Basic leucine zipper transcription activators – tools to improve production and quality of human erythropoietin in Nicotiana benthamiana

Human erythropoietin (hEPO) is one of the most in‐demand biopharmaceuticals, however, its production is challenging. When produced in a plant expression system, hEPO results in extensive plant tissue damage and low expression. It is demonstrated that the modulation of the plant protein synthesis machinery enhances hEPO production. Co‐expression of basic leucine zipper transcription factors with hEPO prevents plant tissue damage, boosts expression, and increases hEPO solubility. bZIP28 co‐expression up‐regulates genes associated with the unfolded protein response, indicating that the plant tissue damage caused by hEPO expression is due to the native protein folding machinery being overwhelmed and that this can be overcome by co‐expressing bZIP28. Graphical and Lay Summary Plants represent a cost‐effective platform for producing high demand biopharmaceuticals, such as human erythropoietin (hEPO). Despite the inherent challenges associated with plant‐based production, the modulation of the plant protein synthesis machinery via basic leucine zipper (bZIP) transcription factors significantly enhanced outcomes. This strategic intervention increased biomass yield (see images of plants), reduced tissue damage (see images of plants and depicted by green versus red circles at sequential days post‐infiltration), amplified hEPO expression (see immuno‐blot), and improved the solubility of hEPO.