The study was aimed to investigate by differential proteomics heat stress response mechanisms in tomato anthers collected from flowers of Solanum lycopersicum cv Saladette (SAL), already annotated as a thermo-tolerant tomato genotype and flowers from Solanum lycopersicum cv M82, reported as a thermo-sensitive genotype. Proteins constitutively present at higher or lower level in the tolerant genotype and/or proteins whose abundance was modulated by growth under high temperature in both genotypes were identified, thus leading to define processes involved in the heat stress response and responsible for efficient reduction of the adverse effects of HS. Results showed differences in the abundance of ninety-six proteins and their functional classification highlighted that heat stress mainly affected metabolic pathways, such as energy metabolism (glycolysis and sucrose degradation), nitrogen assimilation and amino acid biosynthesis and modulated the expression of proteins involved in the folding and degradation machinery and ROS detoxification systems