Inferring signalling networks from images

Abstract

The mapping of signalling networks is one of biology's most important goals. However, given their size, complexity and dynamic nature, obtaining comprehensive descriptions of these networks has proven extremely challenging. A fast and cost-effective means to infer connectivity between genes on a systems-level is by quantifying the similarity between high-dimensional cellular phenotypes following systematic gene depletion. This review describes the methodology used to map signalling networks using data generated in the context of RNAi screens.

Keywords: Heterogeneity; RNAi; image analysis; morphological signatures; signalling networks.

Figure 1

Depleting different members of the same signalling complex leads to similar cellular phenotypes. (A) Kc167 Drosophila cells treated with dsRNA targetting Rho1, Pbl, RacGAP50C or Pavarotti, have similar cellular phenotypes. Cells were treated with RNAi and then fixed, stained with DAPI (red), phalloidin (blue), and anti-alpha tubulin antibody (green), and then imaged using an Opera QEHS microscope (PerkinElmer). The appearance of large, multinucleated, round cells suggests cytokinesis, but not mitosis, has failed during cell division. Scale bars are equal to 20 μm. (B) Rho1, Pbl, RacGAP50C and Pavarotti act as part of the same signalling complex during cytokinesis. All proteins localize to the presumptive cleavage furrow and promote actomyosin contractile ring assembly. Red arrows indicate there is evidence for active regulation of one protein by another, such activation/inhibition or control of localization.

Figure 2

Texture features are useful descriptors of cellular phenotypes. (A) Untreated (left panels) or Taxol treated MCF-10A cells (right panels) were stained with DAPI (red) and antialpha tubulin antibody (yellow). Cells were imaged on an Opera QEHS microscope. ‘SER Edge’ filtered image generated in Columbus (PerkinElmer). (B) The SER Edge feature is calculated on a single cell basis as the pixel intensity within a single object (cell segment) averaged over the corresponding object. Scales bar are equal to 20 μm.