Abstract
O-linked N-acetylglucosamine (O-GlcNAc) is an emerging post-translation modification that couples metabolism with cellular signal transduction by crosstalking with phosphorylation and ubiquitination to orchestrate various biological processes. Herein we show that it modifies the N6-methyladenosine (m6A)-mRNA reader YTHDF1 and fine-tunes its nuclear translocation by the exportin protein Crm1. First we present evidence that YTHDF1 interacts with the sole O-GlcNAc transferase (OGT). Second, we verified the YTHDF1 O-GlcNAcylation sites to be Ser196/Ser197/Ser198, as described in previous numerous chemoproteomic studies. Then we constructed the O-GlcNAc-deficient YTHDF1-S196AS197FS198A (AFA) mutants, which significantly attentuated O-GlcNAc signals. Moreover, we revealed that YTHDF1 is a nucleocytoplasmic protein, whose nuclear export is mediated by Crm1. Furthermore, O-GlcNAcylation increases the cytosolic portion of YTHDF1 by enhancing binding with Crm1, thus upregulating the downstream target (e.g. c-Myc) expression. Molecular dynamics simulations suggest that O-GlcNAcylation at S197 might promote the binding between the nuclear export signal motif and Crm1 through increasing hydrogen bonding. Mouse xenograft assays further demonstrate that YTHDF1-AFA mutants decreased the colon cancer mass and size via decreasing c-Myc expression. In sum, we found that YTHDF1 is a nucleocytoplasmic protein, whose cytosolic localization is dependent on O-GlcNAc modification. We propose that the OGT-YTHDF1-c-Myc axis might underlie colorectal cancer tumorigenesis.
Competing Interest Statement
The authors have declared no competing interest.
Abbreviation
- (TMG)
- Thiamet-G
- (MS)
- Mass spectrometry
- (IP)
- Immunoprecipitation
- (IB)
- Immunoblotting
- (5S-G)
- Acetyl-5S-GlcNAc
- (O-GlcNAc)
- O-linked β-N-acetylglucosamine
- (OGT)
- O-GlcNAc transferase
- (m6A)
- mRNA N6-methyladenosine
- (ETD)
- electron transfer dissociation