Biology subjects

Nguyen, L. V.

Publications and source records attributed to Nguyen, L. V..

7 recordsLinked to original sources

bioRxiv2026

The molecular architecture of tunneling nanotubes

Tunneling nanotubes (TNTs) are thin intercellular bridges that mediate the exchange of proteins, organelles, and nucleic acids between neighboring cells. They are enriched in tumor cells, implicated in chemotherapy resistance, induced in models of aggregation-based diseases, and their formation is stimulated by viruses that use TNTs to enhance infection. Despite their broad relevance and therapeutic potential, TNT morphology and function remain poorly understood, owing to the absence of clear morphological criteria and the limitations of light microscopy. Here, we establish two complementary systems to study TNT formation and function: stimulation with the pseudorabies viral kinase US3 to model viral transmission, and treatment of acute monocytic leukemia THP-1 cells with daunorubicin to model chemotherapy resistance. Using live-cell imaging, we characterize cytoskeletal organization and bidirectional lysosome transport in both contexts, and apply cryo-correlative light and electron microscopy (cryo-CLEM) with cryogenic electron tomography (cryo-ET) to visualize TNTs in their native state at molecular resolution. We show that TNTs display a rich molecular architecture, comprising actin filaments, microtubules, intermediate filaments, active ribosomes, and diverse organelles including multivesicular bodies, autophagosomes, and lysosomes. Sub-nanometer microtubule reconstructions reveal mixed polarity within individual TNTs, suggesting that both connected cells actively contribute to TNT formation and cargo trafficking. This organization is conserved across both systems, implying that TNT biogenesis reflects a shared cellular program rather than a context-specific response. Our findings provide the first structural framework for TNTs, revealing an unexpectedly rich molecular architecture and laying the groundwork for understanding how TNTs orchestrate intercellular communication in disease.

cell biology↗

bioRxiv2026

Chemotherapy reshapes cellular clonal landscape via persister programs in breast cancer xenografts

Tumor ecosystems evolve in response to chemotherapy, but how treatment reshapes clonal architecture and the transcriptional programs of distinct cell states remains unclear. To investigate the response to chemotherapy by individual single cell-derived clones, we used a high-complexity lentiviral barcoding strategy by genetically labelling with unique, heritable and expressible barcode sequences individual cells from four human breast cancer patient-derived tumor xenograft models. We tracked 3,248 single cell-derived clones across 39 xenografts and profiled 676,292 cells with single cell RNA sequencing. The most striking changes following chemotherapy occurred in non-responsive xenografts, attributed to the emergence of previously minor cell clones that are chemotherapy resistant. In a triple negative model, epithelial and mesenchymal cell states showed distinct sensitivities to chemotherapy. ER+/HER2-models activated stress adaptive programs in response to carboplatin, highlighting DUSP1 and KLF4 as markers of platinum tolerance and a slow cycling, persister-like state. Together, these findings reveal that chemotherapy triggers an immediate and substantial reorganization of the cellular clonal landscape, driven by the selective engagement of stress response programs in cell clones that survive treatment.

cancer biology↗

bioRxiv2025

Selectivity profiles and substrate recognition of Rab phosphorylating kinases

The Rab GTPase switch-2 region is a hotspot for post-translational modifications. Its phosphorylation can determine whether individuals develop Parkinsons disease or not. Other modifications of the same region are catalysed by enzymes from bacterial pathogens when they infect human cells. Here, we profiled a set of kinases including LRRK1, LRRK2, DYRK1A, MST1, and TBK1 for their capability of phosphorylating Rab GTPases. We identified several novel kinase:Rab pairs, such as LRRK1:Rab43 and TBK1:Rab29. Further, we comprehensively assessed what makes a Rab GTPase a good kinase substrate, considering the Rab nucleotide binding state and the Rab primary sequence. In a systematic mutational study, Rab variants with modulated phosphorylation properties were established, leading to the identification of a LRRK2 recognition patch in the Rab 3 helix. A Glu to Arg exchange in that patch increased the phosphorylation 18-fold indicating that Rabs are suboptimal LRRK2 substrates. Given that this effect is also observed in a cellular model, we propose that our variants will be excellent tools for analysing the physiological function of Rab phosphorylation.

biochemistry↗

bioRxiv2023

Development of LRRK2 designed ankyrin-repeat proteins

Leucine rich repeat kinase 2 (LRRK2) is a large multidomain protein containing two catalytic domains, a kinase and a GTPase, as well protein interactions domains, including a WD40 domain. The association of increased LRRK2 kinase activity with both the familial and sporadic forms of Parkinsons disease (PD) has led to intense interest in determining its cellular function. However, small molecule probes that can bind to LRRK2 and report on or affect its activity are needed. Here, we identified a series of high-affinity LRRK2-binding designed ankyrin-repeat proteins (DARPins). One of these DARPins (E11) bound to the LRRK2 WD40 domain with high affinity. LRRK2 bound to DARPin E11 showed improved behavior on cryo-EM grids, resulting in higher resolution LRRK2 structures. DARPin E11 did not affect the catalytic activity of a truncated form of LRRK2 in vitro but decreased the phosphorylation of Rab8A, a LRRK2 substrate, in cells. We also found that DARPin E11 disrupts the formation of microtubule-associated LRRK2 filaments in cells, which are known to require WD40-based dimerization. Thus, DARPin E11 is a new tool to explore the function and dysfunction of LRRK2 and guide the development of LRRK2 kinase inhibitors that target the WD40 domain instead of the kinase.

biochemistry↗

bioRxiv2021

The impact of low input DNA on the reliability of DNA methylation as measured by the Illumina Infinium MethylationEPIC BeadChip

BackgroundDNA methylation (DNAm) is commonly assayed using the Illumina Infinium MethylationEPIC BeadChip, but there is currently little published evidence to define the lower limits of the amount of DNA that can be used whilst preserving data quality. Such evidence is valuable for analyses utilising precious or limited DNA sources. Materials and methodsWe use a single pooled sample of DNA in quadruplicate at three dilutions to define replicability and noise, and an independent population dataset of 328 individuals (from a community-based study including US-born non-Hispanic Black and white persons) to assess the impact of total DNA input on the quality of data generated using the Illumina Infinium MethylationEPIC BeadChip. ResultsData are less reliable and more noisy as DNA input decreases to 40ng, with clear reductions in data quality; however samples with a total input as low as 40ng pass standard quality control tests, and we observe little evidence that low input DNA obscures the associations between DNAm and two phenotypes, age and smoking status. ConclusionsDNA input as low as 40ng can be used with the Illumina Infinium MethylationEPIC BeadChip, provided quality checks and sensitivity analyses are undertaken.

bioinformatics↗

bioRxiv2021

Pre-treatment of Nile tilapia (Oreochromis niloticus) with ozone nanobubbles improve efficacy of heat-killed Streptococcus agalactiae immersion vaccine

Nanobubble technology has shown appealing technical benefits and potential applications in aquaculture. We recently found that treatment with ozone nanobubbles (NB-O3) activated expression of several immune-related genes leading to effective response to subsequent exposure to fish pathogens. In this study, we investigated whether pre-treatment of Nile tilapia (Oreochromis niloticus) with NB-O3 can enhance specific immune responses and improve efficacy of immersion vaccination against Streptococcus agalactiae. Spleen and head kidney of fish in the vaccinated groups showed a substantial upregulation in expression levels of three immunoglobulin classes(IgM, IgD, and IgT) compared with the unvaccinated control groups. At day 21 post-immunization, the relative expression was greatest (approx. 3.2 to 4.1 folds). Both systemic and mucosal IgM antibodies were elicited in vaccinated groups. As the result, the cumulative survival rate of the vaccinated groups was found to be higher than that of the unvaccinated groups, with a relative percent survival (RPS) ranging from 52.9-70.5%. However, fish in the vaccinated groups that received pre-treatment with NB-O3, bacterial antigen uptakes, expression levels of IgM, IgD, and IgT, as well as the specific-IgM antibody levels and percent survival, were all slightly or significantly higher than that of the vaccinated group without pre-treatment with NB-O3. Taken together, our findings suggest that utilizing pre-treatment with NB-O3 may improve the immune response and efficacy of immersion vaccination in Nile tilapia. HighlightsO_LIImmune response and efficacy of a heat-killed Streptococcus agalactiae immersion vaccine for Nile tilapia with and without pre-treatment with NB-O3 were accessed. C_LIO_LIBacterial antigen uptake in the NB-O3-VAC compared to the AT-VAC groups was increased 1.32 and 1.80-fold at 3 and 6 h post-vaccination, respectively. C_LIO_LIVaccinated group that received pre-treatment with NB-O3 had slightly to significantly higher levels of IgM, IgD, and IgT mRNA expression; IgM levels; and survival rate. C_LIO_LIPre-treatment with NB-O3 may be a novel strategy for improving efficacy of immersion vaccine in aquaculture C_LI

molecular biology↗

bioRxiv2021

Ozone nanobubble treatments improve survivability of Nile tilapia (Oreochromis niloticus) challenged with a pathogenic multidrug-resistant Aeromonas hydrophila

Multidrug-resistant (MDR) bacteria has rapidly increased in aquaculture, which highlights the risk of production losses due to diseases and potential public health concerns. Previously, we reported that ozone nanobubbles (NB-O3) were effective at reducing concentrations of pathogenic bacteria in water and modulating fish immunity against pathogens; however, multiple treatments with direct NB-O3 exposures caused alterations to the gills of exposed-fish. Here, we set up a modified recirculation system (MRS) assembled with an NB-O3 device (MRS-NB-O3) to investigate whether MRS-NB-O3 were 1) safe for tilapia (Oreochromis niloticus), 2) effective at reducing bacterial load in rearing water, and 3) improved survivability of Nile tilapia following an immersion challenge with a lethal dose of MDR Aeromonas hydrophila. The results indicated no behavioral abnormalities or mortality of Nile tilapia during the 14 day study using the MRS-NB-O3 system. In the immersion challenge, although high bacterial concentration (~2 x 107 CFU/mL) was used, multiple NB-O3 treatments in the first two days reduced the bacteria between 15.9% to 35.6% of bacterial load in water while bacterial concentration increased 13.1% to 27.9% in the untreated control. There was slight up-regulation of non-specific immune-related genes in the gills of the fish receiving NB-O3 treatments. Most importantly, this treatment significantly improved survivability of Nile tilapia with relative percent survival (RPS) of 64.7 - 66.7% in treated fish and surviving fish developed specific antibody against MDR A. hydrophila. In summary, the result suggests that NB-O3 is a promising alternative to antibiotics to control bacterial diseases, including MDR bacteria, and has high potential for application in recirculation aquaculture system (RAS). HighlightsO_LIMultiple treatments of NB-O3 in a modified recirculation system (MRS) were relatively safe for juvenile Nile tilapia C_LIO_LINB-O3 treatments in MRS significantly improved survivability of Nile tilapia challenged with multidrug-resistant (MDR) A. hydrophila with RPS of 64.7 - 66.7% C_LIO_LIConcentration of MDR A. hydrophila in MRS was reduced by 15.9 to 35.6% following each NB-O3 treatment, and increased by 13.1 to 27.9 % in untreated control C_LIO_LISurviving fish developed specific antibody IgM against MDR A. hydrophila C_LIO_LINB-O3 is a promising non-antibiotic approach to control diseases caused by MDR A. hydrophila C_LI

microbiology↗