Efficient coproduction of multiple proteins or their subunits is important in many facets of life sciences and biotechnology. Here， we report a novel approach that exploits the synergy between an engineered mini-intein and an ubiquitin variant to achieve coordinated coexpression of multiple proteins in eukaryotic hosts， from a single open reading frame that encodes a polyprotein precursor consisting of proteins of interest (POIs) connected by an intervening intein-ubiquitin fusion domain. The intein variant mediates highly active autocatalytic cleavage at its N-terminus， whereas the endogenous deubiquitinases cleave at ubiquitin's C-terminus， leading to the release of the POIs. Using fluorescent reporter proteins for proof-of-concept， utility of the intein-ubiquitin domain was validated in higher plants and yeast systems. Essentially complete release of the POIs was achieved as demonstrated with western blots. Proteins expressed using the intein-ubiquitin system potentially preserve their intended sequences， which is important for preventing alteration of POI function.