Live-imaging of adhesion-dependent axonal branch competition in the Drosophila brain

• Applicant: Dr. Jakob Rentsch
• Subject Area: Developmental Neurobiology ; Molecular Biology and Physiology of Neurons and Glial Cells
• Term: since 2026
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 577138588

The formation of synapses is of fundamental importance for brain development. However, many steps occur before synaptogenesis in a highly specific order. This is crucial, as many neurons taken out of their developmental context have been shown to form synapses with other neurons, including themselves. Developmental steps prior to synapse formation include the growth of axons and dendrites, often in elaborate patterns and with major effects on neuronal interactions and thus availability as potential synaptic partners. Such transient neuronal interactions can contribute significantly to connectivity, even though no trace of these events may be left in the adult. Hence, to understand how a neuron chooses a specific synaptic partner, the underlying developmental processes should be investigated based on live dynamics at the time when they occur, rather than only in the adult and in fixed preparations. The brain of the fly Drosophila melanogaster is an excellent model system to study such processes, as the entire intact fly brain can be cultured and simultaneously live-observed during all relevant stages of development of brain wiring. The core idea of my proposal is to live-observe dynamic neuronal interactions via FRET technology in the developing fly brain. I show that this technology works in principle in the chosen neurons of the fly visual system (L4 interneurons) and that all methods are in place to perform the experiments. I suggest an improved version for sparse genetic labeling of interacting neurons, which is generally useful for Drosophila researchers (Objective 1). Moreover, I propose to employ cell-contact FRET to investigate the function of the immunoglobulin superfamily protein rst. Rst has been suggested to be an axon guidance molecule during pupal development. Preliminary data indicate that rst promotes an inter-cell competition between L4 axons by an unknown mechanism. I have formulated a specific hypothesis that can be tested with the live-imaging developed in the preliminary data (Objective 2). Lastly, cell contacts are necessary for synapse formation, but to what extent they determine partner choice has remained controversial. I propose to employ cell-contact FRET between L4 and two postsynaptic partner neurons (Tm2 and Tm9) to investigate how increased or decreased branch dynamics and adhesion affect synapse numbers and partnerships in the adult. These experiments address a fundamental question in the field of neurodevelopment about the relative contributions to synapse formation of availability through adjacencies versus molecular specification (Objective 3).

• DFG Programme: Position