O-GlcNAcylation of NONO is Critical for Paraspeckle Component Assembly and Contributes to Tumorigenesis

Non-POU domain-containing octamer-binding protein (NONO) is a multifunctional member of the Drosophila behavior/human splicing (DBHS) protein family with DNA- and RNA-binding activity. Recent studies revealed that NONO is O-GlcNAcylated and dysregulated in various types of cancer. Excessive O-GlcNAcylation of target proteins has also been implicated in tumorigenesis. However, it is not known whether O-GlcNAcylation of NONO promotes cancer cell growth, and little is known about the effect of O-GlcNAcylation on the biological properties of NONO. This study identifies Thr440 as the primary NONO O-GlcNAcylation site and demonstrates its crucial role in the assembly of paraspeckles, an important subnuclear compartment that facilitates NONO-dependent transcriptional regulation in mammalian cells. We demonstrated that O-GlcNAcylation of NONO is required to maintain the expression of genes related to GO category “microtubule cytoskeleton organization involved in mitosis” and to suppress the expression of genes related to GO category “cellular response to type I interferon”. Moreover, stable depletion of NONO O-GlcNAcylation or NONO knockout significantly inhibited the growth of colon cancer cells. These findings highlight the importance of NONO and its O-GlcNAcylation status in modulating cell cycle progression, immune response, and tumorigenesis.