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Parallel global profiling of plant TOR dynamics reveals a conserved role for LARP1 in protein translation

ABSTRACT: TARGET OF RAPAMYCIN (TOR) is a deeply conserved protein kinase that coordinates eukaryotic metabolism with nutrient availability. In mammals, TOR specifically promotes translation of ribosomal protein mRNAs when amino acids are available to support protein synthesis. The mechanisms controlling translation downstream from TOR remain contested, however, and are largely unexplored in plants. Here, we took parallel global profiling approaches to define the in planta TOR-regulated proteome, and phosphoproteome. We found that TOR regulates ribosome biogenesis in plants at multiple levels. To investigate this further, we focused on a putative TOR substrate identified in our phosphoproteome: LARP1, a eukaryotic RNA-binding protein that is proposed to mediate TOR translational control of 5′TOP mRNAs in humans.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Arabidopsis Thaliana (mouse-ear Cress)

TISSUE(S): Plant Cell, Seedling

SUBMITTER: Maria Regina Scarpin

LAB HEAD: Jacob O. Brunkard

PROVIDER: PXD019904 | Pride | 2021-01-05

REPOSITORIES: Pride

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	20190412_Berkeley_Exp8_Global_TMT1_F01.raw	Raw
	20190412_Berkeley_Exp8_Global_TMT1_F02.raw	Raw
	20190412_Berkeley_Exp8_Global_TMT1_F03.raw	Raw
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	20190412_Berkeley_Exp8_Global_TMT1_F05.raw	Raw

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Parallel global profiling of plant TOR dynamics reveals a conserved role for LARP1 in protein translation

Project description:TARGET OF RAPAMYCIN (TOR) is a deeply conserved protein kinase that coordinates eukaryotic metabolism with nutrient availability. In mammals, TOR specifically promotes translation of ribosomal protein mRNAs when amino acids are available to support protein synthesis. The mechanisms controlling translation downstream from TOR remain contested, however, and are largely unexplored in plants. Here, we took parallel global profiling approaches to define the in planta TOR-regulated proteome, and phosphoproteome. We found that TOR regulates ribosome biogenesis in plants at multiple levels. To investigate this further, we focused on a putative TOR substrate identified in our phosphoproteome: LARP1, a eukaryotic RNA-binding protein that is proposed to mediate TOR translational control of 5′TOP mRNAs in humans.

2021-01-05 | PXD019942 | Pride

eIF4A1 enhances LARP1-mediated translational repression during mTORC1 inhibition

Project description:Eukaryotic translation initiation factor (eIF) 4A — a DEAD-box RNA-binding protein — plays an essential role in translation initiation. Recent reports have suggested helicase-dependent and helicase-independent functions for eIF4A, but the multifaceted roles of eIF4A have not been fully explored. Here, we show that eIF4A1 enhances translational repression during the inhibition of mechanistic target of rapamycin complex 1 (mTORC1), an essential kinase complex controlling cell proliferation. RNA pulldown followed by sequencing revealed that eIF4A1 preferentially binds to mRNAs containing terminal oligopyrimidine (TOP) motifs (TOP mRNAs), whose translation is rapidly repressed upon mTORC1 inhibition. This selective interaction depends on a La-related RNA-binding protein, LARP1. Ribosome profiling revealed that deletion of EIF4A1 attenuated the translational repression of TOP mRNAs upon mTORC1 inactivation. Moreover, eIF4A1 increases the affinity between TOP mRNAs and LARP1 and thus ensures stronger translational repression upon mTORC1 inhibition. Our data show the multimodality of eIF4A1 in modulating protein synthesis through an inhibitory binding partner and provide a unique example of the repressive role of a universal translational activator.

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Nitrogen starvation leads to TOR kinase-mediated downregulation of fatty acid synthesis in the algae Chlorella sorokiniana and Chlamydomonas reinhardtii

Project description:The accumulation of triacylglycerol (TAG) as a storage compound in eukaryotic algae has been the subject of extensive studies over the last 25 years. The model industrial alga Chlorella sorokiniana accumulates TAG and other storage compounds under nitrogen (N)-limited growth. Previously we used transcriptomics to explore the regulation of TAG synthesis in C. sorokiniana. Surprisingly, our analysis showed that the expression of several key genes encoding enzymes involved in plastidic fatty acid synthesis are significantly repressed. Metabolic labeling with radiolabeled acetate showed that de novo fatty acid synthesis is indeed downregulated under N-limitation. Likewise, inhibition of the Target of Rapamycin kinase (TOR), a key regulator of metabolism and growth, decreased fatty acid synthesis. We compared the changes induced in the quantitative proteome and phosphoproteome of cells under N-limitation or TOR inhibition and found extensive overlap between the N-limited and TOR-inhibited conditions. We also detected variations in the phosphorylation of TOR complex proteins under N-limitation, indicating that TOR signaling is altered. Our results indicate that under N-limitation there is significant metabolic remodeling, including fatty acid synthesis, mediated by TOR signaling. We find that TOR-mediated metabolic remodeling of fatty acid synthesis under N-limitation is conserved in the chlorophyte algae Chlorella sorokiniana and Chlamydomonas reinhardtii.

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Project description:To shed light on the TOR signaling network in crop plants, we investigated the global transcriptome changes in rice suspension cells (cultivar Kitaake) treated with the TOR-specific inhibitor rapamycin.

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