May Elizabeth J, Gaudet Rachelle
Department of Molecular and Cellular Biology, Harvard University, 52 Oxford Street, Cambridge, MA 02138, USA.
bioRxiv. 2025 Jun 6:2024.09.23.614616. doi: 10.1101/2024.09.23.614616.
Proteins that transmit molecules and signals across the plasma membrane are crucial in cell biology because they enable cells to sense and respond to their surroundings. A major challenge for studying cell-surface proteins is that often they do not fold or traffic properly to the plasma membrane when produced in heterologous cells. We developed a strategy for quantifying surface localization from fluorescence microscopy images of surface-stained cells. Using clustered protocadherins, a protein family important for cell-cell recognition during neuronal development, we found that surface delivery levels vary among clustered protocadherin isoforms and between wildtype and engineered variants. Quantifying these differences provides evidence that dimerization is not tightly coupled to surface delivery for clustered protocadherins. This work establishes a generalizable framework for screening proteins and variants of interest for proper cell surface localization.
能够在质膜上传递分子和信号的蛋白质在细胞生物学中至关重要，因为它们使细胞能够感知周围环境并做出反应。研究细胞表面蛋白面临的一个主要挑战是，当在异源细胞中产生时，它们往往不能正确折叠或运输到质膜。我们开发了一种从表面染色细胞的荧光显微镜图像中量化表面定位的策略。使用成簇原钙黏蛋白（一种在神经元发育过程中对细胞间识别很重要的蛋白质家族），我们发现表面递送水平在成簇原钙黏蛋白异构体之间以及野生型和工程变体之间存在差异。对这些差异进行量化提供了证据，表明二聚化与成簇原钙黏蛋白的表面递送没有紧密联系。这项工作建立了一个可推广的框架，用于筛选感兴趣的蛋白质及其变体以实现正确的细胞表面定位。
