Structure and dynamics of the Arabidopsis O-fucosyltransferase SPINDLY.

Abstract

SPINDLY (SPY) in Arabidopsis thaliana is a novel nucleocytoplasmic protein O-fucosyltransferase (POFUT), which regulates diverse developmental processes. Sequence analysis indicates that SPY is distinct from ER-localized POFUTs and contains N-terminal tetratricopeptide repeats (TPRs) and a C-terminal catalytic domain resembling the O-linked-N-acetylglucosamine (GlcNAc) transferases (OGTs). However, the structural feature that determines the distinct enzymatic selectivity of SPY remains unknown. Here we report the cryo-electron microscopy (cryo-EM) structure of SPY and its complex with GDP-fucose, revealing distinct active-site features enabling GDP-fucose instead of UDP-GlcNAc binding. SPY forms an antiparallel dimer instead of the X-shaped dimer in human OGT, and its catalytic domain interconverts among multiple conformations. Analysis of mass spectrometry, co-IP, fucosylation activity, and cryo-EM data further demonstrates that the N-terminal disordered peptide in SPY contains trans auto-fucosylation sites and inhibits the POFUT activity, whereas TPRs 1-5 dynamically regulate SPY activity by interfering with protein substrate binding.

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Published Version (Please cite this version)

10.1038/s41467-023-37279-1

Publication Info

Kumar, Shivesh, Yan Wang, Ye Zhou, Lucas Dillard, Fay-Wei Li, Carly A Sciandra, Ning Sui, Rodolfo Zentella, et al. (2023). Structure and dynamics of the Arabidopsis O-fucosyltransferase SPINDLY. Nature communications, 14(1). p. 1538. 10.1038/s41467-023-37279-1 Retrieved from https://hdl.handle.net/10161/26950.

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Sun

Tai-ping Sun

Professor of Biology

The diterpenoid phytohormone gibberellin (GA) plays pivotal roles in regulating growth and development throughout the life cycle of higher plants.  Mutations affecting GA biosynthesis or GA response were the key to control plant stature in wheat and rice that led to dramatically increased grain yield and contributed greatly to the success of the ‘Green Revolution’ in the 1960s.  By multi-faceted approaches using the reference plant Arabidopsis, my lab has made major breakthroughs in elucidating the sites and regulatory mechanisms of GA biosynthesis, and the conserved molecular events of GA perception and the early GA signaling pathway.  We identified the nuclear transcriptional regulators DELLA proteins, which function as master growth repressors by inhibiting all aspects of GA responses.  Binding of GA to its nuclear receptor GID1 enhances the GID1-DELLA interaction, which in turn leads to the rapid proteolysis of DELLA through the ubiquitin-proteasome pathway, and allows transcriptional reprogramming of GA-responsive genes.  We and other researchers further showed that GA-GID1-DELLA is a key regulatory module that controls plant growth by integrating internal developmental cues, and external biotic and abiotic signals (light, cold, salt and pathogen stresses).  DELLA proteins play a central role in these processes via direct protein-protein interactions with key transcription factors.  Our recent studies using genetic and physiological analyses together with chemical biology methods indicate that DELLA’s binding affinity to interacting proteins are oppositely regulated by two novel O-linked glycosylations on specific Ser/Thr residues: O-linked N-acetylglucosamine (O-GlcNAc) modification reduces DELLA activity, whereas O-fucosylation enhances DELLA activity.  We are investigating the global functions of O-GlcNAcylation and O-fucosylation in regulating plant development.


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