Growing and dividing: how O-GlcNAcylation leads the way

Abstract

Cell cycle errors can lead to mutations, chromosomal instability, or death; thus, the precise control of cell cycle progression is essential for viability. The nutrient-sensing posttranslational modification, O-GlcNAc, regulates the cell cycle allowing one central control point directing progression of the cell cycle. O-GlcNAc is a single N-acetylglucosamine sugar modification to intracellular proteins that is dynamically added and removed by O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA), respectively. These enzymes act as a rheostat to fine-tune protein function in response to a plethora of stimuli from nutrients to hormones. O-GlcNAc modulates mitogenic growth signaling, senses nutrient flux through the hexosamine biosynthetic pathway, and coordinates with other nutrient-sensing enzymes to progress cells through Gap phase 1 (G₁). At the G₁/S transition, O-GlcNAc modulates checkpoint control, while in S Phase, O-GlcNAcylation coordinates the replication fork. DNA replication errors activate O-GlcNAcylation to control the function of the tumor-suppressor p53 at Gap Phase 2 (G₂). Finally, in mitosis (M phase), O-GlcNAc controls M phase progression and the organization of the mitotic spindle and midbody. Critical for M phase control is the interplay between OGT and OGA with mitotic kinases. Importantly, disruptions in OGT and OGA activity induce M phase defects and aneuploidy. These data point to an essential role for the O-GlcNAc rheostat in regulating cell division. In this review, we highlight O-GlcNAc nutrient sensing regulating G₁, O-GlcNAc control of DNA replication and repair, and finally, O-GlcNAc organization of mitotic progression and spindle dynamics.

Keywords: O-GlcNAc; O-GlcNAc transferase; O-GlcNAcase; cell cycle; cyclin; mTOR; mini-chromosome complex; nutrient sensing; p53; spindle.

Figure 1

Overview of the cell cycle. The cell cycle begins with gap phase 1 (G₁) followed by DNA synthesis phase (S), Gap phase 2 (G₂), and division during the mitotic phase (M). Cyclins, cyclin dependent kinases (CDK), and checkpoints regulate cell cycle progression in each phase. Mitogenic (ERK) signaling via growth factors, elevated CDK4/6-cyclin D activity, and increased mTORC1 function are essential for G₁ progression. After disruption of retinoblastoma protein (pRB)–E2F interaction at the G₁/S checkpoint, S phase utilizes the mini-chromosome complex (MCM) to synthesize DNA. DNA synthesis errors are checked at S and G2, and activation of the transcription factor p53 is essential to arrest the cell cycle by increasing cyclin-dependent kinase inhibitor p21 while errors are fixed. During mitosis, cyclin B bound to CDK1 regulates mitosis via phosphorylation, and other mitotic kinases such as the Aurora Kinase B complex directs cell division. ERK, extracellular activated protein kinase; mTORC, mammalian target of rapamycin complex.

Figure 2

Entry into G₁is facilitated by O-GlcNAc. Mitogenic signals activate ERK1/2 to phosphorylate the c-MYC transcription factor. Activation of c-MYC induces cyclin D expression. Cyclin D then binds to and activate CKD4/6. O-GlcNAcylation modulates ERK function, c-MYC transcriptional activity, and cyclin D stability. ERK, extracellular activated protein kinase.

Figure 3

Interplay between nutrient-sensing pathways drive G1 progression. UDP-GlcNAc, derived from the hexosamine biosynthetic pathway (HBP), is used by OGT to modify several G₁ proteins including AMPK. OGT interacts and modifies other nutrient sensors such as the MTOR–AMPK axis and FASN. However, AMPK activation inhibits GFAT, the rate liming enzyme of the HBP. Together, these sensors gauge nutrient availability to induce G₁ progression. AMPK, AMP-activated protein kinase; FASN, fatty acid synthase; GFAT, glutamine: fructose-6-phosphate amidotransferase; OGT, O-GlcNAc transferase.

Figure 4

O-GlcNAc stabilizes DNA replication machinery and modifies p53 stabilization. At the G₁/S checkpoint, pRB is O-GlcNAcylated and contributes to checkpoint regulation. S phase entry is characterized by DNA synthesis and replication in preparation for mitosis. O-GlcNAc enhances the ability of the mini-chromosome complex (MCM) to bind chromatin and induce helicase activity, thereby permitting DNA replication. Errors in replication are sensed by p53, a key checkpoint sensor in G₂. p53 is degraded when bound to MDM2. Phosphorylation of p53 prevents MDM2 association and degradation; O-GlcNAc inhibits phosphorylation, increasing the stability of p53. Once stable, p53 induces transcription of p21. p21 is an inhibitor of CDKs and prevents cell cycle progression. OGT or OGA overexpression can directly induce p21 expression and increase p53 stabilization. OGT, O-GlcNAc transferase; OGA, O-GlcNAcase; pRb, retinoblastoma protein.

Figure 5

O-GlcNAc influences mitosis via O-GlcNAcylation of critical mitotic mediators. At prophase, overexpression of OGT or OGA modifies CDK1-cyclin B activity via inhibition of PLK1, ultimately inhibiting mitotic entry. Emerin is reciprocally O-GlcNAcylated and phosphorylated, and Emerin O-GlcNAcylation affects degradation of the nuclear envelope during prophase. The metaphase to anaphase transition is activated by APC/C, and O-GlcNAcylation of CDH1 is required to activate this transition. Chromatin condensation during metaphase is regulated in part by O-GlcNAcylation of histones or O-GlcNAc modulation of other epigenetic marks. Overexpression of OGT or OGA increases chromatin acetylation and decreases methylation. OGT, O-GlcNAc transferase; OGA, O-GlcNAcase.

Figure 6

O-GlcNAcylation controls spindle assembly/disassembly. Spindle dynamics are tightly regulated by AurB which interacts with OGT and OGA. AurB localizes at the spindle and is regulated by MYPT1 and PP1; however, MYPT1 O-GlcNAcylation alters this inhibition. Furthermore, EWS associates with AurB at the spindle, and loss of OGA affects this interaction. O-GlcNAcylated NUMA inhibits NUMA phosphorylation, disturbing the organization of the mitotic poles. Several spindle and mitotic regulatory proteins are O-GlcNAcylated, and disruptions in the O-GlcNAc rheostat result in aneuploidy. EWS, Ewing sarcoma; OGT, O-GlcNAc transferase; OGA, O-GlcNAcase; PP1, protein phosphatase 1.