Protein profile analysis of salt-responsive proteins in leaves and roots in two cultivars of creeping bentgrass differing in salinity tolerance

Knowledge of stress-responsive proteins is critical for further understanding the molecular mechanisms of stress tolerance. The objectives of this study were to establish a proteomic map for a perennial grass species, creeping bentgrass (A. stolonifera L.), and to identify differentially expressed, salt-responsive proteins in two cultivars differing in salinity tolerance. Plants of two cultivars (‘Penncross’ and ‘Penn-A4’) were irrigated daily with water (control) or NaCl solution to induce salinity stress in a growth chamber. Salinity stress was obtained by adding NaCl solution of 2, 4, 6, and 8 dS m⁻¹ in the soil daily for 2-day intervals at each concentration, and then by watering soil with 10 dS m⁻¹ solution daily for 28 days. For proteomic map, using two-dimensional electrophoresis (2-DE), approximately 420 and 300 protein spots were detected in leaves and roots, respectively. A total of 148 leaf protein spots and 40 root protein spots were excised from the 2-DE gels and subjected to mass spectrometry analysis. In total, 106 leaf protein spots and 24 root protein spots were successfully identified. Leaves had more salt-responsive proteins than roots in both cultivars. The superior salt tolerance in ‘Penn-A4’, indicated by shoot extension rate, relative water content, and cell membrane stability during the 28-day salinity stress could be mainly associated with its higher level of vacuolar H⁺-ATPase in roots and UDP-sulfoquinovose synthase, methionine synthase, and glucan exohydrolase in leaves, as well as increased accumulation of catalase and glutathione S-transferase in leaves. Our results suggest that salinity tolerance in creeping bentgrass could be in part controlled by an alteration of ion transport through vacuolar H⁺-ATPase in roots, maintenance of the functionality and integrity of thylakoid membranes, sustained polyamine biosynthesis, and by the activation of cell wall loosening proteins and antioxidant defense mechanisms.