De novo Golgi biogenesis requires coordinated transactivation of a Golgi regulon

The Golgi apparatus expands during differentiation and high secretory demand, yet the transcriptional control of its biogenesis remains poorly defined. Here, we developed a targeted enzymatic ablation method to eliminate the Golgi and trigger de novo organelle formation. Single-cell RNA-seq of cells rebuilding Golgi revealed an orchestrated induction of a broad Golgi gene network coinciding with structural and functional organelle maturation. This gene set spans all Golgi sub-compartments and functions, including glycosylation, trafficking, and ion transport, thus supporting the concept of a unified Golgi regulon, enabling the simultaneous expression of components required for the organelle structural and functional integrity. Through promoter analysis and RNAi screening, we identified CREB3L1 as a key transcriptional regulator critical for Golgi gene activation and organelle reformation. These findings indicate that CREB3L1-dependent transcriptional mechanisms orchestrate a complete Golgi biogenesis program that may be essential for organelle regeneration and for secretory pathway plasticity during physiological remodeling.