Biology subjects

Vu, V.

Publications and source records attributed to Vu, V..

8 recordsLinked to original sources

bioRxiv2026

Chemically induced USP21-KLHL12 association recruits USP21 to COPII assembly

Ubiquitin-specific protease 21 (USP21) is a deubiquitinase implicated in various biological processes and disease states, including cancer. To better understand USP21 protein interaction networks, we used biotin identification proximity labeling, and uncovered USP21 interactions with the cytoskeleton, endosomal, and transcriptional regulators. Notably, a subset of these interactions was modulated by the USP21 selective inhibitor BAY-805. In cells, BAY-805 promoted proximity between USP21 and Kelch like protein KLHL12, a component of the Cullin3 ligase complex. Structural modeling using AlphaFold3, together with hydrogen deuterium exchange mass spectrometry, identified an orthosteric BAY-805 binding site within USP21. Binding of BAY-805 functionally competes with ubiquitin at USP21, thereby enabling USP21 interaction with KLHL12. This BAY-805 induced proximity elicited relocalization of USP21 to KLHL12 containing coat protein complex II (COPII) vesicles. Induced USP21-KLHL12 association led to enlargement of COPII vesicles and impaired transport of large cargoes such as collagen. Overall, our findings reveal a novel chemically induced proximity mechanism in which the monovalent ligand BAY-805 outcompetes ubiquitin from USP21, thereby recruiting USP21 to KLHL12 associated protein complexes.

cell biology↗

bioRxiv2024

Distinct subcircuits within the mesolimbic dopamine system encode the salience and valence of social stimuli

The mesolimbic dopamine (DA) system (MDS) is the canonical "reward" pathway that has been studied extensively in the context of the rewarding properties of sex, food, and drugs of abuse. In contrast, very little is known about the role of the MDS in the processing of the rewarding and aversive properties of social stimuli. Social interactions can be characterized by their salience (i.e., importance) and their rewarding or aversive properties (i.e., valence). Here, we test the novel hypothesis that projections from the medial ventral tegmental area (VTA) to the nucleus accumbens (NAc) core codes for the salience of social stimuli through the phasic release of DA in response to both rewarding and aversive social stimuli. In contrast, we hypothesize that projections from the lateral VTA to the NAc shell codes for the rewarding properties of social stimuli by increasing the tonic release of DA and the aversive properties of social stimuli by reducing the tonic release of DA. Using DA amperometry, which monitors DA signaling with a high degree of temporal and anatomical resolution, we measured DA signaling in the NAc core or shell while rewarding and aversive social interactions were taking place. These findings, as well as additional anatomical and functional studies, provide strong support for the proposed neural circuitry underlying the response of the MDS to social stimuli. Together, these data provide a novel conceptualization of how the functional and anatomical heterogeneity within the MDS detect and distinguish between social salience, social reward, and social aversion. Significance StatementSocial interactions of both positive and negative valence are highly salient stimuli that profoundly impact social behavior and social relationships. Although DA projections from the VTA to the NAc are involved in reward and aversion little is known about their role in the saliency and valence of social stimuli. Here, we report that DA projections from the mVTA to the NAc core signal the salience of social stimuli, whereas projections from the lVTA to the NAc shell signal valence of social stimuli. This work extends our current understanding of the role of DA in the MDS by characterizing its subcircuit connectivity and associated function in the processing of rewarding and aversive social stimuli.

neuroscience↗

bioRxiv2023

Patient-Specific Vascularized Tumor Model: Blocking TAM Recruitment with Multispecific Antibodies Targeting CCR2 and CSF-1R

Tumor-associated inflammation drives cancer progression and therapy resistance, with the infiltration of monocyte-derived tumor-associated macrophages (TAMs) associated with poor prognosis in diverse cancers. Targeting TAMs holds potential against solid tumors, but effective immunotherapies require testing on immunocompetent human models prior to clinical trials. Here, we develop an in vitro model of microvascular networks that incorporates tumor spheroids or patient tissues. By perfusing the vasculature with human monocytes, we investigate monocyte trafficking into the tumor and evaluate immunotherapies targeting the human tumor microenvironment. Our findings demonstrate that macrophages in vascularized breast and lung tumor models can enhance monocyte recruitment via TAM-produced CCL7 and CCL2, mediated by CSF-1R. Additionally, we assess a novel multispecific antibody targeting CCR2, CSF-1R, and neutralizing TGF-{beta}, referred to as CSF1R/CCR2/TGF-{beta} Ab, on monocytes and macrophages using our 3D models. This antibody repolarizes TAMs towards an anti-tumoral M1-like phenotype, reduces monocyte chemoattractant protein secretion, and effectively blocks monocyte migration. Finally, we show that the CSF1R/CCR2/TGF-{beta} Ab inhibits monocyte recruitment in patient-specific vascularized tumor models. Overall, this vascularized tumor model offers valuable insights into monocyte recruitment and enables functional testing of innovative therapeutic antibodies targeting TAMs in the tumor microenvironment (TME).

bioengineering↗

bioRxiv2023

Epigenetic vulnerabilities of leukemia harboring inactivating EZH2 mutations

Epigenetic regulators such as the polycomb repressive complex 2 (PRC2) play a critical role in both normal development and carcinogenesis. Mutations and functional dysregulation of PRC2 complex components such as EZH2 are implicated in various forms of cancer and associated with poor prognosis. This study investigated the epigenetic vulnerabilities of acute myeloid leukemia (AML) and myelodysplastic/myeloproliferative disorders (MDS/MPN) by performing a chemical probe screen in patient cells. Paradoxically, we observed increased sensitivity to EZH2 and EED inhibitors in AML and MDS/MPN patient cells harboring EZH2 mutations. Expression analysis indicated that EZH2 inhibition elicited upregulation of pathways responsible for cell death and growth arrest, specifically in patient cells with mutant EZH2. The identified EZH2 mutations had drastically reduced catalytic activity, resulting in lower cellular H3K27me3 levels and were associated with decreased EZH2 and PRC2 component EED protein levels. Overall, this study provides an important understanding of the role of EZH2 dysregulation in blood cancers and may indicate disease etiology for these poor prognosis AML and MDS/MPN cases.

cancer biology↗

bioRxiv2023

Identifying LasR quorum sensors with improved signal specificity by mapping the sequence-function landscape

Programmable intercellular signaling using components of naturally-occurring quorum sensing can allow for coordinated functions to be engineered in microbial consortia. LuxR-type transcriptional regulators are widely used for this purpose and are activated by homoserine lactone (HSL) signals. However, they often suffer from imperfect molecular discrimination of structurally similar HSLs, causing misregulation within engineered consortia containing multiple HSL signals. Here, we studied one such example, the regulator LasR from Pseudomonas aeruginosa. We elucidated its sequence-function relationship for ligand specificity using targeted protein engineering and multiplexed high-throughput biosensor screening. A pooled combinatorial saturation mutagenesis library (9,486 LasR DNA sequences) was created by mutating six residues in LasRs {beta}5 sheet with single, double, or triple amino acid substitutions. Sort-seq assays were performed in parallel using cognate and non-cognate HSLs to quantify each corresponding sensors response to each HSL signal, which identified hundreds of highly specific variants. Sensor variants identified were individually assayed and exhibited up to 60.6-fold (p = 0.0013) improved relative activation by the cognate signal compared to the wildtype. Interestingly, we uncovered prevalent mutational epistasis and previously unidentified residues contributing to signal specificity. The resulting sensors with negligible signal crosstalk could be broadly applied to engineer bacteria consortia.

synthetic biology↗

bioRxiv2023

Probing the DCAF12 interactions with MAGEA3 and CCT5 C-terminal degrons

DCAF12 is the substrate recognition component of the CRL4 E3 ligase complex that can recognize C-terminal double-glutamic acid degrons to promote degradation of its cognate substrates via the ubiquitin proteasome system. MAGEA3 and CCT5 proteins were reported to be cellular targets of DCAF12. To further characterize the DCAF12 interactions with both MAGEA3 and CCT5, we developed a suite of biophysical and a proximity-based cellular NanoBRET assays showing that both MAGEA3 and CCT5 C-terminal degron peptides interact with DCAF12 in nanomolar affinity in vitro and in cells. Furthermore, we report here the 3.17 [A] cryo-EM structure of DCAF12-DDB1-MAGEA3 complex revealing the key DCAF12 residues involved in C-terminal degron recognition and binding. Our study provides new tools and resources to enable the discovery of small molecule handles targeting the WDR domain of DCAF12 for future PROTAC design and development.

biochemistry↗

bioRxiv2022

PRMT5 regulates ATF4 transcript splicing and oxidative stress response

Protein methyltransferase 5 (PRMT5) symmetrically dimethylates arginine residues leading to regulation of transcription and splicing programs. Although PRMT5 has emerged as an attractive oncology target, the molecular determinants of PRMT5 dependency in cancer remain incompletely understood. Our transcriptomic analysis identified PRMT5 regulation of the activating transcription factor 4 (ATF4) pathway in acute myelogenous leukemia (AML). PRMT5 inhibition resulted in the expression of unstable, intron-retaining ATF4 mRNA that is detained in the nucleus. Concurrently, the decrease in the spliced cytoplasmic transcript of ATF4 led to lower levels of ATF4 protein and downregulation of ATF4 target genes. Upon loss of functional PRMT5, cells with low ATF4 displayed increased oxidative stress, growth arrest, and cellular senescence. Interestingly, leukemia cells with EVI1 oncogene overexpression demonstrated dependence on PRMT5 function. EVI1 and ATF4 regulated gene signatures were inversely correlated. We show that EVI1-high AML cells have reduced ATF4 levels, elevated baseline reactive oxygen species and increased sensitivity to PRMT5 inhibition. Thus, EVI1-high cells demonstrate dependence on PRMT5 function and regulation of oxidative stress response. Overall, our findings identify the PRMT5-ATF4 axis to be safeguarding the cellular redox balance that is especially important in high oxidative stress states, such as those that occur with EVI1 overexpression.

cancer biology↗

bioRxiv2021

Design and execution of a Verification, Validation, and Uncertainty Quantification plan for a numerical model of left ventricular flow after LVAD implantation

Backgroundleft ventricular assist devices (LVADs) are implantable pumps that act as a life support therapy for patients with severe heart failure. Despite improving the survival rate, LVAD therapy can carry major complications. Particularly, the flow distortion introduced by the LVAD in the left ventricle (LV) may induce thrombus formation. While previous works have used numerical models to study the impact of multiple variables in the intra-LV stagnation regions, a comprehensive validation analysis has never been executed. The main goal of this work is to present a model of the LV-LVAD system and to design and follow a verification, validation and uncertainty quantification (VVUQ) plan based on the ASME V&V40 and V&V20 standards to ensure credible predictions. MethodsThe experiment used to validate the simulation is the SDSU cardiac simulator, a bench mock-up of the cardiovascular system that allows mimicking multiple operation conditions for the heart-LVAD system. The numerical model is based on Alya, the BSCs in-house platform for numerical modelling. Alya solves the Navier-Stokes equation with an Arbitrarian Lagrangian-Eulerian (ALE) formulation in a deformable ventricle and includes pressure-driven valves, a 0D Windkessel model for the arterial output and a LVAD boundary condition modeled through a dynamic pressure-flow performance curve. The designed VVUQ plan involves: (a) a risk analysis and the associated credibility goals; (b) a verification stage to ensure correctness in the numerical solution procedure; (c) a sensitivity analysis to quantify the impact of the inputs on the four quantities of interest (QoIs) (average aortic root flow [Formula], maximum aortic root flow [Formula], average LVAD flow [Formula], and maximum LVAD flow [Formula]); (d) an uncertainty quantification using six validation experiments that include extreme operating conditions. ResultsNumerical code verification tests ensured correctness of the solution procedure and numerical calculation verification showed small numerical errors. The total Sobol indices obtained during the sensitivity analysis demonstrated that the ejection fraction, the heart rate, and the pump performance curve coefficients are the most impactful inputs for the analysed QoIs. The Minkowski norm is used as validation metric for the uncertainty quantification. It shows that the midpoint cases have more accurate results when compared to the extreme cases. The total computational cost of the simulations was above 100 [core-years] executed in around three weeks time span in Marenostrum IV supercomputer. ConclusionsThis work details a novel numerical model for the LV-LVAD system, that is supported by the design and execution of a VVUQ plan created following recognised international standards. We present a methodology demonstrating that stringent VVUQ according to ASME standards is feasible but computationally expensive.

bioengineering↗