Abstract
Diverse structures facilitate direct exchange of proteins between cells, including plasmadesmata in plants and tunnelling nanotubes in bacteria and higher eukaryotes. Here we describe a new mechanism of protein transfer, flagellar membrane fusion, in the unicellular parasite Trypanosoma brucei. When fluorescently tagged trypanosomes were co-cultured, a small proportion of double-positive cells were observed. The formation of double-positive cells was dependent on the presence of extracellular calcium and was enhanced by placing cells in medium supplemented with fresh bovine serum. Time-lapse microscopy revealed that double-positive cells arose by bidirectional protein exchange in the absence of nuclear transfer. Furthermore, super-resolution microscopy showed that this process occurred in ≤1 minute, the limit of temporal resolution in these experiments. Both cytoplasmic and membrane proteins could be transferred provided they gained access to the flagellum. Intriguingly, a component of the RNAi machinery (Argonaute) was able to move between cells, raising the possibility that small interfering RNAs are transported as cargo. Transmission electron microscopy showed that shared flagella contained two axonemes and two paraflagellar rods bounded by a single membrane. In some cases flagellar fusion was partial and interactions between cells were transient. In other cases fusion occurred along the entire length of the flagellum, was stable for several hours and might be irreversible. Fusion did not appear to be deleterious for cell function: paired cells were motile and could give rise to progeny while fused. The motile flagella of unicellular organisms are related to the sensory cilia of higher eukaryotes, raising the possibility that protein transfer between cells via cilia or flagella occurs more widely in nature.
Highlights
Intercellular bridges enabling the direct exchange of macromolecules between cells have been described in a diverse set of multicellular and unicellular organisms
Tunnelling nanotubes (TNT) are dynamic, ultrathin membranous structures that have been observed to form de novo when mammalian cells were mixed in culture5
When trypanosomes expressing cytoplasmic DsRED and green fluorescent protein (GFP)-procyclin were mixed together, we observed that DsRED was distributed between two interacting cells, but GFP-procyclin was only transferred to the flagellum of the recipient and not the rest of the cell surface (Figure 4A). These results indicate that proteins on the outer leaflet of the plasma membrane can be transferred between cells, but transfer appears to be restricted to the flagellum
Summary
Intercellular bridges enabling the direct exchange of macromolecules between cells have been described in a diverse set of multicellular and unicellular organisms. These include plasmodesmata in plants, septal pores in fungi and gap junctions and tunnelling nanotubes in animal cells. Tunnelling nanotubes (TNT) are dynamic, ultrathin membranous structures that have been observed to form de novo when mammalian cells were mixed in culture. Tunnelling nanotubes (TNT) are dynamic, ultrathin membranous structures that have been observed to form de novo when mammalian cells were mixed in culture5 They have been implicated in tissue repair, development and electrical coupling of cells and permit the transfer of whole organelles such as lysosomes or mitochondria, over distances up to several cell diameters
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