Methanol, an organic one-carbon (C1) compound, represents an attractive alternative carbon source for microbial fermentation. Despite considerable advancements in methanol utilization by prokaryotes such as Escherichia coli, engineering eukaryotic model organisms such as Saccharomyces cerevisiae into synthetic methylotrophs remains challenging. Here, an engineered module circuit strategy combined with adaptive laboratory evolution was applied to engineer S. cerevisiae to use methanol as the sole carbon source. We revealed that the evolved glyoxylate-based serine pathway plays an important role in methanol-dependent growth by promoting formaldehyde assimilation. Further, we determined that the isoprenoid biosynthetic pathway was upregulated, resulting in an increased concentration of squalene and ergosterol in our evolved strain. These changes could potentially alleviate cell membrane damage in the presence of methanol. This work sets the stage for expanding the potential of exploiting S. cerevisiae as a potential organic one-carbon platform for biochemical or biofuel production.

甲醇是一种有机单碳 (C1) 化合物，是一种很有吸引力的微生物发酵替代碳源。尽管大肠杆菌等原核生物在甲醇利用方面取得了相当大的进步，但将酿酒酵母等真核模型生物工程化为合成甲基营养菌仍然具有挑战性。在这里，结合自适应实验室进化的工程模块电路策略被应用于工程师酿酒酵母使用甲醇作为唯一的碳源。我们揭示了进化的基于乙醛酸盐的丝氨酸途径通过促进甲醛同化在甲醇依赖性生长中起重要作用。此外，我们确定类异戊二烯生物合成途径被上调，导致我们进化菌株中角鲨烯和麦角甾醇的浓度增加。这些变化可能会减轻甲醇存在下的细胞膜损伤。这项工作为扩大利用酿酒酵母作为生化或生物燃料生产的潜在有机单碳平台的潜力奠定了基础。