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Probing cytoskeletal modulation of passive and active intracellular dynamics using nanobody-functionalized quantum dots.

ABSTRACT: The cytoplasm is a highly complex and heterogeneous medium that is structured by the cytoskeleton. How local transport depends on the heterogeneous organization and dynamics of F-actin and microtubules is poorly understood. Here we use a novel delivery and functionalization strategy to utilize quantum dots (QDs) as probes for active and passive intracellular transport. Rapid imaging of non-functionalized QDs reveals two populations with a 100-fold difference in diffusion constant, with the faster fraction increasing upon actin depolymerization. When nanobody-functionalized QDs are targeted to different kinesin motor proteins, their trajectories do not display strong actin-induced transverse displacements, as suggested previously. Only kinesin-1 displays subtle directional fluctuations, because the subset of microtubules used by this motor undergoes prominent undulations. Using actin-targeting agents reveals that F-actin suppresses most microtubule shape remodelling, rather than promoting it. These results demonstrate how the spatial heterogeneity of the cytoskeleton imposes large variations in non-equilibrium intracellular dynamics.

SUBMITTER: Katrukha EA

PROVIDER: S-EPMC5364406 | biostudies-literature | 2017 Mar

REPOSITORIES: biostudies-literature

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Probing cytoskeletal modulation of passive and active intracellular dynamics using nanobody-functionalized quantum dots.

Katrukha Eugene A EA Mikhaylova Marina M van Brakel Hugo X HX van Bergen En Henegouwen Paul M PM Akhmanova Anna A Hoogenraad Casper C CC Kapitein Lukas C LC

Nature communications 20170321

The cytoplasm is a highly complex and heterogeneous medium that is structured by the cytoskeleton. How local transport depends on the heterogeneous organization and dynamics of F-actin and microtubules is poorly understood. Here we use a novel delivery and functionalization strategy to utilize quantum dots (QDs) as probes for active and passive intracellular transport. Rapid imaging of non-functionalized QDs reveals two populations with a 100-fold difference in diffusion constant, with the faste ...[more]

PMID: 28322225

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Project description:Nanotechnology is a growing industry with wide ranging applications in consumer product and technology development. In the biomedical field, nanoparticles are finding increasing use as imaging agents for biomolecular labeling and tumor targeting. The nanoparticle physiochemical properties must be tailored for the specific application. For example, nanoparticle chemical and physical stability in the biological milieu (no oxidation, aggregation, agglomeration or toxicity) are often required. Nanoparticles used for biomolecular fluorescent imaging should also have high quantum yield (QY). The aim of this paper is to examine the QY, stability, and cell toxicity of a series of positive, negative and neutral surface charge quantum dot (QD) nanoparticles. Simple protocols are described to prepare water soluble QDs by modifying the surface with thiol containing antioxidant ligands and polymers keeping the QD core/shell composition constant. The ligands used to produce negatively charged QDs include glutathione (GSH), N-acetyl-L-cysteine (NAC), dihydrolipoic acid (DHLA), tiopronin (TP), bucilliamine (BUC), and mercaptosuccinic acid (MSA). Ligands used to produce positively charged QDs include cysteamine (CYS) and polyethylenimine (PEI). Dithiothreitol (DTT) was used to produce neutral charged QDs. Commercially available nonaqueous octadecylamine (ODA) capped QDs served as the starting material. Our results suggest that QD uptake and cytotoxicity are both dependent on surface ligand coating composition. The negative charged GSH coated QDs show superior performance exhibiting low cytotoxicity, high stability, high QY and therefore are best suited for bioimaging applications. PEI coated QD also show superior performance exhibiting high QY and stability. However, they are considerably more cytotoxic due to their high positive charge which is an advantageous property that can be exploited for gene transfection and/or tumor targeting applications. The synthetic procedures described are straightforward and can be easily adapted in most laboratory settings.

Dataset Information

Probing cytoskeletal modulation of passive and active intracellular dynamics using nanobody-functionalized quantum dots.

Publications

Probing cytoskeletal modulation of passive and active intracellular dynamics using nanobody-functionalized quantum dots.

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