Protein Glutamylation: From Cytoskeleton to Signaling

ABSTRACT Protein glutamylation is a reversible post‐translational modification (PTM) that involves the enzymatic addition of glutamate moieties to, and their removal from, target proteins, thereby modulating their activity, stability, localization, and molecular interactions. Initially characterized as a defining feature of the tubulin code that governs microtubule organization, ciliary function, and neuronal integrity, glutamylation is now recognized as a broader regulatory mechanism extending far beyond the cytoskeleton. This review highlights the conceptual shift from tubulin‐centered regulation to broader cellular signaling functions and summarizes recent advances in the enzymatic machinery that mediates the addition and removal of glutamate moieties, including the tubulin tyrosine ligase‐like (TTLL) family and cytosolic carboxypeptidases (CCPs). We discuss the established roles of tubulin glutamylation in ciliogenesis, neurodevelopment, cell division, and cytoskeletal organization. We then focus on the expanding repertoire of nontubulin substrates, illustrating how glutamylation of key signaling molecules—including cGAS, KLF4, IL‐7Rα, BAP1, TAB1, and YAP1—modulates innate immunity, cell‐fate determination, hematopoietic homeostasis, metabolic adaptation, tumor progression, and immune evasion. By integrating evidence across cytoskeletal biology, cell signaling, and disease mechanisms, this review proposes that glutamylation functions as a versatile signaling‐related PTM rather than merely as a component of the tubulin code. Finally, we discuss emerging therapeutic opportunities for targeting TTLL and CCP enzymes, as well as the potential of glutamylation pathways and substrates as biomarkers and pharmacological targets for human disease.