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Glycan sulfation patterns define autophagy flux at axon tip via PTPRσ-cortactin axis

Abstract

Chondroitin sulfate (CS) and heparan sulfate (HS) are glycosaminoglycans that both bind the receptor-type protein tyrosine phosphatase PTPRσ, affecting axonal regeneration. CS inhibits axonal growth, while HS promotes it. Here, we have prepared a library of HS octasaccharides and, together with synthetic CS oligomers, we found that PTPRσ preferentially interacts with CS-E—a rare sulfation pattern in natural CS—and most HS oligomers bearing sulfate and sulfamate groups. Consequently, short and long stretches of natural CS and HS, respectively, bind to PTPRσ. CS activates PTPRσ, which dephosphorylates cortactin—herein identified as a new PTPRσ substrate—and disrupts autophagy flux at the autophagosome–lysosome fusion step. Such disruption is required and sufficient for dystrophic endball formation and inhibition of axonal regeneration. Therefore, sulfation patterns determine the length of the glycosaminoglycan segment that bind to PTPRσ and define the fate of axonal regeneration through a mechanism involving PTPRσ, cortactin and autophagy.

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Fig. 1: The synthetic oligosaccharides used in this study
Fig. 2: Determination of the receptor-binding structures of sulfated glycans by SPR
Fig. 3: Sulfated glycans regulate PTPRσ
Fig. 4: Abnormal accumulation of autophagosomes in dystrophic endballs by disrupted autophagy flux
Fig. 5: Disruption of autophagy flux induce dystrophic endballs
Fig. 6: Cortactin is a substrate for PTPRσ

Data availability

Source data for Fig. 4a and Supplementary Fig. 5 have been provided as Supplementary Videos 14. All other data that support the conclusions are available from the authors on request.

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Acknowledgements

We thank T. Kuboyama, T. Tojima and H. Kamiguchi (RIKEN BSI) for their technical guidance with the primary culture of DRG neurons. The expression vector for cortactin was kindly provided by M. Yoshida (RIKEN). We also thank N. Sugiura (Aichi Medical University) for his critical comment on the biochemical data. We wish to acknowledge the Division for Medical Research Engineering, Nagoya University School of Medicine, for the technical support during the electron microscopy and SPR assay. This work was supported by a Grant-in-Aid for Scientific Research on Innovative Areas (KAKENHHI Grant No. 23110002 to K.K.) and a Grant-in-Aid for Scientific Research (KAKENHHI Grant No. 16H05139 to K.K.) from the Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan, and by a Grant-in-Aid for Young Scientists (KAKENHI Grant No. 26860209 to K.S.) from the Japan Society for the Promotion of Science (JSPS), Japan and by the Ministry of Science and Technology, Taiwan (grant nos. MOST 106-2745-M-001-001-ASP, MOST 106-2113-M259-009, MOST 106-0210-01-15-02 and MOST 106-2113-M-001-009-MY2 to S.C.H) and Academia Sinica (grant no. AS-IA-104-L04 to S.C.H.).

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Contributions

K.S., T.O. and K.K. designed and performed biological experiments. Y.-C.K., C.-F.T., M.M.L.Z. and A.B. synthesized HS oligosaccharides under supervision by S.C.H. Y.G., S.N. and H.K. contributed biochemical experiments. M.M., Y.I., and K.U. contributed in vivo SCI experiments. J.-I.T. synthesized CS oligosaccharides. K.S., T.O., M.M.L.Z. and K.K. wrote the paper. All authors discussed the results and commented on the manuscript.

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Correspondence to Shang-Cheng Hung or Kenji Kadomatsu.

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Supplementary information

Supplementary Information

Supplementary Tables 1 and 2, Supplementary Figs. 1–19, Supplementary Video captions.

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Supplementary Note

Synthetic Procedures

Supplementary Video 1

Live imaging of growth cones.

Supplementary Video 2

Live imaging of a dystrophic endball.

Supplementary Video 3

Live imaging of retrograde transport of autophagosomes in growth cones.

Supplementary Video 4

Live imaging of retrograde transport of autophagosomes in a dystrophic endball.

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Sakamoto, K., Ozaki, T., Ko, YC. et al. Glycan sulfation patterns define autophagy flux at axon tip via PTPRσ-cortactin axis. Nat Chem Biol 15, 699–709 (2019). https://doi.org/10.1038/s41589-019-0274-x

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