Phosphorylation of Ribosomal Protein RPS6 Integrates Light Signals and Circadian Clock Signals

Abstract

The translation of mRNA into protein is tightly regulated by the light environment as well as by the circadian clock. Although changes in translational efficiency have been well documented at the level of mRNA-ribosome loading, the underlying mechanisms are unclear. The reversible phosphorylation of RIBOSOMAL PROTEIN OF THE SMALL SUBUNIT 6 (RPS6) has been known for 40 years, but the biochemical significance of this event remains unclear to this day. Here, we confirm using a clock-deficient strain of Arabidopsis thaliana that RPS6 phosphorylation (RPS6-P) is controlled by the diel light-dark cycle with a peak during the day. Strikingly, when wild-type, clock-enabled, seedlings that have been entrained to a light-dark cycle are placed under free-running conditions, the circadian clock drives a cycle of RPS6-P with an opposite phase, peaking during the subjective night. We show that in wild-type seedlings under a light-dark cycle, the incoherent light and clock signals are integrated by the plant to cause an oscillation in RPS6-P with a reduced amplitude with a peak during the day. Sucrose can stimulate RPS6-P, as seen when sucrose in the medium masks the light response of etiolated seedlings. However, the diel cycles of RPS6-P are observed in the presence of 1% sucrose and in its absence. Sucrose at a high concentration of 3% appears to interfere with the robust integration of light and clock signals at the level of RPS6-P. Finally, we addressed whether RPS6-P occurs uniformly in polysomes, non-polysomal ribosomes and their subunits, and non-ribosomal protein. It is the polysomal RPS6 whose phosphorylation is most highly stimulated by light and repressed by darkness. These data exemplify a striking case of contrasting biochemical regulation between clock signals and light signals. Although the physiological significance of RPS6-P remains unknown, our data provide a mechanistic basis for the future understanding of this enigmatic event.

Thermal imaging as a noninvasive technique for analyzing circadian rhythms in plants

“…Since the leaf temperature rhythms we observed were under circadian control, we expected that they would be affected by mutations in key oscillator genes. Overexpression of CCA1 ( CCA1 ‐ox) has been reported to cause the loss or attenuation of normal circadian rhythms (Wang & Tobin, ; Daniel et al ., ; Dakhiya et al ., ; Enganti et al ., ). Consistent with previously published reports (Green et al ., ), CCA1 ‐ox plants were still able to respond directly to environmental cues; Fig.…”

“…), suggesting that CCA1 ‐ox plants retain some timing mechanism. The retention of attenuated thermal rhythms in CCA1 ‐ox plants is consistent with data presented in several previous studies showing weak rhythmicity in CCA1 ‐ox plants even in free‐running conditions (Daniel et al ., ; Dakhiya et al ., ; Enganti et al ., ).…”

ErbB-3 BINDING PROTEIN 1 Regulates Translation and Counteracts RETINOBLASTOMA RELATED to Maintain the Root Meristem

“…S6 kinase (S6K) is a key downstream component of TOR signaling that regulates translation and ribosome biogenesis (Deprost et al, 2007;Schepetilnikov et al, 2013). Thus growth-stimulatory factors such as light, sugar, and auxin activate TOR and affect the translation apparatus in different ways (Liu et al, 2013;Dobrenel et al, 2016b;Chen et al, 2018;Enganti et al, 2018;Schepetilnikov and Ryabova, 2018). Correspondingly, elevated TOR expression boosts plant growth in Arabidopsis (Arabidopsis thaliana; Deprost et al, 2007) and increases yield in crop plants, specifically in limiting conditions (Bakshi et al, 2019).…”

Light dependent activation of the GCN2 kinase under cold and salt stress is mediated by the photosynthetic status of the chloroplast

“…Absorbance at 254 nm was recorded using an ISCO UA 5 absorbance/fluorescence monitor and individual data points were extracted using the DATA acquisition software (DATAQ instruments). Polysome-to-monosome (P/M) ratios were calculated as described (Enganti et al, 2018). For polysome profiling with salt stressed tissue, 150 mg of tissue powder was resuspended in 100μl of polysome extraction buffer and centrifuged at 13,000 rpm for 5 minutes at 4 °C.…”