Temperature is an important factor influencing the distribution of marine picocyanobacteria. However, molecular responses contributing to temperature preferences are poorly understood in these important primary producers. We compared the temperature acclimation of a tropical Synechococcus strain WH8102 with temperate strain BL107 at 18 °C relative to 22 °C and examined their global protein expression, growth patterns, photosynthetic efficiency and lipid composition. Global protein expression profiles demonstrate the partitioning of the proteome into major categories: photosynthesis (>40%), translation (10-15%) and membrane transport (2-8%) with distinct differences between and within strains grown at different temperatures. At low temperature, growth and photosynthesis of strain WH8102 was significantly decreased, while BL107 was largely unaffected. There was an increased abundance of proteins involved in protein biosynthesis at 18 °C for BL107. Each strain showed distinct differences in lipid composition with higher unsaturation in strain BL107. We hypothesize that differences in membrane fluidity, abundance of protein biosynthesis machinery and the maintenance of photosynthesis efficiency contribute to the acclimation of strain BL107 to low temperature. Additional proteins unique to BL107 may also contribute to this strain's improved fitness at low temperature. Such adaptive capacities are likely important factors favoring growth of temperate strains over tropical strains in high latitude niches.