Flagellated cells are of great evolutionary importance across animal and plant species. Unlike higher plants, flagellated cells are involved in reproduction of macro-algae as well as in early diverging land plants. Euglena gracilis is an emerging flagellated model organism. The current study reports that a specific calmodulin (CaM2) involved in gravitaxis of E. gracilis interacts with an evolutionary conserved flagellar protein, EgPCDUF4201. The subsequent molecular analysis showed clearly that EgPCDUF4201 is also involved in gravitaxis. We performed subcellular localization of CaM2 using immunoblotting and indirect immunofluorescence. By employing yeast two-hybrid screen, EgPCDUF4201 was identified as an interaction partner of CaM2. The C-terminus of EgPCDUF4201 is responsible for the interaction with CaM2. Silencing of N- and C-terminus of EgPCDUF4201 using RNAi resulted in an impaired gravitaxis. Moreover, indirect immunofluorescence assay showed that EgPCDUF4201 is a flagella associated protein. The current study specifically addressed some important questions regarding the signal transduction chain of gravitaxis in E. gracilis. Besides the fact that it improved the current understanding of gravity sensing mechanisms in E. gracilis, it also gave rise to several interesting research questions regarding the function of the domain of unknown function 4201 in flagellated cells.