Comparative analysis of salt‐responsive phosphoproteins in maize leaves using Ti⁴⁺‐IMAC enrichment and ESI‐Q‐TOFMS

Abstract

Salinity is one of the most common abiotic stresses encountered by plants. Reversible protein phosphorylation is involved in plant defense processes against salinity stress. Here, we performed global phosphopeptide mapping through enrichment by our synthesized PVA-phosphate-Ti(4+) IMAC coupled with subsequent identification by ESI-Q-TOF MS. A total of 104 peptide sequences containing 139 phosphorylation sites were determined from 70 phosphoproteins of the control leaves. In contrast, 124 phosphopeptides containing 143 phosphorylated sites from 92 phosphoproteins were identified in salt-stressed maize leaves. Compared with the control, 47 proteins were phosphorylated, 25 were dephosphorylated, and 45 overlapped. Among the 72 differential phosphoproteins, 35 were known salt stress response proteins and the rest had not been reported in the literature. To dissect the differential phosphorylation, gene ontology annotations were retrieved for the differential phosphoproteins. The results revealed that cell signaling pathway members such as calmodulin and 14-3-3 proteins were regulated in response to 24-h salt stress. Multiple putative salt-responsive phosphoproteins seem to be involved in the regulation of photosynthesis-related processes. These results may help to understand the salt-inducible phosphorylation processes of maize leaves.

“Omics” of Maize Stress Response for Sustainable Food Production: Opportunities and Challenges

“…14-3-3 protein levels were found to decrease in roots and to be phosphorylated in leaves (Hu et al, 2013) under salt stress. The changes in 14-3-3 proteins may affect the plasma membrane ATPase activity and lead to a change in the apoplastic pH in response to phosphorylation.…”

“…The changes in 14-3-3 proteins may affect the plasma membrane ATPase activity and lead to a change in the apoplastic pH in response to phosphorylation. Another salt-responsive phosphoproteomic analysis has been performed in maize leaves using Ti 4 + -IMAC enrichment and ESI-Q-TOF MS (Hu et al, 2013). Seven novel phosphoproteins were identified: phototropin-1 (a translationally controlled tumor protein homolog), rad23 (a DNA repair protein), protein phosphatase inhibitor 2-containing protein, pyruvate orthophosphate dikinase, p-pyruvate carboxylase, and dehydrin, which seem to be involved in the regulation of photosynthesis-related processes.…”

“…PTMs are important for regulating protein function, subcellular localization, and protein stability. Thus far, the only PTM that has been studied is protein phosphorylation during maize salt stress (Hu et al, 2013;Zörb et al, 2010) and mechanical wounding (Lewandowska-Gnatowska et al, 2011).…”

Molecular response of canola to salt stress: insights on tolerance mechanisms

“…PTMs regulate protein function, subcellular localization, correct folding, and stability (Gong et al 2015;Wu et al 2016). The studies on PTMs of stressed plants show that the protein phosphorylation and glycosylation are accelerated under stressful conditions (Hu et al 2013;Mustafa & Komatsu 2014). Moreover, PTMs have commonly been observed in gel-based proteomic studies, in which one protein presents in more than one locations (spots) on the gel (Bandehagh et al 2011;Gharelo-Shokri et al 2016;Mann & Jensen 2003).…”

“…They compared the increases between second leaf (young leaves) and third leaf (older leaves) and conclude that the younger leaves show significantly more increase in their salt-responsive proteins. Another study, by Hu et al (2013), indicated that the expression levels of BnBDC1, BnLEA4, BnMPK3, and BnNAC2 are upregulated 1h after salt stress, while their expression is downregulated 3, 6, and 12h after the stress. Jia et al (2015) …”