---
title: "CellphoneDB
author: "Alex Germanos, Sonali Arora"
date: "Feb 24, 2022"
output: 
  html_document:
    toc: true
    theme: united
---



## Introduction

In this vignette, we analyze our data with cellphoneDb and then make figures to visualize the interactions.

##  cellphoneDb on our scRNASeq data
First we build the docker image of cellphonedb

```{}
singularity build mycellphonedb.sif docker://ydevs/cellphonedb:2.1

```
Next, we prepare our data in a format that is acceptable by cellphoneDb

```{r}

library(biomaRt)
human = useMart("ensembl", dataset = "hsapiens_gene_ensembl")
mouse = useMart("ensembl", dataset = "mmusculus_gene_ensembl")

library(Seurat)
seurat = readRDS("data/seurat_cleaned.allcells.rds")
scale_data = seurat@assays$RNA@scale.data
meta = cbind(cell = rownames(seurat@meta.data), cell_type=seurat@meta.data$cell_types)

# Basic function to convert human to mouse gene names
convert_Mouse_to_Human <- function(genes){
  df =  getLDS(attributes = c("mgi_symbol"), filters = "mgi_symbol",
               values = genes ,mart = mouse,
               attributesL = c("hgnc_symbol"), martL = human, uniqueRows=T)
  df
}
gene_df = convert_Mouse_to_Human(rownames(scale_data))

# next keep only unique genes.
goi = unique(gene_df[,2])
keep_idx2 = match(goi, gene_df[,2])

scale_data = scale_data[keep_idx2, ]
gene_df = gene_df[keep_idx2, ]

rownames(scale_data) = gene_df[, "HGNC.symbol"]

want_cells = c("Basal", "Progenitor", "Differentiated", "Macrophages", "Dendritic cells", "Neutrophils", "T-cells")
want_idx = which(meta_all$cell_type %in% want_cells)

scale_data = scale_data[, want_idx]
# make one for each genotype.

test_df = seurat@meta.data
test_df = test_df[which(test_df$cell_types %in% want_cells), ]

wt_intact_idx= rownames(test_df[which(test_df$genotype=="WT_intact"), ])
pten_intact_idx= rownames(test_df[which(test_df$genotype=="PTEN_intact"), ])
pten_cx_idx= rownames(test_df[which(test_df$genotype=="PTEN_castrate"), ])
pten_cx_4ebp1_idx= rownames(test_df[which(test_df$genotype=="PTEN_castrate_4EBP1"), ])

wt_intact = scale_data[, match(wt_intact_idx, colnames(scale_data))]
pten_intact = scale_data[, match(pten_intact_idx, colnames(scale_data))]
pten_cx = scale_data[, match(pten_cx_idx, colnames(scale_data))]
pten_cx_4ebp1 = scale_data[, match(pten_cx_4ebp1_idx, colnames(scale_data))]

meta_wt_intact = meta[ match(wt_intact_idx, meta[,1]), ]
meta_pten_intact = meta[ match(pten_intact_idx, meta[,1]), ]
meta_pten_cx = meta[ match(pten_cx_idx, meta[,1]), ]
meta_pten_cx_4ebp1 = meta[ match(pten_cx_4ebp1_idx, meta[,1]), ]

write.table(wt_intact, "~/hsiehlab/Sonali/tools/cellphoneDB/scale_counts_wt_intact.txt", 
            sep ="\t", quote=FALSE, row.names=TRUE, col.names=TRUE)
write.table(pten_intact, "~/hsiehlab/Sonali/tools/cellphoneDB/scale_counts_pten_intact.txt", 
            sep ="\t", quote=FALSE, row.names=TRUE, col.names=TRUE)
write.table(pten_cx, "~/hsiehlab/Sonali/tools/cellphoneDB/scale_counts_pten_cx.txt", 
            sep ="\t", quote=FALSE, row.names=TRUE, col.names=TRUE)
write.table(pten_cx_4ebp1, "~/hsiehlab/Sonali/tools/cellphoneDB/scale_counts_pten_cx_4ebp1.txt", 
            sep ="\t", quote=FALSE, row.names=TRUE, col.names=TRUE)

write.table(meta_wt_intact, "~/hsiehlab/Sonali/tools/cellphoneDB/meta_wt_intact.txt", 
            sep ="\t", quote=FALSE, row.names=FALSE, col.names=TRUE)
write.table(meta_pten_intact, "~/hsiehlab/Sonali/tools/cellphoneDB/meta_pten_intact.txt", 
            sep ="\t", quote=FALSE, row.names=FALSE, col.names=TRUE)
write.table(meta_pten_cx, "~/hsiehlab/Sonali/tools/cellphoneDB/meta_pten_cx.txt", 
            sep ="\t", quote=FALSE, row.names=FALSE, col.names=TRUE)
write.table(meta_pten_cx_4ebp1, "~/hsiehlab/Sonali/tools/cellphoneDB/meta_pten_cx_4ebp1.txt", 
            sep ="\t", quote=FALSE, row.names=FALSE, col.names=TRUE)
```

Finally, we run cellphoneDb on each genotype. 


```{}
 singularity run --bind ${PWD}:/DATA -W /DATA mycellphonedb.sif cellphonedb method statistical_analysis meta_wt_intact.txt scale_counts_wt_intact.txt --counts-data hgnc_symbol --project-name=wt_intact --threads 8

 singularity run --bind ${PWD}:/DATA -W /DATA mycellphonedb.sif cellphonedb method statistical_analysis meta_pten_intact.txt scale_counts_pten_intact.txt --counts-data hgnc_symbol --project-name=pten_intact --threads 8

 singularity run --bind ${PWD}:/DATA -W /DATA mycellphonedb.sif cellphonedb method statistical_analysis meta_pten_cx.txt scale_counts_pten_cx.txt --counts-data hgnc_symbol --project-name=pten_cx --threads 8

 singularity run --bind ${PWD}:/DATA -W /DATA mycellphonedb.sif cellphonedb method statistical_analysis meta_pten_cx_4ebp1.txt scale_counts_pten_cx_4ebp1.txt --counts-data hgnc_symbol --project-name=pten_cx_4ebp1 --threads 8

```