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Abstract Macrofossils with unambiguous biogenic origin and predating the one-billion-year-old multicellular fossilsBangiomorphaandProterocladusinterpreted as crown-group eukaryotes are quite rare.Horodyskiais one of these few macrofossils, and it extends from the early Mesoproterozoic Era to the terminal Ediacaran Period. The biological interpretation of this enigmatic fossil, however, has been a matter of controversy since its discovery in 1982, largely because there was no evidence for the preservation of organic walls. Here we report new carbonaceous compressions ofHorodyskiafrom the Tonian successions (~950–720 Ma) in North China. The macrofossils herein with bona fide organic walls reinforce the biogenicity ofHorodyskia. Aided by the new material, we reconstructHorodyskiaas a colonial organism composed of a chain of organic-walled vesicles that likely represent multinucleated (coenocytic) cells of early eukaryotes. Two species ofHorodyskiaare differentiated on the basis of vesicle sizes, and their co-existence in the Tonian assemblage provides a link between the Mesoproterozoic (H.moniliformis) and the Ediacaran (H.minor) species. Our study thus provides evidence that eukaryotes have acquired macroscopic size through the combination of coenocytism and colonial multicellularity at least ~1.48 Ga, and highlights an exceptionally long range and morphological stasis of this Proterozoic macrofossils.