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Article: Multiplexed 3D Analysis of Immune States and Niches in Human Tissue.

Yapp, Clarence / Nirmal, Ajit J / Zhou, Felix / Maliga, Zoltan / Tefft, Juliann B / Llopis, Paula Montero / Murphy, George F / Lian, Christine G / Danuser, Gaudenz / Santagata, Sandro / Sorger, Peter K

bioRxiv : the preprint server for biology

2024

Abstract: Tissue homeostasis and the emergence of disease are controlled by changes in the proportions of resident and recruited cells, their organization into cellular neighbourhoods, and their interactions with acellular tissue components. Highly multiplexed ... ...

Abstract	Tissue homeostasis and the emergence of disease are controlled by changes in the proportions of resident and recruited cells, their organization into cellular neighbourhoods, and their interactions with acellular tissue components. Highly multiplexed tissue profiling (spatial omics)
Language	English
Publishing date	2024-03-28
Publishing country	United States
Document type	Preprint
DOI	10.1101/2023.11.10.566670
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Article ; Online: Cell wall synthesis and remodelling dynamics determine division site architecture and cell shape in Escherichia coli.

Navarro, Paula P / Vettiger, Andrea / Ananda, Virly Y / Llopis, Paula Montero / Allolio, Christoph / Bernhardt, Thomas G / Chao, Luke H

Nature microbiology

2022 Volume 7, Issue 10, Page(s) 1621–1634

Abstract: The bacterial division apparatus catalyses the synthesis and remodelling of septal peptidoglycan (sPG) to build the cell wall layer that fortifies the daughter cell poles. Understanding of this essential process has been limited by the lack of native ... ...

Abstract	The bacterial division apparatus catalyses the synthesis and remodelling of septal peptidoglycan (sPG) to build the cell wall layer that fortifies the daughter cell poles. Understanding of this essential process has been limited by the lack of native three-dimensional views of developing septa. Here, we apply state-of-the-art cryogenic electron tomography (cryo-ET) and fluorescence microscopy to visualize the division site architecture and sPG biogenesis dynamics of the Gram-negative bacterium Escherichia coli. We identify a wedge-like sPG structure that fortifies the ingrowing septum. Experiments with strains defective in sPG biogenesis revealed that the septal architecture and mode of division can be modified to more closely resemble that of other Gram-negative (Caulobacter crescentus) or Gram-positive (Staphylococcus aureus) bacteria, suggesting that a conserved mechanism underlies the formation of different septal morphologies. Finally, analysis of mutants impaired in amidase activation (ΔenvC ΔnlpD) showed that cell wall remodelling affects the placement and stability of the cytokinetic ring. Taken together, our results support a model in which competition between the cell elongation and division machineries determines the shape of cell constrictions and the poles they form. They also highlight how the activity of the division system can be modulated to help generate the diverse array of shapes observed in the bacterial domain.
MeSH term(s)	Amidohydrolases ; Cell Division ; Cell Shape ; Cell Wall ; Escherichia coli/physiology ; Peptidoglycan
Chemical Substances	Peptidoglycan ; Amidohydrolases (EC 3.5.-)
Language	English
Publishing date	2022-09-12
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Research Support, N.I.H., Extramural
ISSN	2058-5276
ISSN (online)	2058-5276
DOI	10.1038/s41564-022-01210-z
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Article: Toxicity, uptake, and nuclear translocation of ingested micro-nanoplastics in an in vitro model of the small intestinal epithelium

DeLoid, Glen M. / Cao, Xiaoqiong / Bitounis, Dimitrios / Singh, Dilpreet / Llopis, Paula Montero / Buckley, Brian / Demokritou, Philip

Food and chemical toxicology. 2021 Dec., v. 158

2021

Abstract: Despite mounting evidence of increasing micro- and nanoplastics (MNPs) in natural environments, food, and drinking water, little is known of the potential health hazards of MNPs ingestion. We assessed toxicity and uptake of environmentally relevant MNPs ... ...

Abstract	Despite mounting evidence of increasing micro- and nanoplastics (MNPs) in natural environments, food, and drinking water, little is known of the potential health hazards of MNPs ingestion. We assessed toxicity and uptake of environmentally relevant MNPs in an in vitro small intestinal epithelium (SIE). Test MNPs included 25 and 1000 nm polystyrene (PS) microspheres (PS25 and PS1K); 25, 100, and 1000 nm carboxyl modified PS spheres (PS25C, PS100C, and PS1KC), and secondary MNPs from incinerated polyethylene (PEI). MNPs were subjected to 3-phase digestion to mimic transformations in the gastrointestinal tract (GIT) and digestas applied to the SIE. Carboxylated MNPs significantly reduced viability and increased permeability to 3 kD dextran. Uptake of carboxyl PS materials was size dependent, with significantly greater uptake of PS25C. Fluorescence confocal imaging showed some PS25C agglomerates entering cells independent of endosomes (suggesting diffusion), others within actin shells (suggesting phagocytosis), and many free within the epithelial cells, including agglomerates within nuclei. Pre-treatment with the dynamin inhibitor Dyngo partially reduced PS25 translocation, suggesting a potential role for endocytosis. These findings suggest that ingestion exposures to MNPs could have serious health consequences and underscore the urgent need for additional detailed studies of the potential hazards of ingested MNPs.
Keywords	actin ; carboxylation ; dextran ; digestion ; digestive tract ; dynamins ; endosomes ; fluorescence ; ingestion ; intestinal mucosa ; microparticles ; models ; nanoplastics ; permeability ; phagocytosis ; polyethylene ; polystyrenes ; toxicity ; toxicology ; viability
Language	English
Dates of publication	2021-12
Publishing place	Elsevier Ltd
Document type	Article
ZDB-ID	782617-5
ISSN	1873-6351 ; 0278-6915
ISSN (online)	1873-6351
ISSN	0278-6915
DOI	10.1016/j.fct.2021.112609
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Article ; Online: Toxicity, uptake, and nuclear translocation of ingested micro-nanoplastics in an in vitro model of the small intestinal epithelium.

DeLoid, Glen M / Cao, Xiaoqiong / Bitounis, Dimitrios / Singh, Dilpreet / Llopis, Paula Montero / Buckley, Brian / Demokritou, Philip

Food and chemical toxicology : an international journal published for the British Industrial Biological Research Association

2021 Volume 158, Page(s) 112609

Abstract	Despite mounting evidence of increasing micro- and nanoplastics (MNPs) in natural environments, food, and drinking water, little is known of the potential health hazards of MNPs ingestion. We assessed toxicity and uptake of environmentally relevant MNPs in an in vitro small intestinal epithelium (SIE). Test MNPs included 25 and 1000 nm polystyrene (PS) microspheres (PS25 and PS1K); 25, 100, and 1000 nm carboxyl modified PS spheres (PS25C, PS100C, and PS1KC), and secondary MNPs from incinerated polyethylene (PEI). MNPs were subjected to 3-phase digestion to mimic transformations in the gastrointestinal tract (GIT) and digestas applied to the SIE. Carboxylated MNPs significantly reduced viability and increased permeability to 3 kD dextran. Uptake of carboxyl PS materials was size dependent, with significantly greater uptake of PS25C. Fluorescence confocal imaging showed some PS25C agglomerates entering cells independent of endosomes (suggesting diffusion), others within actin shells (suggesting phagocytosis), and many free within the epithelial cells, including agglomerates within nuclei. Pre-treatment with the dynamin inhibitor Dyngo partially reduced PS25 translocation, suggesting a potential role for endocytosis. These findings suggest that ingestion exposures to MNPs could have serious health consequences and underscore the urgent need for additional detailed studies of the potential hazards of ingested MNPs.
MeSH term(s)	Actins ; Biological Transport ; Caco-2 Cells ; Cell Nucleus ; Endocytosis ; Environmental Exposure/adverse effects ; Epithelial Cells/metabolism ; HT29 Cells ; Humans ; Intestinal Mucosa/metabolism ; Intestine, Small/metabolism ; Microplastics/metabolism ; Microplastics/toxicity ; Microspheres ; Nanostructures ; Optical Imaging ; Particle Size ; Permeability ; Polyethylene/chemistry ; Polystyrenes/metabolism ; Polystyrenes/toxicity ; Water Pollutants, Chemical/metabolism ; Water Pollutants, Chemical/toxicity
Chemical Substances	Actins ; Microplastics ; Polystyrenes ; Water Pollutants, Chemical ; Polyethylene (9002-88-4)
Language	English
Publishing date	2021-10-18
Publishing country	England
Document type	Journal Article
ZDB-ID	782617-5
ISSN	1873-6351 ; 0278-6915
ISSN (online)	1873-6351
ISSN	0278-6915
DOI	10.1016/j.fct.2021.112609
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Article ; Online: A biologist's guide to planning and performing quantitative bioimaging experiments.

Senft, Rebecca A / Diaz-Rohrer, Barbara / Colarusso, Pina / Swift, Lucy / Jamali, Nasim / Jambor, Helena / Pengo, Thomas / Brideau, Craig / Llopis, Paula Montero / Uhlmann, Virginie / Kirk, Jason / Gonzales, Kevin Andrew / Bankhead, Peter / Evans, Edward L / Eliceiri, Kevin W / Cimini, Beth A

PLoS biology

2023 Volume 21, Issue 6, Page(s) e3002167

Abstract: Technological advancements in biology and microscopy have empowered a transition from bioimaging as an observational method to a quantitative one. However, as biologists are adopting quantitative bioimaging and these experiments become more complex, ... ...

Abstract	Technological advancements in biology and microscopy have empowered a transition from bioimaging as an observational method to a quantitative one. However, as biologists are adopting quantitative bioimaging and these experiments become more complex, researchers need additional expertise to carry out this work in a rigorous and reproducible manner. This Essay provides a navigational guide for experimental biologists to aid understanding of quantitative bioimaging from sample preparation through to image acquisition, image analysis, and data interpretation. We discuss the interconnectedness of these steps, and for each, we provide general recommendations, key questions to consider, and links to high-quality open-access resources for further learning. This synthesis of information will empower biologists to plan and execute rigorous quantitative bioimaging experiments efficiently.
MeSH term(s)	Microscopy ; Image Processing, Computer-Assisted
Language	English
Publishing date	2023-06-27
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Research Support, N.I.H., Extramural
ZDB-ID	2126776-5
ISSN	1545-7885 ; 1544-9173
ISSN (online)	1545-7885
ISSN	1544-9173
DOI	10.1371/journal.pbio.3002167
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Article ; Online: MethodsJ2: a software tool to capture metadata and generate comprehensive microscopy methods text.

Ryan, Joel / Pengo, Thomas / Rigano, Alex / Llopis, Paula Montero / Itano, Michelle S / Cameron, Lisa A / Marqués, Guillermo / Strambio-De-Castillia, Caterina / Sanders, Mark A / Brown, Claire M

Nature methods

2021 Volume 18, Issue 12, Page(s) 1414–1416

MeSH term(s)	Cell Nucleus/ultrastructure ; Chromatin/ultrastructure ; Humans ; Image Processing, Computer-Assisted/methods ; Image Processing, Computer-Assisted/standards ; Medical Informatics/statistics & numerical data ; Metadata/statistics & numerical data ; Microscopy/methods ; Software
Chemical Substances	Chromatin
Language	English
Publishing date	2021-10-28
Publishing country	United States
Document type	Letter ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
ZDB-ID	2169522-2
ISSN	1548-7105 ; 1548-7091
ISSN (online)	1548-7105
ISSN	1548-7091
DOI	10.1038/s41592-021-01290-5
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Article ; Online: Hepatitis C virus NS3-4A protease regulates the lipid environment for RNA replication by cleaving host enzyme 24-dehydrocholesterol reductase.

Tallorin, Lorillee / Villareal, Valerie A / Hsia, Chih-Yun / Rodgers, Mary A / Burri, Dominique J / Pfeil, Marc-Philipp / Llopis, Paula Montero / Lindenbach, Brett D / Yang, Priscilla L

The Journal of biological chemistry

2020 Volume 295, Issue 35, Page(s) 12426–12436

Abstract: Many RNA viruses create specialized membranes for genome replication by manipulating host lipid metabolism and trafficking, but in most cases, we do not know the molecular mechanisms responsible or how specific lipids may impact the associated membrane ... ...

Abstract	Many RNA viruses create specialized membranes for genome replication by manipulating host lipid metabolism and trafficking, but in most cases, we do not know the molecular mechanisms responsible or how specific lipids may impact the associated membrane and viral process. For example, hepatitis C virus (HCV) causes a specific, large-fold increase in the steady-state abundance of intracellular desmosterol, an immediate precursor of cholesterol, resulting in increased fluidity of the membrane where HCV RNA replication occurs. Here, we establish the mechanism responsible for HCV's effect on intracellular desmosterol, whereby the HCV NS3-4A protease controls activity of 24-dehydrocholesterol reductase (DHCR24), the enzyme that catalyzes conversion of desmosterol to cholesterol. Our cumulative evidence for the proposed mechanism includes immunofluorescence microscopy experiments showing co-occurrence of DHCR24 and HCV NS3-4A protease; formation of an additional, faster-migrating DHCR24 species (DHCR24) in cells harboring a HCV subgenomic replicon RNA or ectopically expressing NS3-4A; and biochemical evidence that NS3-4A cleaves DHCR24 to produce DHCR24
MeSH term(s)	Cell Line, Tumor ; Hepacivirus/enzymology ; Hepacivirus/genetics ; Humans ; Lipid Metabolism ; Membrane Lipids/genetics ; Membrane Lipids/metabolism ; Nerve Tissue Proteins/genetics ; Nerve Tissue Proteins/metabolism ; Oxidoreductases Acting on CH-CH Group Donors/genetics ; Oxidoreductases Acting on CH-CH Group Donors/metabolism ; Proteolysis ; RNA, Viral/biosynthesis ; RNA, Viral/genetics ; Serine Proteases/genetics ; Serine Proteases/metabolism ; Viral Nonstructural Proteins/genetics ; Viral Nonstructural Proteins/metabolism
Chemical Substances	Membrane Lipids ; Nerve Tissue Proteins ; RNA, Viral ; Viral Nonstructural Proteins ; Oxidoreductases Acting on CH-CH Group Donors (EC 1.3.-) ; DHCR24 protein, human (EC 1.3.1.-) ; NS3-4A serine protease, Hepatitis C virus (EC 3.4.-) ; Serine Proteases (EC 3.4.-)
Language	English
Publishing date	2020-07-08
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
ZDB-ID	2997-x
ISSN	1083-351X ; 0021-9258
ISSN (online)	1083-351X
ISSN	0021-9258
DOI	10.1074/jbc.RA120.013455
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Article ; Online: CAR T cell killing requires the IFNγR pathway in solid but not liquid tumours.

Nature

2022 Volume 604, Issue 7906, Page(s) 563–570

Abstract: Chimeric antigen receptor (CAR) therapy has had a transformative effect on the treatment of haematologic ... ...

Abstract	Chimeric antigen receptor (CAR) therapy has had a transformative effect on the treatment of haematologic malignancies
MeSH term(s)	Cell Death ; Glioblastoma/genetics ; Glioblastoma/therapy ; Humans ; Immunotherapy, Adoptive ; Receptors, Chimeric Antigen ; T-Lymphocytes/pathology
Chemical Substances	Receptors, Chimeric Antigen
Language	English
Publishing date	2022-04-13
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Research Support, N.I.H., Extramural ; Research Support, U.S. Gov't, Non-P.H.S.
ZDB-ID	120714-3
ISSN	1476-4687 ; 0028-0836
ISSN (online)	1476-4687
ISSN	0028-0836
DOI	10.1038/s41586-022-04585-5
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Article ; Online: Suppression of amber codons in Caulobacter crescentus by the orthogonal Escherichia coli histidyl-tRNA synthetase/tRNAHis pair.

Ko, Jae-hyeong / Llopis, Paula Montero / Heinritz, Jennifer / Jacobs-Wagner, Christine / Söll, Dieter

PloS one

2013 Volume 8, Issue 12, Page(s) e83630

Abstract: While translational read-through of stop codons by suppressor tRNAs is common in many bacteria, archaea and eukaryotes, this phenomenon has not yet been observed in the α-proteobacterium Caulobacter crescentus. Based on a previous report that C. ... ...

Abstract	While translational read-through of stop codons by suppressor tRNAs is common in many bacteria, archaea and eukaryotes, this phenomenon has not yet been observed in the α-proteobacterium Caulobacter crescentus. Based on a previous report that C. crescentus and Escherichia coli tRNA(His) have distinctive identity elements, we constructed E. coli tRNA(His) CUA, a UAG suppressor tRNA for C. crescentus. By examining the expression of three UAG codon- containing reporter genes (encoding a β-lactamase, the fluorescent mCherry protein, or the C. crescentus xylonate dehydratase), we demonstrated that the E. coli histidyl-tRNA synthetase/tRNA(His) CUA pair enables in vivo UAG suppression in C. crescentus. E. coli histidyl-tRNA synthetase (HisRS) or tRNA(His) CUA alone did not achieve suppression; this indicates that the E. coli HisRS/tRNA(His) CUA pair is orthogonal in C. crescentus. These results illustrate that UAG suppression can be achieved in C. crescentus with an orthogonal aminoacyl-tRNA synthetase/suppressor tRNA pair.
MeSH term(s)	Ampicillin Resistance/genetics ; Caulobacter crescentus/genetics ; Caulobacter crescentus/metabolism ; Codon, Terminator ; Escherichia coli/genetics ; Escherichia coli/metabolism ; Gene Expression ; Gene Expression Regulation, Bacterial ; Histidine-tRNA Ligase/metabolism ; RNA, Transfer, His/genetics ; RNA, Transfer, His/metabolism
Chemical Substances	Codon, Terminator ; RNA, Transfer, His ; Histidine-tRNA Ligase (EC 6.1.1.21)
Language	English
Publishing date	2013-12-30
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't ; Research Support, U.S. Gov't, Non-P.H.S.
ISSN	1932-6203
ISSN (online)	1932-6203
DOI	10.1371/journal.pone.0083630
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Article ; Online: VLDLR and ApoER2 are receptors for multiple alphaviruses.

Clark, Lars E / Clark, Sarah A / Lin, ChieYu / Liu, Jianying / Coscia, Adrian / Nabel, Katherine G / Yang, Pan / Neel, Dylan V / Lee, Hyo / Brusic, Vesna / Stryapunina, Iryna / Plante, Kenneth S / Ahmed, Asim A / Catteruccia, Flaminia / Young-Pearse, Tracy L / Chiu, Isaac M / Llopis, Paula Montero / Weaver, Scott C / Abraham, Jonathan

Nature

2021 Volume 602, Issue 7897, Page(s) 475–480

Abstract: Alphaviruses, like many other arthropod-borne viruses, infect vertebrate species and insect vectors separated by hundreds of millions of years of evolutionary history. Entry into evolutionarily divergent host cells can be accomplished by recognition of ... ...

Abstract	Alphaviruses, like many other arthropod-borne viruses, infect vertebrate species and insect vectors separated by hundreds of millions of years of evolutionary history. Entry into evolutionarily divergent host cells can be accomplished by recognition of different cellular receptors in different species, or by binding to receptors that are highly conserved across species. Although multiple alphavirus receptors have been described
MeSH term(s)	Animals ; LDL-Receptor Related Proteins ; Ligands ; Mice ; Mosquito Vectors ; Receptors, LDL ; Semliki forest virus/metabolism ; Sindbis Virus/physiology
Chemical Substances	LDL-Receptor Related Proteins ; Ligands ; Receptors, LDL ; VLDL receptor ; low density lipoprotein receptor-related protein 8
Language	English
Publishing date	2021-12-20
Publishing country	England
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
ZDB-ID	120714-3
ISSN	1476-4687 ; 0028-0836
ISSN (online)	1476-4687
ISSN	0028-0836
DOI	10.1038/s41586-021-04326-0
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