Leveraging proteome and phosphoproteome to unravel the molecular mechanisms of legume–rhizobia symbiosis

Rapid advances in next-generation sequencing technology coupled with growing computing power have enabled the development of several high-throughput “omic” approaches over the last two decades. Prime among these approaches are quantitative proteomics and phosphoproteomics, which deal with levels of protein and post-translational protein modifications, respectively. The field of plant proteomics is relatively young compared to that of animal proteomics; nevertheless, its impact in the field of plant sciences cannot be overstated. The inherent complexity of the plant tissue, including the cell wall, phenolic compounds, and various pigments, has made sample preparation for plant proteomics and phosphoproteomics challenging. Nevertheless, new techniques have been developed, and techniques for microbial and animal proteomics have been successfully adapted and modified for studying plant proteins. These proteomic and phosphoproteomic approaches have not only provided insight into global proteome changes during plant development and stress tolerance (both biotic and abiotic) but also provided invaluable clues about the potential regulatory mechanisms governing such processes. The legume community has also applied these proteomic techniques to investigate several aspects of legume biology, including the establishment and development of legume–rhizobia symbiosis. In this chapter, decades of technological advances in the Medicago truncatula proteome and phosphoproteome utilizing both non-targeted and targeted (selected reaction monitoring, SRM) approaches are discussed. We also discuss the salient features of the M. truncatula protein atlas, M. truncatula acetylome, and proteogenomic survey, which was performed utilizing the large proteomic and phosphoproteomic databases. We believe that these high-throughput proteomic approaches will enable the community not only to identify new components in the early symbiotic signaling pathway but also understand the late stages of symbiotic nodule development and function. These evolving technologies will further our understanding of symbiotic associations in legumes and bring us one step closer to the “holy grail” of engineering nitrogen-fixing root nodule symbiosis in non-leguminous crops. These perspectives are discussed along with applications and future directions of M. truncatula proteomic and phosphoproteomic studies in this chapter.