Sp1, a transcription factor, enhances GSDME expression to promote pyroptosis. GSDME, cleaved by caspase 3, is crucial for tumor defense and chemotherapy response. This study identifies Sp1 as a positive regulator of GSDME-mediated pyroptosis. Sp1 binds to the GSDME promoter at -36 to -28 site, promoting GSDME transcription. Sp1 knockdown or inhibition reduces chemotherapy-induced pyroptosis. The regulation synergizes with STAT3 activity and antagonizes DNA methylation but has minimal effect on GSDMD-mediated pyroptosis or TNF-induced necroptosis. Sp1 regulates GSDME expression, which in turn modulates pyroptosis. Sp1 knockdown decreases GSDME expression, reducing pyroptosis and drug resistance. Sp1 also influences GSDME expression through interactions with STAT3 and DNA methylation. Sp1 is essential for GSDME-mediated pyroptosis but not for other necrosis-associated gene expressions. Sp1 regulation of GSDME provides a new mechanism for controlling pyroptosis, which may be a target for treating inflammatory diseases and cancer. The study highlights the importance of Sp1 in GSDME expression and pyroptosis, offering potential therapeutic strategies for cancer and inflammatory conditions.Sp1, a transcription factor, enhances GSDME expression to promote pyroptosis. GSDME, cleaved by caspase 3, is crucial for tumor defense and chemotherapy response. This study identifies Sp1 as a positive regulator of GSDME-mediated pyroptosis. Sp1 binds to the GSDME promoter at -36 to -28 site, promoting GSDME transcription. Sp1 knockdown or inhibition reduces chemotherapy-induced pyroptosis. The regulation synergizes with STAT3 activity and antagonizes DNA methylation but has minimal effect on GSDMD-mediated pyroptosis or TNF-induced necroptosis. Sp1 regulates GSDME expression, which in turn modulates pyroptosis. Sp1 knockdown decreases GSDME expression, reducing pyroptosis and drug resistance. Sp1 also influences GSDME expression through interactions with STAT3 and DNA methylation. Sp1 is essential for GSDME-mediated pyroptosis but not for other necrosis-associated gene expressions. Sp1 regulation of GSDME provides a new mechanism for controlling pyroptosis, which may be a target for treating inflammatory diseases and cancer. The study highlights the importance of Sp1 in GSDME expression and pyroptosis, offering potential therapeutic strategies for cancer and inflammatory conditions.