About

Luiza Mendonça, PhD, is an Assistant Professor at the University of Minnesota Medical School. Her research employs integrative imaging approaches, including cryo-electron microscopy (cryoEM/ET), cryo-focused ion beam/scanning electron microscopy (cryoFIB/SEM), and correlative light and electron microscopy (CLEM), to investigate cellular pathways exploited by RNA viruses. Her work aims to identify novel molecular targets for therapeutic interventions, with the potential to reveal molecular weak links that can be exploited to combat viral diseases such as AIDS and COVID-19. Dr. Mendonça has presented her research at various symposia, including the Schachtele Symposium at the University of Minnesota Dental School and the Institute for Molecular Virology, demonstrating her active engagement in the scientific community.

Research topics

• Biology
• Medicine
• Virology
• Immunology
• Molecular biology

Selected publications

• ROCKY MOUNTAIN SPOTTED FEVER INDUCED MYOCARDITIS

  Journal of the American College of Cardiology · 2025-03-30

  articleOpen access
  PublisherDOI

• High-Yield and Quantitative Purification Method for HIV Which Minimizes Forces Applied to Virions Utilized to Investigate Maturation of HIV-1 via Cryo-Electron Tomography

  Viruses · 2025-03-03 · 2 citations

  articleOpen access

  HIV is a lentivirus characterized by its cone shaped mature core. Visualization and structural examination of HIV requires the purification of virions to high concentrations. The yield and integrity of these virions are crucial for ensuring a uniform representation of all viral particles in subsequent analyses. In this study, we present a method for the purification of HIV virions which minimizes the forces applied to virions while maximizing the efficiency of collection. This method, which relies on virion sedimentation simulations, allows us to capture between 1000 and 5000 HIV virions released from individual HEK293 cells after transfection with the NL4.3 HIV backbone. We utilized this approach to investigate HIV core formation from several constructs: pNL4-3(RT:D185A&D186A) with an inactive reverse transcriptase, NL4.3(IN: V165A&R166A) with a type-II integrase mutation, and NL4.3(Ψ: Δ(105–278)&Δ(301–332)) featuring an edited Ψ packaging signal. Notably, virions from NL4.3(Ψ: Δ(105–278)&Δ(301–332)) displayed a mixed population, comprising immature virions, empty cores, and cores with detectable internal density. Conversely, virions derived from NL4.3(IN: V165A&R166A) exhibited a type II integrase mutant phenotype characterized by empty cores and RNP density localized around the cores, consistent with previous studies. In contrast, virions released from pNL4-3(RT:D185A&D186A) displayed mature cores containing detectable RNP density. We suggest that the sedimentation simulations developed in this study can facilitate the characterization of enveloped viruses.

  PublisherOA PDFDOI

• Characterization of bi-segmented and tri-segmented recombinant Pichinde virus particles

  Journal of Virology · 2024-09-12 · 3 citations

  articleOpen access

  Mammarenaviruses include several highly virulent pathogens (e.g., Lassa virus) capable of causing severe hemorrhagic fever diseases for which there are no approved vaccines and limited treatment options. Mammarenaviruses are enveloped, bi-segmented ambisense RNA viruses. There is limited knowledge about cellular proteins incorporated into progeny virion particles and their potential biological roles in viral infection. Pichinde virus (PICV) is a prototypic arenavirus used to characterize mammarenavirus replication and pathogenesis. We have developed a recombinant PICV with a tri-segmented RNA genome as a viral vector platform. Whether the tri-segmented virion differs from the wild-type bi-segmented one in viral particle morphology and protein composition has not been addressed. In this study, recombinant PICV (rPICV) virions with a bi-segmented (rP18bi) and a tri-segmented (rP18tri) genome were purified by density-gradient ultracentrifugation and analyzed by cryo-electron microscopy and mass spectrometry. Both virion types are pleomorphic with spherical morphology and have no significant difference in size despite rP18tri having denser particles. Both virion types also contain similar sets of cellular proteins. Among the highly enriched virion-associated cellular proteins are components of the endosomal sorting complex required for transport pathway and vesicle trafficking, such as ALIX, Tsg101, VPS, CHMP, and Ras-associated binding proteins, which have known functions in virus assembly and budding. Other enriched cellular proteins include peripheral and transmembrane proteins, chaperone proteins, and ribosomal proteins; their biological roles in viral infection warrant further analysis. Our study provides important insights into mammarenavirus particle formation and aids in the future development of viral vectors and antiviral discovery.IMPORTANCEMammarenaviruses, such as Lassa virus, are enveloped RNA viruses that can cause severe hemorrhagic fever diseases (Lassa fever) with no approved vaccine and limited therapeutic options. Cellular proteins incorporated into progeny virion particles and their biological roles in mammarenavirus infection have not been well characterized. Pichinde virus (PICV) is a prototypic mammarenavirus used as a surrogate model for Lassa fever. We used cryo-electron microscopy and proteomic analysis to characterize the morphology and protein contents of the purified PICV particles that package either two (bi-segmented) or three (tri-segmented) genomic RNA segments. Our results demonstrate a similar virion morphology but different particle density for the bi- and tri-segmented viral particles and reveal major virion-associated cellular proteins. This study provides important insights into the virus-host interactions that can be used for antiviral development and optimizing arenavirus-based vaccine vectors.

  PublisherOA PDFDOI

• Enhanced Yield and Gentle Purification of HIV for Cryo-Electron Tomography Analysis of Virion Maturation

  bioRxiv (Cold Spring Harbor Laboratory) · 2024-12-12

  preprintOpen access

  Abstract HIV is a lentivirus characterized by the formation of its mature core. Visualization and structural examination of HIV requires purification of virions to high concentrations. The yield and integrity of these virions are crucial for ensuring a uniform representation of all viral particles in subsequent analyses. In this study, we present a method for purification of HIV virions which minimizes forces applied to virions while maximizing the efficiency of collection. This method allows us to capture between 1,000 and 5,000 HIV virions released from individual HEK293 cells after transfection with the NL4.3 HIV backbone, a 10 fold advantage over other methods. We utilized this approach to investigate HIV core formation from several constructs: pNL4-3(RT:D 185 A&D 186 A) with an inactive reverse transcriptase, NL4.3(IN: V 165 A&R 166 A) with a type-II integrase mutation, and NL4.3(Ѱ: Δ(105-278)&Δ(301-332)) featuring an edited Ѱ packaging signal. Notably, virions from NL4.3(Ѱ: Δ(105-278)&Δ(301-332)) displayed a mixed population, comprising immature virions, empty cores, and cores with detectable internal density. Conversely, virions derived from NL4.3(IN: V 165 A&R 166 A) exhibited a type II integrase mutant phenotype characterized by empty cores and RNP density localized around the cores, consistent with previous studies. In contrast, virions released from pNL4-3(RT:D 185 A&D 186 A) displayed mature cores containing detectable RNP density. We suggest that the purification methods developed in this study can significantly facilitate the characterization of enveloped viruses.

  PublisherOA PDFDOI

• Editorial: HTLV-1: addressing unmet research needs, volume II

  Frontiers in Microbiology · 2023-10-26

  editorialOpen accessSenior authorCorresponding

  signficantly higher. HTLV transmission occurs by verQcal transmission (predominantly via breastmilk), or horizontally through sexual intercourse, blood transfusions or shared syringes.The current therapeuQcs for HTLV-associated diseases have very limited benefits.Therefore, the development of new therapies is of crucial importance. More basic, applied and clinical research is needed in order to develop HTLV vaccines and novel treatment opQons against ATL and HAM/TSP. This research topic aims to present the recent discoveries related to potenQal novel drugs for HTLV-1-related diseases, new research on the link between anQsense proteins and diseases, as well as clinical updates on how HTLV-1 infecQon affects pregnancy outcomes. Komatsu et al. examined the clinical data of HTLV-1-infected and seroconverted pregnant mothers, who enlisted in the database of a prefecture-wide antenatal adult T-cell leukemia prevenQon program from 2011-2018 in Nagasaki, Japan. They produced the first report with chronological data on HTLV-1 proviral load on this populaQon. While InfecQon had no adverse effects on pregnancy outcomes, a posiQve correlaQon between the increase of proviral load and the number of parity in HTLV-1 posiQve women was observed. This report underscores the importance of though monitoring for HTLV-1 posiQve mothers and infants.Ernzen et al. evaluated the potenQal of an inhibitor of the Protein arginine methyltransferase 5 (PRMT5) to hinder ATL inducQon. PRMT5 is involved in epigeneQcs regulaQon and is overexpressed in HTLV-1-transformed cell lines. They demonstrated that the PRMT5-inhibitor EPZ015666 could prevent the HTLV-1-driven immortalizaQon of cells and exhibited selecQve toxicity in HTLV-1 transformed cell lines as well as in HTLV-1-infected primary T-cells in a dose-and Qme-dependent manner. Moreover, the treatment employing the PRMT5inhibitor significantly reduces the mortality rate in murine models. The authors conclude that PRMT5 is implicated in the HTLV-1-mediated T-cell transformaQon and disease pathogenesis in vivo and that EPZ015666 may have therapeuQc value for treatment of ATL, highlighQng the importance of the PRMT5 as a potenQal new target to the development of anQ-ATL drugs.Another study aimed to address the therapeuQcal potenQal of histone methyl transferase inhibitors in HTLV-1-associated myelopathy (HAM). Koseki et al. tested if EZH1/EZH2 (enhancer of zeste homolog 1/2) inhibitors could reduce the cell proliferaQon, HTLV-1 proviral load and modulate cytokine producQon. They conclude that EZH1/2 dual inhibitors can reduce the proliferaQon of HTLV-1-infected cells and HAM-derived peripheral blood mononuclear cells (PBMC) in a dose-dependent fashion. AddiQonally, the drugs could suppress the exacerbated immune response in infected cells, as observed by the increase of IL-10, although the levels of IFN-gamma and TNF-alfa were not reduced. These data indicates that the EZH1/2 dual inhibitors may be effecQve therapeuQc agents for HAM. To further confirm these promising data, the authors suggest the conducQon of proof-of-concept clinical trials in paQents with HAM.Several retroviruses share an intriguing feature: they have an open reading frame on the negaQve strand of their genomes, which results in natural anQsense transcripts. Lin et al.summarized the main recent findings regarding the anQsense transcripts of HTLV-1, HTLV-2, and HIV-1, shedding light on the role of these transcripts and proteins on viral lifecycle, and also on the cellular factors that regulate their transcripQon. They highlighted that anQsense mRNAs of HTLV-1, HTLV-2 and HIV-1 are transcribed from 3' LTR and have mulQple transcripQon start sites.All of them have their expression driven by TATA-less promoters and these anQsense transcripts can give rise to proteins. This review also called afenQon to the role of myocyte enhancer factor-2 (MEF-2) protein family on the regulaQon of HBZ (HTLV-1 basic leucine zipper factor) transcripQon. The authors also provided and overview of the role of HBZ in ATL disorder: HBZ has several host interacQng partners involved in the promoQon of T-cell proliferaQon, transiQon of G1/S phase, transcripQonal factors and coacQvators (such as p300/CPB) and it is considered an inhibitor of tax-mediated transcripQon, which is a funcQon also played by its HTLV-2 anQsense counterpart, the protein APH-2 (HTLV-2 anQsense protein). The authors indicate the importance of further research to elucidate the mechanism of MEF-2 family in HBZ regulaQon and neoplasia.They also suggest anQsense genes may be potenQal targets for therapeuQc intervenQons. To date, CRISPR silencing of HBZ was deleterious for ATL cell lines, xenograhs and paQent samples, encouraging further studies in vivo models.Liu et al. invesQgated the potenQal interacQon between HBZ, the nucleolar proteins nucleophosmin (NPM1/B23) and nucleolin (C23) and their implicaQons on the pathogenesis of ATL. They have proven the interacQon between NPM1/B23 with HBZ in the nucleolar region in different cell lines. The authors also observed C23 interacQon with HBZ. InteresQngly, an isoform of HBZ, sHBZ, can interact with its own shbz mRNA. As both proteins are founded in nucleolar regions and interacts with each other, they hypothesize that NPM1/B23 may form a complex with HBZ and C23. AddiQonally, downregulaQon of NPM1/B23 led to a reducQon of sHBZ expression along with an increase of Tax expression. In this work, the authors presented novel interacQon partners for HBZ, and suggested that these interacQons may be involved in HBZ funcQons, contribuQng to ATL development, calling afenQon to the importance of further studies to understand the outcome of these interacQons in the pathogenesis of ATL.In summary, this special issue offers valuable insights into potenQal targets and promising drugs that could broaden the treatment opQons for diseases caused by HTLV-1. AddiQonally, it presents recent discoveries related to seroconversion's effects on pregnancy outcomes, explores new partners of the anQsense protein HBZ, and sheds light on its role in ATL pathogenesis. These findings collecQvely enhance our understanding of the mechanisms underlying HTLV-1-related disorders.

  PublisherOA PDFDOI

• JoVE Video Dataset

  2023-12-16

  articleOpen accessSenior author

  In situ cellular cryotomography is a powerful technique for studying complex objects in their native frozen-hydrated cellular context, making it highly relevant to cellular biology and virology. The potential of combining cryotomography with other microscopy modalities makes it a perfect technique for integrative and correlative imaging. However, sample preparation for in situ cellular tomography is not straightforward, as cells do not readily attach and stretch over the electron microscopy grid. Additionally, the grids themselves are fragile and can break if handled too forcefully, resulting in the loss of imageable areas. The geometry of tissue culture dishes can also pose a challenge when manipulating the grids with tweezers. Here, we describe the tips and tricks to overcome these (and other) challenges and prepare good-quality samples for in situ cellular cryotomography and correlative imaging of adherent mammalian cells. With continued advances in cryomicroscopy technology, this technique holds enormous promise for advancing our understanding of complex biological systems.

  PublisherOA PDFDOI

• Cryo-electron tomography of ChAdOx spikes (AZD2816)

  EMPIAR dataset · 2023-03-10

  datasetOpen access

  EMPIAR, the Electron Microscopy Public Image Archive centered at EMBL-EBI, is a public resource for raw electron microscopy images related to EMDB, contains micrographs, particle sets and tilt-series.

  PublisherDOI

• ChAdOx1 COVID vaccines express RBD open prefusion SARS-CoV-2 spikes on the cell surface

  bioRxiv (Cold Spring Harbor Laboratory) · 2023-05-24 · 2 citations

  preprintOpen access

  Abstract Vaccines against SARS-CoV-2 have been proven to be an effective means of decreasing COVID-19 mortality, hospitalization rates, and transmission. One of the vaccines deployed worldwide is ChAdOx1 nCoV-19, which uses an adenovirus vector to drive the expression of the original SARS-CoV-2 spike on the surface of transduced cells. Using cryo-electron tomography and subtomogram averaging, we determined the native structures of the vaccine product expressed on cell surfaces in situ . We show that ChAdOx1-vectored vaccines expressing the Beta SARS-CoV-2 variant produce abundant native prefusion spikes predominantly in one-RBD-up conformation. Furthermore, the ChAdOx1 vectored HexaPro stabilized spike yields higher cell surface expression, enhanced RBD exposure, and reduced shedding of S1 compared to the wild-type. We demonstrate in situ structure determination as a powerful means for studying antigen design options in future vaccine development against emerging novel SARS-CoV-2 variants and broadly against other infectious viruses.

  PublisherOA PDFDOI

• HIV-2 Immature Particle Morphology Provides Insights into Gag Lattice Stability and Virus Maturation

  Journal of Molecular Biology · 2023-05-05 · 14 citations

  articleOpen access
  PublisherOA PDFDOI

• Sample Preparation for <em>In Situ</em> Cryotomography of Mammalian Cells

  Journal of Visualized Experiments · 2023-12-15 · 1 citations

  articleOpen accessSenior author

  In situ cellular cryotomography is a powerful technique for studying complex objects in their native frozen-hydrated cellular context, making it highly relevant to cellular biology and virology. The potential of combining cryotomography with other microscopy modalities makes it a perfect technique for integrative and correlative imaging. However, sample preparation for in situ cellular tomography is not straightforward, as cells do not readily attach and stretch over the electron microscopy grid. Additionally, the grids themselves are fragile and can break if handled too forcefully, resulting in the loss of imageable areas. The geometry of tissue culture dishes can also pose a challenge when manipulating the grids with tweezers. Here, we describe the tips and tricks to overcome these (and other) challenges and prepare good-quality samples for in situ cellular cryotomography and correlative imaging of adherent mammalian cells. With continued advances in cryomicroscopy technology, this technique holds enormous promise for advancing our understanding of complex biological systems.

  PublisherOA PDFDOI

Frequent coauthors

• Peijun Zhang

  Diamond Light Source

  113 shared

• Tao Ni

  75 shared

• Anna-Sophia Krebs

  University of Oxford

  72 shared

• Andrew Howe

  South London and Maudsley NHS Foundation Trust

  67 shared

• Yuewen Sheng

  Diamond Light Source

  66 shared

• Laura C. Zanetti-Domingues

  Science and Technology Facilities Council

  66 shared

• Mohamed A. Koronfel

  62 shared

• Maria Harkiolaki

  University of Warwick

  62 shared