Quantitative ubiquitylomics approach for characterizing the dynamic change and extensive modulation of ubiquitylation in rice seed germination

Abstract

Summary During seed germination, cells embark on extensive post‐transcriptional and post‐translational modifications (PTM), providing a perfect platform to study these events in embryo rebooting from relative quiescenct to highly active state. PR‐619, a deubiquitylase inhibitor, delayed the rice seed germination and resulted in the accumulation of ubiquitylated proteins, which indicated the protein ubiquitylation is involved in this process. Using the K‐Ɛ‐GG antibody enrichment method integrated with high‐resolution mass spectrometry, a list of 2576 lysine ubiquitylated (Kub) sites in 1171 proteins was compiled for rice embryos at 0, 12 and 24 h after imbibition (HAI). Of these, the abundance of 1419 Kub sites in 777 proteins changed significantly. Most of them substantially increased within the first 12 HAI, which is similar to the dynamic state previously observed for protein phosphorylation, implying that the first 12 HAI are essential for subsequent switch during rice seed germination. We also quantitatively analyzed the embryo proteome in these samples. Generally, a specific protein’s abundance in the ubiquitylome was uncorrelated to that in the proteome. The differentially ubiquitinated proteins were greatly enriched in the categories of protein processing, DNA and RNA processing/regulation related, signaling, and transport. The DiGly footprint of the Kub sites was significantly reduced on K48, a linkage typically associated with proteasome‐mediated degradation. These observations suggest ubiquitylation may modulate the protein function more than providing 26S degradation signals in the early stage of rice seed germination. Revealing this comprehensive ubiquitylome greatly increases our understanding of this critical PTM during seed germination.

An Updated Overview on the Regulation of Seed Germination

“…Protein ubiquitination is another important protein modification known to negatively affect the stability of proteins with a crucial role in germination, such as DELLAs [14,229], ABI3 [230] and ABI5 [231,232]. Changes in ubiquitination were detected in more than 1000 proteins during rice seed germination and most changes occurred at 12 hai, as observed previously for phosphorylation [233]. Sumoylation has also been demonstrated to play a role in this process by providing stability to ABI5 and MYB30 TFs.…”

Dynamic Responses of Barley Root Succinyl-Proteome to Short-Term Phosphate Starvation and Recovery

“…At the protein level, only four proteins overlapped between DFASPs and DAPs, which belonged to different metabolic processes (Supplementary Table 9). A quantitative ubiquitylomics analysis of rice seed germination revealed that protein abundance in the ubiquitylome is not correlated with that in the proteome (He et al, 2020). Consistent with this phenomenon, we did not find a significant correlation between the overlapping succinylations and proteins (Supplementary Figure 5).…”

Exogenous Gibberellin Promotes the Germination of Phellodendron Chinense Schneid Seeds by Regulating the Expression of Phytohormone Related Genes and Anabolic Proteins

“…The protein kinases [38,39], protein phosphatases, and transcription factor [40,41] classes of protein serve to regulate many physiological processes [42]. Seed germination marks the starting point of plant growth and development, which is regulated by some proteins [43]. These proteins regulate catabolism [44], energy metabolism [45], anabolism, and defense reactions [46].…”