Integrating growth and architecture: Cell cycle expression of polarity genes

Sandhya Payankaulam , Stephanie L. Hickey, and David N. Arnosti

This repository contains the results generated in this study as well as the code necessary for reproducing all of the analyses in figure 1 and the supplemental figures.

Data

The WT Drosophila Wing imaginal disc 10x scRNA-seq data generated and described in Deng et. al. 2019 can be fond here. The early Drosophila embryo Drop-seq data generated and described in Karaiskos et. al. 2017 can be found here.

Downloading raw data

The specific data used in this study can be downloaded from GEO by running:

$ sh ./run/download_data.sh

download_data.sh creates a Raw_data to contain the downloaded files.

Analysis

The following analyses were completed using R version 4.0.0. To install the necessary packages install R and run:

CAUTION this will install Seurat V3.2.3, replacing any other version of Seurat in your R library $ Rscript ./run/install_packages.R

Preprocessing

The data was preprocessed and normalized using:

$ Rscript ./run/preprocess.R

preprocess.R creates a normalized_data directory which contains R objects with normalized gene expression matrices.

Phase assignment

Each cell was assigned a putative cell-cycle phase label using Seurat V3's CellCycleScoring function. We used a custom list of cell-cycle phase marker genes, and the citations linking these genes to specific cell-cycle phases can be found in supplemental table 1 included with the manuscript. To recreated our labels run:

$ Rscript ./run/phase_assignment.R

phase_assignment.R creates a results folder and outputs:

• results/20200922_genes_of_interest_detected_in_dataset.csv, a table that lists the labels for polarity and cell-cycle genes and whether or not they are expressed in a particular single-cell data set.
• Seurat objects that contain the phase assignments for each cell in the @meta.data slot

Figures and Tables

All of the tables and figures from this manuscript are available in results and can be recreated by running:

$ Rscript ./run/figure1_plots.R

• figure1_plots.R outputs the contents of results/figure1
• these plots were used in figure 1 (wing disc data) and figure S5 (embryo data)

and

$ Rscript ./run/suplemental_figures.R

suplemental_figures.R outputs the contents of results/supplemental

• *_fisher_pval.csv and *_fisher_FDR.csv list the p-values or FDRs associated with the stacked bar plots (Figures 1C and E, S1, S3, S5C and E, S6, S8)
• *_kw_dunns.csv lists the p-values and FDRs associated with the expression vs. phase dot plots and heatmaps (Figures 1B, D and F, S2, S4, S5B, D and F, S7, S9)