Prompt Title: research plan

It is an interesting observation that ETTIN forms liquid-like droplets in the floral meristem cells. Given that ETTIN directly interacts with the auxin phytohormone via its C-terminal disordered region, it is reasonable to hypothesize that auxin may play a role in the phase separation behavior of ETTIN. Phase separation has been shown to play an essential role in various cellular processes, and the involvement of auxin in this context could shed light on novel regulatory mechanisms.

To explore whether auxin induces or affects the phase separation of ETTIN, you could consider the following experiments:

1. In vitro phase separation assays: Purify ETTIN protein and study its phase separation behavior in vitro in the presence and absence of auxin. Observe if there is a difference in the droplet formation, droplet size, or the concentration threshold required for phase separation. This will provide a direct assessment of auxin's effect on ETTIN phase separation.

2. In vivo imaging: Perform live-cell imaging of the floral meristem cells expressing fluorescently-tagged ETTIN in the presence and absence of exogenously applied auxin. You could also use auxin biosynthesis inhibitors or manipulate endogenous auxin levels using genetic approaches. Analyze if the changes in auxin levels affect the formation or properties of ETTIN droplets in vivo.

3. Assessing the role of ETTIN's disordered region: Generate ETTIN mutants lacking the C-terminal disordered region or mutations that disrupt auxin binding. Examine the phase separation behavior of these mutant proteins in vitro and in vivo. This will help you understand the contribution of the disordered region and auxin binding in the phase separation process.

4. Investigate the involvement of other cellular factors: As other proteins or cellular factors might also modulate ETTIN phase separation, investigate potential interaction partners, post-translational modifications, or cellular conditions that could influence the phase separation process. This can be done using methods such as co-immunoprecipitation, mass spectrometry, or proteomic approaches.

By conducting these experiments, you will be able to elucidate the role of auxin in the phase separation behavior of ETTIN and potentially uncover novel regulatory mechanisms involving phase separation and phytohormone signaling.