Abstract
Cell-penetrating peptides (CPPs) can translocate across cell membranes, and thus have great potential for the cellular delivery of macromolecular cargoes. However, the mechanism of this cellular uptake process is not yet fully understood. In this study, a time-lapse single-particle light-sheet microscopy technique was implemented to obtain a parallel visualization of the translocating process of individual human immunodeficiency virus 1 (HIV-1) transactivator of transcription (Tat) peptide conjugated quantum dots (TatP-QDs) in complex cellular terrains. Here, TatP-QDs served as nanoscale dynamic pens, which depict remarkable trajectory aggregates of TatP-QDs on the cell surface. Spectral-embedding analysis of the trajectory aggregates revealed a manifold formed by isotropic diffusion and a fraction of directed movement, possibly caused by interaction between the Tat peptides and heparan sulfate groups on the plasma membrane. Further analysis indicated that the membrane deformation induced by Tat-peptide attachment increased with the disruption of the actin framework in cytochalasin D (cyto D)-treated cells, yielding higher interactions on the TatP-QDs. In native cells, the Tat peptides can remodel the actin framework to reduce their interaction with the local membrane environment. Characteristic hot spots for interaction were detected on the membrane, suggesting that a funnel passage may have formed for the Tat-coated particles. This finding offers valuable insight into the cellular delivery of nanoscale cargo, suggesting an avenue for direct therapeutic delivery.
Highlights
Numerous barriers can be encountered during the delivery of drugs [1], which seriously reduce the efficacy of drugs and increase their off-target toxicity
We developed a single-particle light-sheet microscopy to capture the translocation of Tat peptide (TatP)-quantum dots (QDs) across the plasma membranes of living cells, which was found to occur preferentially at selected plasma-membrane regions
We prepared a cell-culture medium containing 5 μM TatP and 5 nM TatP-QDs to study the initial rates of cell-penetrating peptides (CPPs) uptake at room temperature
Summary
Numerous barriers can be encountered during the delivery of drugs [1], which seriously reduce the efficacy of drugs and increase their off-target toxicity. These barriers include filtration by the kidney and premature clearance by the reticulo-endothelial system. Upon arriving at target tissues, drug molecules must further cross plasma membranes to reach the sites of action. It is desirable to render drug molecules for crossing cellular membranes directly and thereby avoids the complications of vesicle-mediated internalization pathways. CPP-based membrane-translocation strategies have attracted considerable attention, as they can offer a high-delivery yield, low toxicity, and the possibility of tailoring their functions [3].
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