New genes and mechanisms establishing Left-Right Asymmetry in Drosophila – DRO-ASY

We propose a basic research project aiming at a better understanding of Left-Right (L/R) asymmetry establishment, using Drosophila as a model system. Breaking L/R symmetry in Bilateria embryos is a major event in body plan organization. During development, L/R asymmetry controls polarized morphogenesis leading to the asymmetric positioning of unique organs (heart, liver, stomach) and asymmetrical morphogenesis of bilateral ones (lungs, brain). In addition, L/R asymmetry controls the directional looping of tubular organs (heart tube, gut). Handedness is thus essential for the correct morphogenesis and function of visceral organs and of the brain. It is estimated that 1/5,000 - 1/10,000 persons suffer from L/R defects (situs inversus, heterotaxia, isomerism), which are responsible for a number of complex congenital heart defects (CHDs), misrotation of the intestine and spontaneous miscarriage. L/R asymmetry defects are also associated with polycystic renal disease and Kartagener syndrome.

Our laboratory initiated the study of L/R asymmetry in Drosophila, through the identification of the situs inversus gene myosin ID (myoID). MyoID is a dextral determinant controlling clockwise looping of L/R organs like the genitalia, gut and testis. In myoID mutant flies, organs are completely inverted and adopt sinistral (counter clockwise) orientation. Recently, genetic screening allowed the identification of the HOX gene Abdominal-B as a major gene controlling the expression of MyoID in the genitalia, as well as the activity of the sinistral pathway. Abd-B mutant genitalia do not rotate, indicating that Abd-B is an upstream transcription factor capable of controlling the transition from symmetry (no rotation) to asymmetry (dextral or sinistral rotation).

In this project, we propose to explore new important questions, through the following tasks:

Task 1 – Characterization of novel L/R pathway genes: genetic and two-hybrid screening allowed the identification of novel genes involved in Drosophila L/R development, namely the profilin gene (chickadee), V-ATPases and the Kinesin-II microtubule motor (Drosophila orthologs of KIF3A and KIF3B). Most Drosophila cells (except some neurons) do not have primary cilia therefore suggesting a non-ciliary function of Kinesin-II. This part may help drawing important parallels with vertebrates and characterize potential unifying/conserved mechanisms to establish L/R asymmetry.

Task 2 - Functional conservation of MyoID in vertebrates: work in collaboration (unpublished) showed that myoID morpholinos can lead to specific L/R defects in Xenopus, therefore suggesting a functional conservation of MyoID in vertebrates. In this part, we will further investigate the role of MyoID in zebrafish, potentially opening new avenues on the mechanisms determining L/R asymmetry in vertebrates.

Task 3 – Identification of sinistral pathway factors: our work revealed the existence of two parallel pathways, dextral (MyoID) and sinistral (unknown), which are both controlled by Abd-B. Based on our recent results and understanding, we propose to identify the putative factor(s) involved in sinistral determination by i) identifying the Abd-B target genes in the genitalia L/R organizer using molecular approaches (DAM-ID and TRAP methods ) and ii) through a specific genetic screen to identify sinistral activities, based on the rescue of the Abd-B loss-of-function phenotype.

This project should provide new information on i) the molecular nature and mechanism of dextral and sinistral development in Drosophila, ii) the functional conservation of myosin ID as a dextral determinant in vertebrates. In addition, studying the role of Kinesin-II and V-ATPase in Drosophila should provide an evolutionary perspective to the problem of L/R asymmetry determination and indicate whether or not some unifying mechanisms may exist in bilateria.