Disruption of either the auxin transporter PIN-FORMED 1 (PIN1) or the protein kinase PINOID (PID) leads to the development of pin-like inflorescences. Previous studies suggested that PID phosphorylates and activates PIN1. Here we report unexpected findings about the genetic interactions between the two genes. We deleted the first 2/3 of the PIN1 coding sequence using CRISPR/Cas9 and the resulting pin1 mutant (pin1-27) was a strong allele. Surprisingly, heterozygous pin1-27 suppressed three independent pid null mutants whereas homozygous pin1-27 enhanced the phenotypes of the pid mutants during embryogenesis. Furthermore, we show that deletion of either the hydrophilic loop or the second half of PIN1 also abolished PIN1 function, yet those heterozygous pin1 mutants were also capable of rescuing pid nulls. Moreover, we inserted GFP into the hydrophilic loop of PIN1 through CRISPR-mediated homology-directed repair (HDR). The GFP signal and pattern in the PIN1-GFP HDR line are similar to those in the previously reported PIN1-GFP transgenic lines. Interestingly, the PIN1-GFP HDR line also rescued various pid null mutants in a semi-dominant fashion. In addition, the previously reported key phosphorylation sites in PIN1 were still phosphorylated in PIN1-GFP pid plants. We conclude that PID is not directly required for phosphorylation and activation of PIN1.
[doi:10.25345/C5RX93Q2P]
[dataset license: CC0 1.0 Universal (CC0 1.0)]
Keywords: TMT ; phosphorylation ; Auxin ; PIN1 ; PID ; CRISPR/Cas9
Principal Investigators: (in alphabetical order) |
Steven P Briggs, UCSD, USA Yunde Zhao, University of California San Diego, UCSD |
Submitting User: | zhouxinshen |
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