Downstream targets of the metabolic stress sensor SnRK1 - One kinase to rule them all

The evolutionarily conserved SnRK1 (SNF1-related kinase 1) protein kinase (the ortholog of yeast SNF1 and mammalian AMPK) is a major metabolic sensor, allowing acclimatization and energy homeostasis upon changes in energy availability, thereby ensuring plant survival. SnRK1 is activated by low energy stress and repressed by high photosynthetic sugar levels, with the allosteric inhibitor trehalose-6-P acting as a proxy for sucrose supply. To maintain energy homeostasis during energy-depleting stress conditions, SnRK1 not only induces catabolic processes (recovering C and energy from alternative sources), but also suppresses nonessential energy-consuming anabolic processes to redirect the C and energy fluxes to processes vital for plant survival. In this work, we explored plant growth and anthocyanin biosynthesis as energy-intensive processes directly repressed by SnRK1 activity in Arabidopsis thaliana. As a case study of redirection of resources to increased (a)biotic stress tolerance, we identified Plasmodiophora brassicae(clubroot disease) resistance in transgenic lines with increased SnRK1 activity and programmed cell death as a positively regulated target process in the plant’s immune response.

Phenotyping of transgenic Arabidopsis lines with altered SnRK1 activity suggests a role for SnRK1 in growth rate control via repression of cell division activity and turgor-driven cell expansion. Cellular assays with transient overexpression of the cell cycle regulatory machinery in leaf mesophyll protoplasts identified different direct SnRK1 targets involved in the G1/S and G2/M phase transitions, consistent with a novel and perhaps counterintuitive role for SnRK1 as positive regulator of endoreduplication.

Analysis of mutant and transgenic Arabidopsis lines and cellular assays with transient overexpression in leaf mesophyll protoplasts also confirmed SnRK1 as a negative regulator of anthocyanin biosynthesis and identified MYB75transcription factorexpression and MYB75/bHLH/TTG1 (MBW) transcription factorcomplex association and stability as its direct targets. Both the transcriptional and post-translational regulation appear to be mediated by TCP-type transcription factors. Consistently, we also identified these plant-specific transcription factors as part of the SnRK1 signaling network using a forward genetics approach with a tps1mutant suppressor screen based on the restoration of sucrose-induced anthocyanin accumulation and root growth.

Finally, we assessed whether modification of SnRK1 activity is a viable strategy to broadly and more sustainably increase biotic stress tolerance using Plasmodiophora brassicae infection, causing clubroot disease in crucifer species (Brassicaceae). Preliminary qualitative phenotyping indeed showed an increased tolerance of Arabidopsis plants upon overexpression of the SnRK1 catalytic subunit, SnRK1α1, without a significant effect on seed yield and total seed fatty acid content, economically important traits for closely related oleaginous Brassica crops such as rapeseed.

Resistance to clubroot infection in some cases coincides with the onset of a hypersensitive response and several reports point to SnRK1 as a positive regulator of the associated programmed cell death. Using cellular assays, we identified the LSD1/bZIP10 pathway as a likely direct positively regulated target of SnRK1, possibly stimulating cell death upon pathogen recognition or pathogen-dependent energy depletion.

In conclusion, the results obtained in this work contribute to our understanding of the processes targeted by the metabolic stress sensor SnRK1 upon carbon and energy depletion. Detailed elucidation of the downstream signaling network will enable more directed selection and precision breeding of crops with increased yields and enhanced performance in stressful and increasingly unpredictable environments.