# Single-cell analysis of the early Drosophila salivary gland reveals that morphogenetic control involves both the induction and exclusion of gene expression programs

## TL;DR

This study uses single-cell RNA sequencing to show how gene expression changes control the early development of Drosophila salivary glands, including both activating and deactivating specific genes.

## Contribution

The study reveals that morphogenesis involves not only the induction but also the exclusion of gene expression programs, with Tollo playing a critical role.

## Key findings

- A set of genes is upregulated in placodal cells before morphogenetic changes occur.
- Certain genes, like hth, grainyhead, and tollo/toll-8, are selectively excluded from placodal cells during development.
- Maintaining tollo expression disrupts tube morphogenesis, suggesting its exclusion is necessary for proper development.

## Abstract

How tissue shape and therefore function is encoded by the genome remains in many cases unresolved. The tubes of the salivary glands in the Drosophila embryo start from simple epithelial placodes, specified through the homeotic factors Scr/Hth/Exd. Previous work indicated that early morphogenetic changes are prepatterned by transcriptional changes, but an exhaustive transcriptional blueprint driving physical changes was lacking. We performed single-cell-RNAseq-analysis of FACS-isolated early placodal cells, making up less than 0.4% of cells within the embryo. Differential expression analysis in comparison to epidermal cells analyzed in parallel generated a repertoire of genes highly upregulated within placodal cells prior to morphogenetic changes. Furthermore, clustering and pseudotime analysis of single-cell-sequencing data identified dynamic expression changes along the morphogenetic timeline. Our dataset provides a comprehensive resource for future studies of a simple but highly conserved morphogenetic process of tube morphogenesis. Unexpectedly, we identified a subset of genes that, although initially expressed in the very early placode, then became selectively excluded from the placode but not the surrounding epidermis, including hth, grainyhead and tollo/toll-8. We show that maintaining tollo expression severely compromised the tube morphogenesis. We propose tollo is switched off to not interfere with key Tolls/LRRs that are expressed and function in the tube morphogenesis.

How does transcriptional patterning regulate salivary gland morphogenesis? In this study, the authors provide a detailed scRNA-seq characterization of transcriptional changes during early morphogenesis of the Drosophila salivary gland placode, revealing that this process is regulated by both the induction, but also exclusion of regulatory factors, such as Tollo.