Cross-stress transcriptomic integration in Marchantia polymorpha reveals stress-specific expression of regulated cell-death-associated genes

Regulated cell death (RCD) contributes to plant development, stress adaptation, and immunity, but whether RCD-associated genes show conserved transcriptional responses across diverse stresses remains unclear. Here, we performed a comparative reanalysis of publicly available Marchantia polymorpha RNA-seq datasets to investigate recurrent and stress-specific expression of RCD-associated genes. We uniformly processed 36 RNA-seq samples from three independent studies, yielding nine stress-versus-control comparisons: eight abiotic (seven from one study and an independent high-light experiment) and one pathogen infection. Differentially expressed genes (DEGs) were identified for each of the 9 stress datasets compared to their respective controls, which were then examined for shared DEGs among different sets of stress vs. control comparisons. A curated list of 14 RCD-associated genes previously identified in M. polymorpha were used to examine conservation of transcriptional response. Integrated expression data were additionally examined using weighted gene co-expression network analysis (WGCNA) after batch correction of dataset. Of 34 DEGs shared across all nine comparisons, only 12 changed in the same direction in every comparison. The 34 genes included two transcription factors, an AP2/ERF-family factor (Mp5g22160) and a bHLH factor (Mp2g00890), which were upregulated in infection and Schröder high-light but downregulated in all seven Tan et al. comparisons. Among 14 curated RCD-associated genes, 10 were differentially expressed in at least one comparison, and MpZOU2 was the only one regulated in more than one (upregulated in cold, darkness and infection; downregulated in Tan high-light). WGCNA identified 64 co-expression modules, including an infection-associated module enriched in protein ubiquitination, proteasome-mediated degradation, ER-associated degradation, autophagy, and membrane trafficking, and a nitrogen-deficiency and osmotic-stress-associated module enriched in oxidative stress responses and cell-wall organization. Together, these findings indicate that M. polymorpha does not exhibit a uniform transcriptional RCD program across diverse stresses. Instead, RCD-associated genes appear to be selectively recruited within broader, context-dependent stress-response networks. The recurrent regulation of MpZOU2 identifies it as a candidate for functional investigation, while emphasizing that transcript-level changes alone do not establish RCD execution.