Tuning microtubule dynamics to enhance cancer therapy by modulating FER-mediated CRMP2 phosphorylation

Yiyan Zheng,Fiona Chen,Kevin A Myers,Ritika Sethi,Frank Von Delft,C R Taylor ,David Mannion,Ming Wang,Nathanael S Gray,Robert C Bast,Sunanda Dhar,Juliet Goldsmith,Dahai Jiang,Kenta Masuda,Sunila Pradeep,Sandra Herrero-González ,Karin Hellner,Christopher Yau,Anil K Sood,Donatien Chedom Fotso,Lingegowda S Mangala,Mohammad Karaminejadranjbar ,S Knapp ,Ashwag Albukhari,Leticia Campo,D Y P Ferguson ,Abdulkhaliq Alsaadi,Gabriel López-Berestein ,Cristian Rodríguez-Aguayo ,Fabrizio Miranda,Ahmed A Ahmed

doi:10.1038/s41467-017-02811-7

Abstract

Though used widely in cancer therapy, paclitaxel only elicits a response in a fraction of patients. A strong determinant of paclitaxel tumor response is the state of microtubule dynamic instability. However, whether the manipulation of this physiological process can be controlled to enhance paclitaxel response has not been tested. Here, we show a previously unrecognized role of the microtubule-associated protein CRMP2 in inducing microtubule bundling through its carboxy terminus. This activity is significantly decreased when the FER tyrosine kinase phosphorylates CRMP2 at Y479 and Y499. The crystal structures of wild-type CRMP2 and CRMP2-Y479E reveal how mimicking phosphorylation prevents tetramerization of CRMP2. Depletion of FER or reducing its catalytic activity using sub-therapeutic doses of inhibitors increases paclitaxel-induced microtubule stability and cytotoxicity in ovarian cancer cells and in vivo. This work provides a rationale for inhibiting FER-mediated CRMP2 phosphorylation to enhance paclitaxel on-target activity for cancer therapy.

Highlights

Though used widely in cancer therapy, paclitaxel only elicits a response in a fraction of patients
We show how FER phosphorylates Collapsin response mediator protein 2 (CRMP2) at Y479 and Y499 to regulate CRMP2 structure and its interaction with microtubules, respectively
We show that loss of CRMP2 phosphorylation, either via FER depletion or inhibition, significantly sensitizes ovarian cancer cells to paclitaxel treatment

Summary

Results

We set out to identify a microtubule-associated protein whose function: (1) is modulated by an oncogene kinase that could be targeted therapeutically and (2) directly controls microtubule bundling and stability. FER kinase has been reported to promote ovarian cancer metastasis[20]. Previous reports found that FES, the only other family member of FER, is able to phosphorylate CRMP2 at Y3221. CRMP2 has been reported to be a known modulator of microtubule assembly[13]. We tested whether CRMP2 can modulate microtubule bundling and stability and whether this function can be regulated by FER. In vitro kinase assays using a catalytically active glutathione S-transferase (GST)-tagged truncation of FER (residues 541–822) and recombinant CRMP2 (residues 13–516) confirmed that FER phosphorylates CRMP2 (Supplementary Fig. 1a)

64 IB: CRMP2

Discussion

Methods