Abstract text: Seed mucilage is a specialized, pectin-rich extracellular matrix and a valuable model for studying cell wall assembly. Although the role of homogalacturonan (HG) in mucilage structuration is well characterized under standard growth conditions, its response to heat stress remains unexplored. In silico analysis of expression datasets revealed that several seed coat–related genes involved in HG modification are misregulated at high temperature, including PMEI6, PMEI17, PME58, PLL21, and GAUT11, suggesting a key role for HG in the heat stress response. Among these, PMEI6, PME58, and GAUT11 are mucilage-specific genes involved in mucilage structure, whereas PMEI17 and PLL21 remain uncharacterized. To investigate HG dynamics under heat stress, mutant lines were grown at 28 °C after flowering, and mucilage phenotypes were analyzed using biochemical and cytological approaches. Wild-type Col-0 seeds showed a marked reduction (~75%) in mucilage release area, with around 30% failing to release mucilage completely, indicating a strong impact of heat on mucilage organization. All HG-related mutants showed exacerbated defects, with pmei6 most affected. Methylation analyses revealed increased HG methylesterification in WT, pmei6, and pmei17 under heat stress, suggesting that altered HG methylation alone cannot fully explain the phenotype and pointing to broader pectin remodeling in response to elevated temperature.