TOR-inhibitor insensitive-1 (TRIN1) regulates cotyledons greening in Arabidopsis

Abstract

Target of Rapamycin (TOR) is an eukaryotic protein kinase and evolutionally conserved from the last eukaryotic common ancestor (LECA) to humans. The growing evidences have shown that TOR signaling acts as a central controller of cell growth and development. The downstream effectors of TOR have been well-identified in yeast and animals by using the immunosuppression agent rapamycin. However, less is known about TOR in plants. This is largely due to the fact that plants are insensitive to rapamycin. In this study, AZD8055 (AZD), the novel ATP-competitive inhibitor of TOR, was employed to decipher the downstream effectors of TOR in Arabidopsis. One AZD insensitive mutant, TOR-inhibitor insensitive-1 (trin1), was screened from 10,000 EMS-induced mutation seeds. The cotyledons of trin1 can turn green when its seeds were germinated on ½ MS medium supplemented with 2 μM AZD, whereas the cotyledons greening of wild-type (WT) can be completely blocked at this concentration. Through genetic mapping, TRIN1 was mapped onto the long arm of chromosome 2, between markers SGCSNP26 and MI277. Positional cloning revealed that TRIN1 was an allele of ABI4, which encoded an ABA-regulated AP2 domain transcription factor. Plants containing P35S::TRIN1 or P35S::TRIN1-GUS were hypersensitive to AZD treatment and displayed the opposite phenotype observed in trin1. Importantly, GUS signaling was significantly enhanced in P35S::TRIN1-GUS transgenic plants in response to AZD treatment, indicating that suppression of TOR resulted in the accumulation of TRIN1. These observations revealed that TOR controlled seed-to-seedling transition by negatively regulating the stability of TRIN1 in Arabidopsis. For the first time, TRIN1, the downstream effector of TOR signaling, was identified through a chemical genetics approach.

The Arabidopsis TOR Kinase Specifically Regulates the Expression of Nuclear Genes Coding for Plastidic Ribosomal Proteins and the Phosphorylation of the Cytosolic Ribosomal Protein S6

“…These genes were also repressed by ABA but strongly induced by the application of brassinolide. This is consistent with the inhibitory effects of ABA on the growth-promoting hormones like brassinosteroid and with the role of TOR in brassinosteroid (Zhang et al, 2009, 2016) and ABA signaling (Kravchenko et al, 2015; Li et al, 2015), which could result in TOR inhibition. As reported earlier by Pal et al (2013) we found diverse and uncoordinated responses to variations in sugar supply for nuclear genes coding for pRPS while their expression was repressed by nitrogen starvation as observed in Chlamydomonas (Schmollinger et al, 2014).…”

“…About half of the plastid small ribosome subunit (30S) proteins and most of the large subunit (50S) proteins are encoded by the nuclear genome (Yamaguchi and Subramanian, 2000). TOR silencing (Deprost et al, 2007; Xiong and Sheen, 2012; Caldana et al, 2013), inhibition by rapamycin (Sormani et al, 2007; Ren et al, 2011) or by AZD-8055 (Montané and Menand, 2013; Li et al, 2015) as well as mutations affecting the TORC1 complex (Moreau et al, 2012; Kravchenko et al, 2015) consistently result in leaf chlorosis and yellowing. We show here that TOR inhibition results in a coordinated decrease in pRP expression, at the level of protein abundance but also at the total and translated mRNA levels, which could explain these chlorotic phenotypes ( Figure 3 ).…”

The crosstalk between Target of Rapamycin (TOR) and Jasmonic Acid (JA) signaling existing in Arabidopsis and cotton

“…We found that AZD can induce growth retardation of cotton seedlings, and these effects were dose-dependent (as indicated by root length and fresh weight). This suggests that AZD based approach can be used to probe TOR signaling in cotton similar to other plants89.…”

“…Furthermore, we also found the altered expression profiles of auxin and abscisic acid (ABA) related genes under TOR inhibition in cotton; these observations are highly consistent with similar treatments in Arabidopsis 11 (Supplementary Table 3). As the crosstalk between TOR and ABA or auxin signaling pathways have already been previously investigated92930, we focused on JA signaling pathway for further investigation. The expression levels of some randomly-selected genes were verified using QRT-PCR, and the results were consistent with the RNA-Seq analysis (Supplementary Figure 2).…”

“…To explore TOR functions using RAP in higher plants, yeast FKBP12 -based Arabidopsis transgenic plants (BP12-2) were produced, which confer conditional sensitivity to rapamycin7. Recent studies have also revealed that the ATP-competitive TOR specific inhibitors, which were named after active-site TOR inhibitors (asTORis) including AZD8055 (AZD), Torin1 and Torin2, can also effectively inhibit TOR activity and retard plant growth8910. These inhibitors are effective against both TORC1 and TORC2 complexes and have broader kinase-dependent inhibition on TOR than the widely used rapamycin.…”