The posttranslational modifications (PTMs) of proteins, as critical mechanisms for protein regulation, are well known to enhance the functional diversity of the proteome and dramatically participate in complicated biological processes. Recent efforts in the field of cancer biology have illustrated the extensive landscape of PTMs and their crosstalk with a wide range of pro-tumorigenic signaling pathways that decisively contribute to neoplastic transformation, tumor recurrence, and resistance to oncotherapy. Cancer stemness is an emerging concept that maintains the ability of tumor cells to self-renew and differentiate and has been recognized as the root of cancer development and therapy resistance. In recent years, the PTM profile for modulating the stemness of various tumor types has been identified. This breakthrough has shed light on the underlying mechanisms by which protein PTMs maintain cancer stemness, initiate tumor relapse, and confer resistance to oncotherapies. This review focuses on the latest knowledge of protein PTMs in reprogramming the stemness of gastrointestinal (GI) cancer. A deeper understanding of abnormal PTMs in specific proteins or signaling pathways provides an opportunity to specifically target cancer stem cells and highlights the clinical relevance of PTMs as potential biomarkers and therapeutic targets for patients with GI malignancies.