We demonstrate a transcriptional regulatory design algorithm that can boost expression in yeast and mammalian cell lines. The system consists of a simplified transcriptional architecture composed of a minimal core promoter and a synthetic upstream regulatory region (sURS) composed of up to three motifs selected from a list of 41 motifs conserved in the eukaryotic lineage. The sURS system was first characterized using an oligo-library containing 189,990 variants. We validate the resultant expression model using a set of 43 unseen sURS designs. The validation sURS experiments indicate that a generic set of grammar rules for boosting and attenuation may exist in yeast cells. Finally, we demonstrate that this generic set of grammar rules functions similarly in mammalian CHO-K1 and HeLa cells. Consequently, our work provides a design algorithm for boosting the expression of promoters used for expressing industrially relevant proteins in yeast and mammalian cell lines.