Updated project metadata. An endosymbiotic event where an archaeon acquired and intracellular protobacteria underpinned the evolution of mitochondria, a key event for the evolution of eukaryotes. Despite is biological importance, the molecular mechanisms driving the early stages of the host-endosymbiont interactions that lead to the evolution of endosymbiont-derived organelles remains poorly understood. Novymonas esmeraldas is a trypanosomatid that recently acquired a -proteobacterial endosymbiont which is compartmentalized into vacuole-like structures called symbiosomes. The life cycle of the host is not synchronized with that of its endosymbiont resulting in heterogeneous numbers of endosymbionts per cell among the population. In order to elucidate the molecular mechanisms mediating this early stage of the host-endosymbiont interactions we isolated the endosymbionts from Novymonas esmeraldas and analyzed the proteome of isolated endosymbionts, holosymbionts (cells containing endosymbionts) and aposymbionts (cells where endosymbionts where removed by treatment with antibiotics). We show that at least one host-encoded protein localizes to the symbiosome and knockout of the corresponding gene leads to an increase of over 2-fold of endosymbionts per cell. This result suggests that controlling the endosymbiont proliferation by the host is an early step for the establishment and posterior evolution of an endosymbiotic event.