscopy imaging of the tumors showed that Cy5.5-ALN co-localized with the bone matrix surrounding tumor-induced bone, but not with the viable tumor cells. Together, these results suggest that a drug-ALN conjugate is a promising approach for targeted delivery of drug to the tumor-induced bone area in the metastatic foci of prostate cancer.

biochemistry↗

bioRxiv2022

Dexamethasone Enhances CAR T Cell Persistence and Function by Upregulating Interleukin 7 Receptor

Dexamethasone (dex) is a glucocorticoid that is a mainstay for treatment of inflammatory pathologies, including immunotherapy-associated toxicities. Dex suppresses the endogenous immune response and is also believed to suppress the function of chimeric antigen receptor (CAR) T cells. However, recent reports observed higher CAR T cell numbers in patients treated with dex, highlighting the rationale for interrogating the specific effects of dex on CAR T cells. Here, we found that dex did not inhibit CAR T cell expansion or function. A single dose of dex during the manufacturing process upregulated the pro-persistence interleukin 7 receptor (IL7R) on CAR T cells and induced expression of genes involved in activation, migration, and persistence. The ex vivo upregulation of IL7R induced by dex significantly enhanced CAR T cell persistence and anti-tumor efficacy in vivo when combined with exogenous IL-7. Moreover, the combination of dex and IL-7 resulted in increased persistence of CAR T cells and led to complete remission of mice. Overall, our studies in both in vitro and in vivo treatment support a positive role of dex on CAR T cell potency and provide insight into the application of glucocorticoids in cellular anti-cancer therapy.

immunology↗

bioRxiv2022

KDM4A promotes NEPC progression through regulation of MYC expression

Neuroendocrine prostate cancer (NEPC) represents one of the most lethal forms of prostate cancer (PCa) and lacks life-prolonging treatment. The incidence of NEPC is increased due to the widespread use of AR pathway inhibitors (ARPIs) in the treatment of non-metastatic CRPC and hormone-sensitive metastatic tumors. Here, we identified histone lysine demethylase KDM4A as a key player in NEPC progression and an effective therapeutic target. We found that KDM4A mRNA and protein are overexpressed in human and mouse NEPC compared to prostate adenocarcinoma. Knockdown (KD) or knockout (KO) of KDM4A in NEPC cell lines suppressed cancer cell growth in vitro and in vivo. Mechanistically, we found that KDM4A promotes NEPC progression, in part, through direct transcriptional regulation of MYC. We showed that MYC is hyper-activated in human and mouse NEPC. KDM4A KD led to suppression of MYC signaling. MYC KD or inhibition profoundly suppressed NEPC cell proliferation. Furthermore, a potent pan-KDM4 inhibitor QC6352 significantly reduced NEPC cell growth in vitro and in vivo. Taken together, we demonstrated that KDM4A is an important regulator of NEPC progression and targeting KDM4A may potentially be an effective therapeutic strategy for NEPC. SignificanceNeuroendocrine prostate cancer (NEPC) is a highly aggressive prostate cancer subtype that is resistant to potent androgen receptor pathway inhibitors (ARPIs) and currently lacks effective therapeutic options. Histone lysine demethylase KDM4A is an important epigenetic regulator of gene expression in development and cancer. In this study, we show that KDM4A is highly expressed in NEPC and is required for NEPC proliferation, anchorage-independent growth, and in vivo growth, which is in part mediated through the regulation of MYC expression. Importantly, we demonstrate that inhibition of KDM4A significantly impairs NEPC growth in preclinical models. Thus, our findings provide valuable insights into the molecular mechanisms underlying NEPC progression and offer a rationale for clinical trials with KDM4 inhibitor in NEPC patients.

cancer biology↗

bioRxiv2021

Activation of retinoic acid receptor reduces metastatic prostate cancer bone lesions through blocking endothelial-to-osteoblast transition

Metastatic prostate cancer (PCa) in bone induces bone-forming lesions that contribute to progression and therapy resistance. Currently strategies targeting PCa-induced bone formation are lacking. We previously showed that PCa-induced bone originates from endothelial cells (EC) that have undergone endothelial-to-osteoblast (EC-to-OSB) transition in response to tumor-secreted BMP4. Here, we show that activation of retinoic acid receptor (RAR) inhibits EC-to-OSB transition and reduces PCa-induced bone formation. We found that palovarotene, a RAR{gamma} agonist being tested for heterotopic ossification in fibrodysplasia ossificans progressiva, inhibited EC-to-OSB transition and osteoblast mineralization in vitro, and decreased tumor-induced bone formation and tumor growth in several osteogenic PCa models. RAR/{beta}/{gamma} isoform knockdown in 2H11 ECs blocked EC-to-OSB transition and osteoblast mineralization. Pan-RAR agonist ATRA inhibited MycCaP-BMP4-induced bone formation and tumor growth under castration. Furthermore, palovarotene or ATRA reduced plasma Tenascin C, a factor secreted by EC-OSB cells, which may be used to monitor treatment response. Mechanistically, BMP4-activated pSmad1 forms a complex with RAR in the nucleus of 2H11 cells. RAR activation by palovarotene or ATRA causes pSmad1 degradation by recruiting E3-ubiquitin ligase Smurf1 into the nuclear pSmad1/RAR{gamma} complex. Our findings suggest that palovarotene can be repurposed to target PCa-induced bone formation to improve clinical outcomes for bone metastasis.

cancer biology↗

bioRxiv2021

Prostate tumor-induced stromal reprogramming generates Tenascin C that promotes prostate cancer metastasis through YAP/TAZ inhibition

Metastatic prostate cancer (PCa) in bone induces bone-forming lesions that enhance PCa progression. How tumor-induced bone formation enhances PCa progression is not known. We have previously shown that PCa-induced bone originates from endothelial cells (EC) that have undergone endothelial-to-osteoblast (EC-to-OSB) transition by tumor-secreted BMP4. Here, we show that EC-to-OSB transition leads to changes in the tumor microenvironment that increases the metastatic potential of PCa cells. We found that conditioned medium (CM) from EC-OSB hybrid cells increases the migration, invasion and survival of PC3-mm2 and C4-2B4 PCa cells. Quantitative mass spectrometry (iTRAQ) identified Tenascin C (TNC) as one of the major proteins secreted from EC-OSB hybrid cells. TNC expression in tumor-induced osteoblasts was confirmed by immunohistochemistry of MDA-PCa118b xenograft and human bone metastasis specimens. Mechanistically, BMP4 increases TNC expression in EC-OSB cells through the Smad1-Notch/Hey1 pathway. How TNC promotes PCa metastasis was next interrogated by in vitro and in vivo studies. In vitro studies showed that a TNC neutralizing antibody inhibits EC-OSB-CM-mediated PCa cell migration and survival. TNC knockdown decreased, while addition of recombinant TNC or TNC overexpression increased migration and anchorage-independent growth of PC3 or C4-2b cells. When injected orthotopically, PC3-mm2-shTNC clones decreased metastasis to bone, while C4-2b-TNC overexpressing cells increased metastasis to lymph nodes. TNC enhances PCa cell migration through 5{beta}1 integrin-mediated YAP/TAZ inhibition. These studies elucidate that tumor-induced stromal reprogramming generates TNC that enhances PCa metastasis and suggest that TNC may be a target for PCa therapy.

cancer biology↗

bioRxiv2021

The prevalence, predominance, and metabolic potentials of Candidatus Prochlorobium terpiosii in the coral-killing sponge, Terpios hoshinota

Terpios hoshinota is a ferocious, space-competing sponge that kills a variety of stony corals by overgrowth. Outbreaks of this species have led to intense coral reef damage and declines in living corals on the square kilometer scale in many geographical locations. Our large-scale 16S rRNA gene survey across three oceans revealed that the core microbiome of T. hoshinota included operational taxonomic units (OTUs) related to Prochloron, Endozoicomonas, Pseudospirillum, SAR116, Magnetospira, and Ruegeria. A Prochloron- related OTU was the most dominant cyanobacterium in T. hoshinota in the western Pacific Ocean, South China Sea, and Indian Ocean. The complete metagenome-assembled genome of the Prochloron-related cyanobacterium and our pigment analysis revealed that this bacterium had phycobiliproteins and phycobilins and lacked chlorophyll b, inconsistent with the iconic definition of Prochloron. Furthermore, the phylogenetic analyses based on 16S rRNA genes and 120 single-copy genes demonstrated that the bacterium was phylogenetically distinct to Prochloron, strongly suggesting that it should be a sister taxon to Prochloron; we therefore proposed this symbiotic cyanobacterium as a novel species under a new genus: Candidatus Paraprochloron terpiosii. With the recovery of the complete genome, we characterized the metabolic potentials of the novel cyanobacterium in carbon and nitrogen cycling and proposed a model for the interaction between Ca. Pp. terpiosi LD05 and T. hoshinota. In addition, comparative genomics analysis revealed that Ca. Paraprochloron and Prochloron showed distinct features in transporter systems and DNA replication. ImportanceThe finding that one species predominates cyanobacteria in T. hoshinota from different geographic locations indicates that this sponge and Ca. Pp. terpiosi LD05 share a tight relationship. This study builds the foundation for T. hoshinotas microbiome and paves a way for understanding the ecosystem, invasion mechanism, and causes of outbreak of this coral-killing sponge. Also, the first Prochloron-related complete genome enables us to study this bacterium with molecular approaches in the future and broadens our knowledge of the evolution of symbiotic cyanobacteria.

ecology↗

bioRxiv2021

P4HA2-induced prolyl hydroxylation suppresses YAP1-mediated prostate cancer cell migration, invasion, and metastasis

Yes-associated protein 1 (YAP1), a key player in the Hippo pathway, has been shown to play a critical role in tumor progression. However, the role of YAP1 in prostate cancer cell invasion, migration, and metastasis is not well defined. Through functional, transcriptomic, epigenomic, and proteomic analyses, we showed that prolyl hydroxylation of YAP1 plays a critical role in the suppression of cell migration, invasion, and metastasis in prostate cancer. Knockdown (KD) or knockout (KO) of YAP1 led to an increase in cell migration, invasion, and metastasis in prostate cancer cells. Microarray analysis showed that the EMT pathway was activated in Yap1-KD cells. ChIP-seq analysis showed that YAP1 target genes are enriched in pathways regulating cell migration. Mass spectrometry analysis identified P4H prolyl hydroxylase in the YAP1 complex and YAP1 was hydroxylated at multiple proline residues. Proline-to-alanine mutations of YAP1 isoform 3 identified proline 174 as a critical residue, and its hydroxylation suppressed cell migration, invasion, and metastasis. KO of P4ha2 led to an increase in cell migration and invasion, which was reversed upon Yap1 KD. Our study identified a novel regulatory mechanism of YAP1 by which P4HA2-dependent prolyl hydroxylation of YAP1 determines its transcriptional activities and its function in prostate cancer metastasis.

cancer biology↗

bioRxiv2020

Hyperpolyploidization of hepatocyte initiates preneoplastic lesion formation in the liver

Hepatocellular carcinoma (HCC) is the most predominant primary malignancy in the liver. Genotoxic and genetic models have revealed that HCC cells are derived from hepatocytes, but where the critical region for tumor foci emergence is and how this transformation occurs are still unclear. Here, hyperpolyploidization of hepatocytes around the centrilobular (CL) region was demonstrated to be closely linked with the development of HCC cells after diethylnitrosamine treatment. We identified the CL region as a dominant lobule for accumulation of hyperpolyploid hepatocytes and preneoplastic tumor foci formation. We also demonstrated that upregulation of Aurkb plays a critical role in promoting hyperpolyploidization. Increase of AURKB phosphorylation was detected on the midbody during cytokinesis, causing abscission failure and hyperpolyploidization. Pharmacological inhibition of AURKB dramatically reduced nucleus size and tumor foci number surrounding the CL region in diethylnitrosamine-treated liver. Our work reveals an intimate molecular link between pathological hyperpolyploidy of CL hepatocytes and transformation into HCC cells.

pathology↗

bioRxiv2018

RNA-Seq and Protein Mass Spectrometry in Microdissected Kidney Tubules Reveal Signaling Processes that Initiate Lithium-Induced Diabetes Insipidus

ABSTRACT1Lithium salts, used for treatment of bipolar disorder, frequently induce nephrogenic diabetes insipidus (NDI), limiting therapeutic success. NDI is associated with loss of expression of the molecular water channel, aquaporin-2, in the renal collecting duct (CD). Here, we use the methods of systems biology in a well-established rat model of lithium-induced NDI to identify signaling pathways activated at the onset of polyuria. Using single-tubule RNA-Seq, full transcriptomes were determined in microdissected cortical CDs of rats 72 hrs after initiation of lithium chloride (LiCl) administration (vs. time-controls without LiCl). Transcriptome-wide changes in mRNA abundances were mapped to gene sets associated with curated canonical signaling pathways, showing evidence for activation of NF-{kappa}B signaling with induction of genes coding for multiple chemokines as well as most components of the Major Histocompatibility Complex (MHC) Class I antigen-presenting complex. Administration of antiinflammatory doses of dexamethasone to LiCl-treated rats countered the loss of aquaporin-2 protein. RNA-Seq also confirmed prior evidence of a shift from quiescence into the cell cycle with arrest. Time course studies demonstrated an early (12 hrs) increase in multiple immediate early genes including several transcription factors. Protein mass spectrometry in microdissected cortical CDs provided corroborative evidence but also identified decreased abundance of several anti-oxidant proteins. Integration of new data with prior data about lithium effects at a molecular level leads to a signaling model in which lithium increases ERK activation leading to induction of NF-{kappa}B signaling and an inflammatory-like response that represses Aqp2 gene transcription.

systems biology↗