FijiWingsPolarity: An open source toolkit for semi-automated detection of cell polarity

Abstract

Epithelial cells are defined by apical-basal and planar cell polarity (PCP) signaling, the latter of which establishes an orthogonal plane of polarity in the epithelial sheet. PCP signaling is required for normal cell migration, differentiation, stem cell generation and tissue repair, and defects in PCP have been associated with developmental abnormalities, neuropathologies and cancers. While the molecular mechanism of PCP is incompletely understood, the deepest insights have come from Drosophila, where PCP is manifest in hairs and bristles across the adult cuticle and organization of the ommatidia in the eye. Fly wing cells are marked by actin-rich trichome structures produced at the distal edge of each cell in the developing wing epithelium and in a mature wing the trichomes orient collectively in the distal direction. Genetic screens have identified key PCP signaling pathway components that disrupt trichome orientation, which has been measured manually in a tedious and error prone process. Here we describe a set of image processing and pattern-recognition macros that can quantify trichome arrangements in micrographs and mark these directly by color, arrow or colored arrow to indicate trichome location, length and orientation. Nearest neighbor calculations are made to exploit local differences in orientation to better and more reliably detect and highlight local defects in trichome polarity. We demonstrate the use of these tools on trichomes in adult wing preps and on actin-rich developing trichomes in pupal wing epithelia stained with phalloidin. FijiWingsPolarity is freely available and will be of interest to a broad community of fly geneticists studying the effect of gene function on PCP.

A Van Gogh/Vangl tyrosine phosphorylation switch regulates its interaction with core Planar Cell Polarity factors Prickle and Dishevelled

“…The point mutations that lead to loss of binding to a single effector (Y374F only binding Dsh, and V376A only binding to Pk) showed a weaker phenotype as compared to Y374A (Figure 4C-D). This was further supported through quantification of cellular hair orientation, using the FijiWingsPolarity plug-in (Dobens et al, 2018), that revealed a significant variation from the Vang WT control in each case, and also between Vang-Y374A and the other point mutations (Figure 4C-D). Importantly, the difference in phenotype was not due to variation in expression levels, as all genotypes showed equivalent protein levels in larval disc lysates (Figure 4E).…”

“…( D ) Quantification of the percentage of cellular actin hairs oriented within a specific angle range for the indicated genotypes. Angles were determined using plugin FijiWingsPolarity (Dobens et al 2018). Cellular hair orientation angles from 3 independent wings were combined, and data binned for statistical analysis via Chi-squared test.…”

“…Adult wings were collected in PBS containing 0.1% Triton-X-100 (PBST) and incubated for 1 hr at room temperature before mounting in 80% glycerol in PBS. Hair orientation was quantified using the FijiWingsPolarity plugin (Dobens et al, 2018). The quantification was performed on equivalent regions of 3 wings per genotype, and angles binned into 30° degree segments to allow for statistical analysis via a Chi-squared test.…”

PCP Auto Count: A Novel Fiji/ImageJ plug-in for automated quantification of planar cell polarity and cell counting

“…Some are only executable on certain operating systems (e.g. FijiWingsPolarity [40], which works with Mac OS, but not PC). Of those that remain, the underlying approaches to measuring cell orientation differ from PCPA.…”

Tissue polarity and PCP protein function: C. elegans as an emerging model

“…In the fly, the cells of various body parts and appendages are polarized in the proximal-distal direction. This polarity is easily recognized by the orientation of actinbased hairs, formed at the posterior of individual epithelial cells of the wing, notum, and legs [4]. Two main PCP pathways organize proximal-distal polarity in various tissues.…”