AbstractSeed death resulting from hybridization between Arabidopsis thaliana and Arabidopsis arenosa has complex genetic determination and involves deregulation 5 to 8 d after pollination (DAP) of AGAMOUS-LIKE genes and retroelements. To identify causal mechanisms, we compared transcriptomes of compatible and incompatible hybrids and parents at 3 DAP. Hybrids misexpressed endosperm and seed coat regulators and hyperactivated genes encoding ribosomal, photosynthetic, stress-related, and immune response proteins. Regulatory disruption was more severe in Columbia-0 hybrids than in C24 hybrids, consistent with the degree of incompatibility. Maternal loss-of-function alleles for endosperm growth factor TRANSPARENT TESTA GLABRA2 and HAIKU1 and defense response regulators NON-EXPRESSOR OF PATHOGENESIS RELATED1 and SALICYLIC ACID INDUCTION-DEFICIENT2 increased hybrid seed survival. The activation of presumed POLYCOMB REPRESSIVE COMPLEX (PRC) targets, together with a 20-fold reduction in expression of FERTILIZATION INDEPENDENT SEED2, indicated a PRC role. Proximity to transposable elements affected natural variation for gene regulation, but transposon activation did not differ from controls. Collectively, this investigation provides candidates for multigenic orchestration of the incompatibility response through disruption of endosperm development, a novel role for communication between endosperm and maternal tissues and for pathways previously connected to immunity, but, surprisingly, does not identify a role for transposons.