A hierarchical map of regulatory genetic interactions in membrane trafficking.

Endocytosis is critical for cellular physiology and thus is highly regulated. To identify regulatory interactions controlling the endocytic membrane system, we conducted 13 RNAi screens on multiple endocytic activities and their downstream organelles. Combined with image analysis of thousands of single cells per perturbation and their cell-to-cell variability, this created a high-quality and cross-comparable quantitative data set. Unbiased analysis revealed emergent properties of the endocytic membrane system and how its complexity evolved and distinct programs of regulatory control that coregulate specific subsets of endocytic uptake routes and organelle abundances. We show that these subset effects allow the mapping of functional regulatory interactions and their interaction motifs between kinases, membrane-trafficking machinery, and the cytoskeleton at a large scale, some of which we further characterize. Our work presents a powerful approach to identify regulatory interactions in complex cellular systems from parallel single-gene or double-gene perturbation screens in human cells and yeast.