Target of Rapamycin Regulates Development and Ribosomal RNA Expression through Kinase Domain in Arabidopsis    

Abstract

Target of rapamycin (TOR) is a central regulator of cell growth, cell death, nutrition, starvation, hormone, and stress responses in diverse eukaryotes. However, very little is known about TOR signaling and the associated functional domains in plants. We have taken a genetic approach to dissect TOR functions in Arabidopsis (Arabidopsis thaliana) and report here that the kinase domain is essential for the role of TOR in embryogenesis and 45S rRNA expression. Twelve new T-DNA insertion mutants, spanning 14.2 kb of TOR-encoding genomic region, have been characterized. Nine of these share expression of defective kinase domain and embryo arrest at 16 to 32 cell stage. However, three T-DNA insertion lines affecting FATC domain displayed normal embryo development, indicating that FATC domain was dispensable in Arabidopsis. Genetic complementation showed that the TOR kinase domain alone in tor-10/tor-10 mutant background can rescue early embryo lethality and restore normal development. Overexpression of full-length TOR or kinase domain in Arabidopsis displayed developmental abnormalities in meristem, leaf, root, stem, flowering time, and senescence. We further show that TOR, especially the kinase domain, plays a role in ribosome biogenesis by activating 45S rRNA production. Of the six putative nuclear localization sequences in the kinase domain, nuclear localization sequence 6 was identified to confer TOR nuclear targeting in transient expression assays. Chromatin immunoprecipitation studies revealed that the HEAT repeat domain binds to 45S rRNA promoter and the 5# external transcribed spacer elements motif. Together, these results show that TOR controls the embryogenesis, postembryonic development, and 45S rRNA production through its kinase domain in Arabidopsis.

Ribosomal Protein S6, a Target of Rapamycin, Is Involved in the Regulation of rRNA Genes by Possible Epigenetic Changes in Arabidopsis

“…However, our ChIP data suggest that the region of the rDNA promoter to which RPS6 binds is largely different from the binding sites for Arabidopsis TOR reported by Ren et al (11). In contrast to the fact that the activity of TOR has been positively implicated in transcriptional regulation of rDNA in the study of Ren et al (11), as well as in those of yeast and animals (9, 10), our results show an RPS6-dependent mechanism in transcriptional repression of the rDNA. However, it is possible that the RPS6 exists as a component of the nucleolar TOR complex and plays a negative regulator for the rDNA transcription, antagonizing the positive effect provided by the TOR kinase.…”

“…Similar to yeast and animals (9,10), binding of the Arabidopsis TOR to the rDNA promoter has recently been reported (11). However, our ChIP data suggest that the region of the rDNA promoter to which RPS6 binds is largely different from the binding sites for Arabidopsis TOR reported by Ren et al (11).…”

“…RPS6 has been identified as the first protein to be phosphorylated in response to various signals (44,45). The presence of S6K in the nucleolus and demonstration that TOR phosphorylates RPS6 via S6K, which is sensitive to osmotic stress (17), and the direct binding of TOR to rDNA promoter (11), may control optimum transcription of rRNA according to the environmental conditions and gene dosage. Such a mechanism would allow environmental signals to be transduced via TOR to control not only translation but also transcription (particularly of rRNA) for optimum growth of the plant under a given environment (Fig.…”

“…In yeast, the pol I-dependent transcription of 35S rRNA precursor is directly controlled by TOR, which binds to the rDNA promoter via its helix-turn-helix motif (9). More recently, association of mammalian TOR and the Arabidopsis TOR to their respective rDNA promoters has also been reported, and the binding of mammalian TOR was shown to be sensitive to rapamycin treatment (10,11). TOR has also been implicated in the transcriptional activation of a number of ribosomal protein genes that is mediated by the activities of its downstream effector kinase (ribosomal protein S6 kinase) and the c-Myc transcription factor (Sch9 in animals and Sfp1 in yeast, respectively) (3).…”

Translational Regulation of Cytoplasmic mRNAs

“…Moreover, silencing of AtTOR, or inhibition of its activity with rapamycin or torin, causes leaf senescence, arrests organ growth, and reduces ribosomal RNA synthesis and translation. In contrast, overexpression of TOR enhances growth of the root and shoot and increases seed production (Deprost et al, 2007;Ren et al, 2011;Ren et al, 2012;Xiong and Sheen, 2012;Xiong et al, 2013) consistent with a role for AtTOR in growth control.…”