Deletion of a C–terminal intrinsically disordered region of WRINKLED1 affects its stability and enhances oil accumulation in Arabidopsis

Abstract

These authors contributed equally to this work. SUMMARY WRINKLED1 (WRI1) is a key transcription factor governing plant oil biosynthesis. We characterized three intrinsically disordered regions (IDRs) in Arabidopsis WRI1, and found that one C-terminal IDR of AtWRI1 (IDR3) affects the stability of AtWRI1. Analysis by bimolecular fluorescence complementation and yeasttwo-hybrid assays indicated that the IDR3 domain does not determine WRI1 stability by interacting with BTB/POZ-MATH proteins connecting AtWRI1 with CULLIN3-based E3 ligases. Analysis of the WRI1 sequence revealed that a putative PEST motif (proteolytic signal) is located at the C-terminal region of AtWRI1 IDR3 . We also show that a 91 amino acid domain at the C-terminus of AtWRI1 without the PEST motif is sufficient for transactivation. We found that removal of the PEST motif or mutations in putative phosphorylation sites increased the stability of AtWRI1, and led to increased oil biosynthesis when these constructs were transiently expressed in tobacco leaves. Oil content was also increased in the seeds of stable transgenic wri1-1 plants expressing AtWRI1 with mutations in the IDR3-PEST motif. Taken together, our data suggest that intrinsic disorder of AtWRI1 IDR3 may facilitate exposure of the PEST motif to protein kinases. Thus, phosphorylation of the PEST motif in the AtWRI1 IDR3 domain may affect AtWRI1-mediated plant oil biosynthesis. The results obtained here suggest a means to increase accumulation of oils in plant tissues through WRI1 engineering.

Phosphorylation of WRINKLED1 by KIN10 Results in Its Proteasomal Degradation, Providing a Link between Energy Homeostasis and Lipid Biosynthesis

“…Further work will be required to test details of this model. Several previous reports have described proteasomal degradation of WRI1 (Chen et al, 2013;Ma et al, 2015Ma et al, , 2016. One involves a PEST sequence representing a potential phosphorylation site identified in the C-terminal intrinsically disordered region of WRI1 (IDR3-PEST; Figure 11) .…”

Selection for a Zinc-Finger Protein Contributes to Seed Oil Increase during Soybean Domestication

“…Therefore, other mechanisms of downstream genes/components may contribute to the oil accumulation. Recent studies demonstrate that the extension of WRI1 expression in the mid phase of seed development and increased WRI1 stability through C-terminal manipulation or interaction with 14-3-3 proteins play more critical roles in seed oil accumulation than the strength of WRI1 expression and lead to significant enhancement of seed oil content Ma et al, 2015Ma et al, , 2016Kanai et al, 2016). Considering that the function of GmZF351 partially depends on WRI1, it is possible that GmZF351 may fine-tune the temporal expression of WRI1 and lipid biosynthesis genes and/or may facilitate the stabilization of the WRI1 protein as well as the promotion of oil content.…”

Nannochloropsis, a rich source of diacylglycerol acyltransferases for engineering of triacylglycerol content in different hosts

“…Nicotiana benthamiana plants were grown as described in [40]. Four-week-old plants were chosen for transient expression experiments.…”

“…Agrobacterium tumefaciens -mediated transient expression of the full-length NoDGTT5 in N. benthamiana leaves was performed as described previously by [40]. …”