Dataset Information

A systematic High-Content Screening microscopy approach reveals key roles for Rab33b, OATL1 and Myo6 in nanoparticle trafficking in HeLa cells.

ABSTRACT: Synthetic nanoparticles are promising tools for imaging and drug delivery; however the molecular details of cellular internalization and trafficking await full characterization. Current knowledge suggests that following endocytosis most nanoparticles pass from endosomes to lysosomes. In order to design effective drug delivery strategies that can use the endocytic pathway, or by-pass lysosomal accumulation, a comprehensive understanding of nanoparticle uptake and trafficking mechanisms is therefore fundamental. Here we describe and apply an RNA interference-based high-content screening microscopy strategy to assess the intracellular trafficking of fluorescently-labeled polystyrene nanoparticles in HeLa cells. We screened a total of 408 genes involved in cytoskeleton and membrane function, revealing roles for myosin VI, Rab33b and OATL1 in this process. This work provides the first systematic large-scale quantitative assessment of the proteins responsible for nanoparticle trafficking in cells, paving the way for subsequent genome-wide studies.

SUBMITTER: Panarella A

PROVIDER: S-EPMC4931513 | biostudies-literature | 2016 Jul

REPOSITORIES: biostudies-literature

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A systematic High-Content Screening microscopy approach reveals key roles for Rab33b, OATL1 and Myo6 in nanoparticle trafficking in HeLa cells.

Panarella Angela A Bexiga Mariana G MG Galea George G O' Neill Elaine D ED Salvati Anna A Dawson Kenneth A KA Simpson Jeremy C JC

Scientific reports 20160704

Synthetic nanoparticles are promising tools for imaging and drug delivery; however the molecular details of cellular internalization and trafficking await full characterization. Current knowledge suggests that following endocytosis most nanoparticles pass from endosomes to lysosomes. In order to design effective drug delivery strategies that can use the endocytic pathway, or by-pass lysosomal accumulation, a comprehensive understanding of nanoparticle uptake and trafficking mechanisms is therefo ...[more]

PMID: 27374232

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Project description:How cells control their shape and size is a fundamental question of biology. In most bacteria, cell shape is imposed by the peptidoglycan (PG) polymeric meshwork that surrounds the cell. Thus, bacterial cell morphogenesis results from the coordinated action of the proteins assembling and degrading the PG shell. Remarkably, during steady-state growth, most bacteria maintain a defined shape along generations, suggesting that error-proof mechanisms tightly control the process. In the rod-shaped model for the Gram-positive bacterium Bacillus subtilis, the average cell length varies as a function of the growth rate, but the cell diameter remains constant throughout the cell cycle and across growth conditions. Here, in an attempt to shed light on the cellular circuits controlling bacterial cell width, we developed a screen to identify genetic determinants of cell width in B. subtilis. Using high-content screening (HCS) fluorescence microscopy and semiautomated measurement of single-cell dimensions, we screened a library of ∼4,000 single knockout mutants. We identified 13 mutations significantly altering cell diameter, in genes that belong to several functional groups. In particular, our results indicate that metabolism plays a major role in cell width control in B. subtilis. IMPORTANCE Bacterial shape is primarily dictated by the external cell wall, a vital structure that, as such, is the target of countless antibiotics. Our understanding of how bacteria synthesize and maintain this structure is therefore a cardinal question for both basic and applied research. Bacteria usually multiply from generation to generation while maintaining their progenies with rigorously identical shapes. This implies that the bacterial cells constantly monitor and maintain a set of parameters to ensure this perpetuation. Here, our study uses a large-scale microscopy approach to identify at the whole-genome level, in a model bacterium, the genes involved in the control of one of the most tightly controlled cellular parameters, the cell width.

Dataset Information

A systematic High-Content Screening microscopy approach reveals key roles for Rab33b, OATL1 and Myo6 in nanoparticle trafficking in HeLa cells.

Publications

A systematic High-Content Screening microscopy approach reveals key roles for Rab33b, OATL1 and Myo6 in nanoparticle trafficking in HeLa cells.

Similar Datasets

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