Deciphering the role of cytoplasmic domain of Channelrhodopsin in modulation of the interactome and SUMOylome of Chlamydomonas reinhardtii

Translocation of channelrhodopsins (ChRs) is mediated by the intraflagellar transport (IFT) machinery. However, the functional role of the network involving photoreceptors, IFT and other proteins in controlling algal ciliary motility is still not fully delineated. In the current study, we have identified two important motifs at the Cterminus of ChR1, VXPX and LKNE. VXPX is a known ciliary targeting sequence in animals, and LKNE is a wellknown SUMOylation motif. To the best of our knowledge, this study gives prima facie insight into the role of SUMOylation in Chlamydomonas. We prove that VMPS of ChR1 is important for interaction with GTPase CrARL11. We show that SUMO motifs are present in the C-terminus of putative ChR1s from green algae. Performing experiments with n-Ethylmaleimide (NEM) and Ubiquitin-like protease 1 (ULP-1), we show that SUMOylation may modulate ChR1 protein in Chlamydomonas. Experiments with 2D08, a known sumoylation blocker, increased the concentration of ChR1 protein. Finally, we show the endogenous SUMOylated proteins (SUMOylome) of C. reinhardtii, identified by using immunoprecipitation followed by nano-LC-MS/MS detection. This report establishes a link between evolutionarily conserved SUMOylation and ciliary machinery for the maintenance and functioning of cilia across the eukaryotes. Our enriched SUMOylome of C. reinhardtii comprehends the proteins related to ciliary development and photo-signaling, along with the orthologue(s) associated to human ciliopathies as SUMO targets.

Automated summary

In this study, two important motifs, VXPX and LKNE, were identified at the C-terminus of ChR1. VXPX is a ciliary targeting sequence in animals, while LKNE is a well-known SUMOylation motif. The study showed that VMPS of ChR1 is essential for interaction with GTPase CrARL11 and revealed the presence of SUMO motifs in the C-terminus of putative ChR1s from green algae. Additionally, the study demonstrated the potential role of SUMOylation in modulating ChR1 protein in Chlamydomonas.