Systems survey of endocytosis by multiparametric image analysis

Abstract

Endocytosis is a complex process fulfilling many cellular and developmental functions. Understanding how it is regulated and integrated with other cellular processes requires a comprehensive analysis of its molecular constituents and general design principles. Here, we developed a new strategy to phenotypically profile the human genome with respect to transferrin (TF) and epidermal growth factor (EGF) endocytosis by combining RNA interference, automated high-resolution confocal microscopy, quantitative multiparametric image analysis and high-performance computing. We identified several novel components of endocytic trafficking, including genes implicated in human diseases. We found that signalling pathways such as Wnt, integrin/cell adhesion, transforming growth factor (TGF)-beta and Notch regulate the endocytic system, and identified new genes involved in cargo sorting to a subset of signalling endosomes. A systems analysis by Bayesian networks further showed that the number, size, concentration of cargo and intracellular position of endosomes are not determined randomly but are subject to specific regulation, thus uncovering novel properties of the endocytic system.

Dopamine receptor D 3 regulates endocytic sorting by a Prazosin-sensitive interaction with the coatomer COPI

“…In untreated cells, both were trafficked through the endocytic pathway; however, EGF then moved to lysosomes, and transferrin moved to recycling endosomes and eventually, the plasma membrane as expected (14). We showed, in Fig.…”

A novel lentivirus for quantitative assessment of gene knockdown in stem cell differentiation

“…5 In addition to single gene knockdown, investigators developed RNAi libraries to discover gene pathways controlling various cellular processes and facilitate global understanding of cellular behavior. 6 --9 This approach was used to identify genes that are involved in proteasome-mediated proteolysis, 10 virus infection and replication, 11 --14 cytokinesis, 8,9,15,16 endocytosis, 17 stem cell self-renewal 18 or cancer cell proliferation and survival. 19,20 RNAi knockdown requires a good selection strategy to link genetic information to cellular phenotype.…”

Lipid-Mediated Endocytosis

“…It plays a central role in the life of eukaryotic cells by mediating important aspects in the communication between the cell and the outside world, and by serving as a sensitive regulator of cell dynamics and homeostasis (Collinet et al 2010). The endocytosed cargo includes nutrients, receptor-ligand complexes, lipids, antigens, membrane proteins, toxins, and pathogens, to name a few.…”