Exploring endocytic compartment morphology with systematic genetics and single cell image analysis

Mojca Mattiazzi Usaj1,#, Nil Sahin1,2,#, Helena Friesen1, Carles Pons3, Matej Usaj1, Myra Paz Masinas1, Ermira Shuteriqi1, Aleksei Shkurin1,2, Patrick Aloy3,4, Quaid Morris1,2,5,6,7*, Charles Boone1,2,*, and Brenda J. Andrews1,2,8,*

  • 1The Donnelly Centre, University of Toronto, Toronto, Canada
  • 2Department of Molecular Genetics, University of Toronto, Toronto, Canada
  • 3Institute for Research in Biomedicine (IRB Barcelona), the Barcelona Institute for Science and Technology, Barcelona, Catalonia, Spain
  • 4Instituci├│ Catalana de Recerca i Estudis Avan├žats (ICREA), Barcelona, Catalonia, Spain
  • 5Department of Computer Science, University of Toronto, Toronto, Canada
  • 6Vector Institute, Toronto, Canada
  • 7Ontario Institute of Cancer Research, Toronto, Canada
  • 8Lead Contact
  • # These authors contributed equally to this work
  • * Correspondence: brenda.andrews@utoronto.ca, charlie.boone@utoronto.ca, quaid.morris@utoronto.ca

Summary

Endocytosis is a conserved process that mediates the internalization of nutrients and plasma membrane components, including receptors, for sorting to endosomes and the vacuole (lysosome). We combined systematic yeast genetics, high-content screening, and neural network-based image analysis of single cells to screen for genes that influence the morphology of four main endocytic compartments: coat proteins, actin patches, late endosome, and vacuole. This unbiased approach identified 17 mutant phenotypes and ~1600 genes whose perturbation affected at least one of the four compartments. Numerous mutants were associated with multiple phenotypes, indicating that morphological pleiotropy is often seen within the endocytic pathway. Morphological profiles based on the 17 aberrant phenotypes were highly correlated for functionally related genes, enabling prediction of gene function. Incomplete penetrance was prevalent, and single-cell analysis enabled exploration of the mechanisms underlying cellular heterogeneity, which include replicative age, organelle inheritance, and stress response.