Circadian clock-related transcription factors Reveille 1 and Reveille 8 regulate the O-acetylserine cluster genes and sulfur homeostasis.

O-acetylserine (OAS) is the sulfur acceptor in primary sulfate assimilation and the precursor of cysteine, synthesized by serine acetyltransferase (SERAT). Beyond its metabolic role, OAS acts as a signaling molecule, accumulating during sulfur starvation and inducing sulfur-deficiency marker genes. Recently, OAS was also shown to increase transiently after a light-dark transition, triggering the induction of a group of "OAS cluster" genes. Here, we investigated the mechanisms underlying OAS accumulation and the regulation of OAS cluster genes. We found that no single SERAT isoform accounts for the full induction of OAS cluster genes; instead, the loss of any of the 3 major isoforms (SERAT1;1, SERAT2;1, SERAT2;2) reduced induction, with SERAT2;2 playing a predominant role. All 5 isoforms contributed to the response to varying degrees. In addition, 3 transcription factors were required: the sulfur-deficiency regulator SLIM1 and the circadian clock components RVE1 and RVE8. These factors bound to OAS cluster gene promoters and unexpectedly controlled OAS accumulation itself. While RVE1 and RVE8 did not transduce the OAS signal, they contributed to sulfur-deficiency responses at both the transcriptional and metabolic levels. Together, our findings reveal a complex regulatory network involving multiple SERAT isoforms and transcription factors, advancing our understanding of sulfur homeostasis and its integration with light and circadian regulation.