Background: RNA alternative splicing represents a pivotal regulatory mechanism of eukaryotic gene expression, wherein splicing factors (SFs) serve as key regulators. Aberrant SF expression drives oncogenic splice variant production, thereby promoting tumorigenesis and malignant progression. However, the biological functions and potential targets of SFs remain largely underexplored. Methods: Through multi-omics analysis, we identified survival-related splicing factors (SFs) in esophageal cancer and elucidated their biological regulatory networks. To further investigate their downstream splicing targets, we combined alternative splicing events resulting from SF knockdown with those specific to esophageal cancer. Finally, these splicing events were validated through full-length RNA sequencing and confirmed in cancer cells and clinical specimens. Result: We identified six SFs that are highly expressed in esophageal cancer and correlate with poor prognosis. Further analysis revealed that these factors are significantly associated with immune infiltration, cancer stemness, tumor heterogeneity, and drug resistance. CRNKL1 was identified as a hub SFs. The target genes and pathways regulated by these SFs showed substantial overlap, suggesting their coordinated roles in promoting cancer stemness and metastasis. Specifically, alternative splicing of key markers, such as CD44 and CTTN, was regulated by most of these SFs and correlated with poor prognosis. Conclusions: Our study unveils six survival-related SFs that contribute to the aggressiveness of esophageal cancer and CTTN and CD44 alternative splicing may act as common downstream effectors of survival-related SFs. This study provides mechanistic insights into SF-mediated tumorigenesis and highlight novel therapeutic vulnerabilities in esophageal cancer.