Maiolo James R, Ferrer Marc, Ottinger Elizabeth A
Department of Chemistry and Biochemistry, Swarthmore College, 500 College Avenue, Swarthmore, PA, 19081, USA.
Biochim Biophys Acta. 2005 Jun 30;1712(2):161-72. doi: 10.1016/j.bbamem.2005.04.010.
The identification of cell-penetrating peptides (CPPs) as vectors for the intracellular delivery of conjugated molecules such as peptides, proteins, and oligonucleotides has emerged as a significant tool to modulate biological activities inside cells. The mechanism of CPP uptake by the cells is still unclear, and appears to be both endocytotic and non-endocytotic, depending on the CPP and cell type. Moreover, it is also unknown whether cargo sequences have an effect on the uptake and cellular distribution properties of CPP sequences. Here, we combine results from quantitative fluorescence microscopy and binding to lipid membrane models to determine the effect of cargo peptide molecules on the cellular uptake and distribution of the arginine-rich CPPs, R7, and R7W, in live cells. Image analysis algorithms that quantify fluorescence were used to measure the relative amount of peptide taken up by the cell, as well as the extent to which the uptake was endocytotic in nature. The results presented here indicate that fusion of arginine-rich CPPs to peptide sequences reduces the efficiency of uptake, and dramatically changes the cellular distribution of the CPP from a diffuse pattern to one in which the peptides are mostly retained in endosomal compartments.
细胞穿透肽（CPPs）作为肽、蛋白质和寡核苷酸等共轭分子细胞内递送的载体，已成为调节细胞内生物活性的重要工具。细胞摄取CPPs的机制仍不清楚，似乎既有内吞作用又有非内吞作用，这取决于CPP和细胞类型。此外，货物序列是否对CPP序列的摄取和细胞分布特性有影响也尚不清楚。在这里，我们结合定量荧光显微镜和与脂质膜模型结合的结果，以确定货物肽分子对富含精氨酸的CPPs（R7和R7W）在活细胞中的细胞摄取和分布的影响。使用量化荧光的图像分析算法来测量细胞摄取的肽的相对量，以及摄取本质上是内吞作用的程度。此处给出的结果表明，富含精氨酸的CPPs与肽序列融合会降低摄取效率，并显著改变CPP的细胞分布，从弥散模式变为肽大多保留在内体区室的模式。
