Salicylic acid influences the protease activity and posttranslation modifications of the secreted peptides in the moss Physcomitrella patens

Abstract

Plant secretome comprises dozens of secreted proteins. However, little is known about the composition of the whole secreted peptide pools and the proteases responsible for the generation of the peptide pools. The majority of studies focus on target detection and characterization of specific plant peptide hormones. In this study, we performed a comprehensive analysis of the whole extracellular peptidome, using moss Physcomitrella patens as a model. Hundreds of modified and unmodified endogenous peptides that originated from functional and nonfunctional protein precursors were identified. The plant proteases responsible for shaping the pool of endogenous peptides were predicted. Salicylic acid (SA) influenced peptide production in the secretome. The proteasome activity was altered upon SA treatment, thereby influencing the composition of the peptide pools. These results shed more light on the role of proteases and posttranslational modification in the "active management" of the extracellular peptide pool in response to stress conditions. It also identifies a list of potential peptide hormones in the moss secretome for further analysis. KEYWORDS LC-MS/MS, peptidome, Physcomitrella patens, proteases, secretome 1 | INTRODUCTION Biologically active peptides play crucial roles in plant growth and development, including response to biotic and abiotic stress conditions. 1,2 However, the number of peptides in intracellular or extracellular peptidomes as well as their diversity have been underestimated for a long time. The majority of well-studied plant bioactive peptides are generated by processing inactive protein precursors and undergo posttranslational modifications (PTMs) that are essential for their biological activities. 3 Secreted peptide hormones from the cell into the extracellular space include CLAVATA3 (CLV3)/EMBRYO SURROUNDING REGION (CLE) and the C-TERMINALLY ENCODED PEPTIDE (CEP) and cysteine-rich peptides, such as RAPID ALKALINIZATION FACTOR (RALF), which control growth and development processes, cell proliferation, and differentiation. 4,5 The release of peptides from inactive preproteins depends on the activity of specific proteases. 6 According to the structural features of their active centers, plant proteases can be divided into the following classes: serine-, cysteine-, metallo-, threonine-and aspartic-type proteases. 7 For example, the formation of AtRALF1 and AtRALF23 peptides depends on precursor cleavageby subtilisin-like proteinase S1P/SBT6.1 (serine-type), 8-10 and phytaspases (aspartic-type) were shown to cleave prosystemin in tomato. 11 In addition, proteasomes, which have caspase-like, trypsinlike, and chymotrypsin-like activities, 12-17 can be responsible for the generation of peptide pools in animal cells. 18,19 However, it is not known whether proteasomes influence peptide pools in plant cells and secretomes. Plant cells contain a mixture of 26S and 20S proteasomes that perform ubiquitin-dependent and ubiquitinindependent proteolysis. It has been shown that increased activity of the 20S pro...

Thimet Oligopeptidase Biochemical and Biological Significances: Past, Present, and Future Directions

“…With the rapid advance of nano liquid chromatography in tandem with ESI-MS/MS (LC-ESI/MS/MS), peptidomics can start to be used to identify multiple peptides in biological samples, without the need to digest the proteins and characterize the resulting peptides [76][77][78]. InPeps were further sequenced directly from the low molecular weight fraction of cells and/or tissues homogenates using liquid-chromatography in tandem with mass spectrometry (LC-MS/MS), without the need of the substrate capture assay [38,[79][80][81][82][83][84]. To date, multiple research groups have identified InPeps in different human cell lines such as HEK293, MCF7 and SH-SY5Y [85], human tissues [86,87], rodents [76][77][78][88][89][90], zebrafish [91], yeast [81], and plants [82,83], even without using the "substrate capture assay" [38,79,90].…”

Peptidome: Chaos or Inevitability

“…However, the rapid development of modern mass-spectrometry analysis coupled with the explosive growth of genetic data banks has led to the considerable expansion of the list of characterized native peptidomes. Tens of thousands of peptides that significantly differ in their origin, function and properties have been identified in the tissue and biological fluids of multiple organisms [ 10 , 11 , 12 , 13 , 14 , 15 , 16 , 17 ]. Even though the peptidomes of prokaryotic and eukaryotic cells comprise thousands of peptides, the majority of them are generated during protein degradation [ 18 ].…”

“…In this review, we summarize our knowledge about the generation of different types of peptides, their precursors and biological function. In addition, we analyzed selected peptidome datasets ( Table 1 ) from plants [ 15 , 16 , 62 ], bacteria [ 63 ], humans [ 12 , 64 ] and yeast [ 65 ] to identify some common trends in their composition and physicochemical properties.…”

The Role of Peptide Signals Hidden in the Structure of Functional Proteins in Plant Immune Responses

“…A recent study by Filippova et al showed that a substantial portion (about 10%) of the moss Physcomitrella patens secretome peptides contains modifications, for example, hydroxylated proline. The authors of this study also suggest that such peptides might be generated inside the cell and then released via different pathways into the extracellular space or might be cleaved out from their precursors in the extracellular space [24]. However, the role of intracellular and extracellular peptide pools in plant immune response is still poorly understood.…”

“…The most abundant types of proteases shaping the extracellular pool of endogenous peptides in the moss Physcomitrella patens are serine-type (subtilisin-like) proteases, mealloproteases, and aspartic-type proteases. Treatment with salicylic acid leads to an increase in the number of endogenous peptides potentially cleaved by serine-type and aspartic-type proteases [24]. Evidence is emerging about the important role of the plant ubiquitin—proteasome system (UPS) in defense and stress hormone turnover regulation [33,34].…”