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Probing the 14-3-3 Isoform-Specificity Profile of Interactions Stabilized by Fusicoccin A

preprint
revised on 03.04.2020 and posted on 06.04.2020 by James Frederich, Ananya Sengupta, Josue Liriano, Ewa A. Bienkiewicz, Brian G. Miller
Fusicoccin A (FC) is a fungal phytotoxin that stabilizes protein–protein interactions (PPIs) between 14-3-3 adapter proteins and their phosphoprotein interaction partners. In recent years, FC has emerged as an important chemical probe of human 14-3-3 PPIs implicated in cancer and neurological diseases. These previous studies have established the structural requirements for FC-induced stabilization of 14-3-3·client phosphoprotein complexes; however, the effect of different 14-3-3 isoforms on FC activity has not been systematically explored. This is a relevant question for the continued development of FC variants because there are seven distinct isoforms of 14-3-3 in humans. Despite their remarkable sequence and structural similarities, a growing body of experimental evidence supports both tissue-specific expression of 14-3-3 isoforms and isoform-specific functions in vivo. Herein, we report the isoform-specificity profile of FC in vitrousing recombinant human 14-3-3 isoforms and a focused library of fluorescein-labeled hexaphosphopeptides mimicking the C-terminal 14-3-3 recognition domains of client phosphoproteins targeted by FC in cell culture. Our results reveal modest isoform preferences for individual client phospholigands and demonstrate that FC differentially stabilizes PPIs involving 14-3-3s. Together, these data provide strong motivation for the development of non-natural FC variants with enhanced selectivity for individual 14-3-3 isoforms.

Funding

R01-GM125926, R01-GM133843, R01-GM115388

History

Email Address of Submitting Author

jfrederich@fsu.edu

Institution

Florida State University

Country

United States

ORCID For Submitting Author

0000-0003-2077-4436

Declaration of Conflict of Interest

authors declare no conflict of interest

Version Notes

updated on 04/03/2020 to reflect a typo in the structure of FC

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