Cellular uptake of a cationic amphiphilic fluorophore in the form of assemblies via Clathrin-dependent endocytosis

Abstract Cellular uptake is a key step for the organic fluorophore molecules to label eukaryotic cells. Small amphiphilic fluorophores in aqueous solution present a rather complicated system, where individual molecules (monomers) or nano-scale assemblies co-exist in the solution. How and in which form do they enter into cells, and how they further gather in cells? These questions are still remain unknown to the best of our knowledge. In order to unravel the above problems, we herein studied the cellular uptake of a cationic amphiphile TPE-11, which shows strong aggregation induced emission effects in aqueous solution. Significant differences of fluorescent signals were obtained when concentration of TPE-11 applied to label Hela cells was below and above its critical micellar concentration (CMC). To elucidate the pathways for assembly forms entering into inner cells, we screened candidate proteins involving in internalization of TPE-11 in HeLa cells by using proteomics analysis, liquid chromatography plus mass spectrometry. The results indicate that TPE-11 mainly takes form of assemblies to enter into cells via Clathrin-dependent endocytosis (CDE). The result was confirmed by the specific inhibitor of the CDE pathway and knockdown of Clathrin with shRNA interfering. The cellular uptake demonstrated by TPE-11 might also apply to analogous cationic amphiphilic molecules.