Deciphering the nanomaterial uptake in a bystander manner invoked by co-administration of functionalized nanoparticles

摘要 纳米材料应用中一个重要但研究不足的方面是纳米材料的细胞生物学。 通过细胞和组织进行运输。以克服细胞膜并到达它们的作用部位 在体内，纳米颗粒(NPs)通常与穿透细胞的配体形成络合物。这类配体包括多肽 比如TAT(反式激活转录)。这些配体与细胞表面受体结合，以激活 NP摄取的内吞过程。然而，这些功能化的NP的影响以及由此产生的 内吞过程对细胞摄取其他NPs的影响尚不清楚。在这项建议中，我的目标是解决这个问题 通过研究TAT偶联NP(TAT-NP)来解决这个问题。 通过追踪包裹着几种细胞穿透肽的NPs，包括Tat，我们之前发现了一种 新的受体依赖的巨噬细胞吞噬(MP)途径摄取NP。MP有相对较大的内吞 空泡(直径200 nm)，因此可以更容易地吞噬纳米颗粒大小的货物。在这里，我们 发现TAT-NPs通过调用该MP途径可以将无法进入细胞的旁观者NPs带入细胞 细胞本身。这种旁观者的摄取依赖于TAT与其受体硫酸乙酰肝素的相互作用 (HS)蛋白多糖，通过HS依赖的MP途径发生。我们进一步表明，它只是 对于NP类型的旁观者货物是活性的，其活性受到细胞外半胱氨酸(Cys)的极大刺激。 在这里，我们的目标是剖析这个旁观者的理解过程。在目标1中，我们将确定先决条件 引发了旁观者的兴趣。在目标2和目标3中，我们将利用遗传筛选和其他方法来 确定调节这种旁观者吸收的因素，并确定其货物选择性和 Cys规则。在目标4中，我们将探索其在生物医学应用中的潜在用途。我的建议 研究具有巨大的潜力，可以揭示纳米材料运输的重要细胞机制，促进 细胞内递送的效率，并开辟了研究细胞生物学、新陈代谢和 纳米材料的输送。

Abstract One important yet understudied aspect in nanomaterial applications is the cell biology of nanomaterial transport into cells and through tissues. To overcome the cell membranes and reach their sites of action within, nanoparticles (NPs) are often complexed with cell-penetrating ligands. Such ligands include peptides like TAT (Trans-Activating Transcription). These ligands engage with cell surface receptors to invoke endocytic processes for NP uptake. However, the impact of these functionalized NPs and the resulting endocytic process on the cellular uptake of other NPs remains unknown. In this proposal, I aim to tackle this problem by studying TAT-coupled NPs (TAT-NP). By tracing NPs coated with several cell-penetrating peptides, including TAT, we previously discovered a novel receptor-dependent macropinocytosis (MP) pathway for NP uptake. MP has relatively large endocytic vacuoles (>200 nm in diameter) and thus can more readily engulf cargo as large as nanoparticles. Here, we found that TAT-NPs, by invoke this MP pathway, can bring into cells bystander NPs that are unable to enter the cells by themselves. This bystander uptake depends on TAT interaction with its receptor, heparan sulfate (HS) proteoglycans, and occurs through the HS-dependent MP pathway. We further showed that it is only active for NP-type bystander cargo, and its activity is greatly stimulated by Cysteine (Cys) outside the cells. Here, we aim to dissect out this bystander uptake process. In the Aim 1, we will determine the prerequisites for initiating the bystander uptake. In the Aim 2 and 3, we will utilize genetic screening and other methods to determine the factors that mediate this bystander uptake, and define its properties of cargo selectivity and Cys regulation. In the Aim 4, we will explore the potential use in biomedical applications. My proposed studies hold great potential for unveiling important cellular machineries for nanomaterial transport, boosting the efficiency of intracellular delivery, and opening up a new avenue to study cell biology, metabolism and nanomaterial delivery.