Executioner caspase is proximal to Fasciclin 3 which facilitates non-lethal activation in Drosophila olfactory receptor neurons

Reviewer #1 (Public review):

Summary:

In this manuscript, Muramoto and colleagues have examined a mechanism by which the executioner caspase Drice is activated in a non-lethal context in Drosophila. The authors have comprehensively examined this in the Drosophila olfactory receptor neurons using sophisticated techniques. In particular, they had to engineer a new reporter by which non-lethal caspase activation could be detected. The authors conducted a proximity labeling experiment and identified Fasciclin 3 as a key protein in this context. While the removal of Fascilin 3 did not block non-lethal caspase activation (likely because of redundant mechanisms), its overexpression was sufficient to activate non-lethal caspase activation.

Strengths:

While non-lethal functions of caspases have been reported in several contexts, far less is known about the mechanisms by which caspases are activated in these non-lethal contexts. So, the topic is very timely. The overall detail of this work is impressive and the results for the most part are well-controlled and justified.

Weaknesses:

The behavioral results shown in Figure 6 need more explanation and clarification (more details below). As currently shown, the results of Figure 6 seem uninterpretable. Also, overall presentation of the Figures and description in legends can be improved.

Reviewer #2 (Public review):

In this study, the authors investigate the role of caspases in neuronal modulation through non-lethal activation. They analyze proximal proteins of executioner caspases using a variety of techniques, including TurboID and a newly developed monitoring system based on Gal4 manipulation, called MASCaT. They demonstrate that overexpression of Fas3G promotes the non-lethal activation of caspase Dronc in olfactory receptor neurons. In addition, they investigate the regulatory mechanisms of non-lethal function of caspase by performing a comprehensive analysis of proximal proteins of executioner caspase Drice. It is important to point out that the authors use an array of techniques from western blot to behavioral experiments and also that the generated several reagents, from fly lines to antibodies.

This is an interesting work that would appeal to readers of multiple disciplines. As a whole these findings suggest that overexpression of Fas3G enhances a non-lethal caspase activation in ORNs, providing a novel experimental model that will allow for exploration of molecular processes that facilitate caspase activation without leading to cell death.