Fluid shear stress is crucial for maintenance of a properly functioning endothelium. In this study we demonstrate that the KLF2 transcription factor is greatly induced by pulsatile shear stress in murine microvascular endothelial cells. The promoter elements responsible for the induction were studied by transfection with luciferase-reporter plasmids including the 5′ flanking region of the murine KLF2 gene. Deletion analysis reveals that the responses are regulated by a region from -157 to -95 bp from the start site of transcription. Furthermore, shear stress induces specific nuclear binding within this region. These results define a novel shear stress response region that is highly conserved between mouse and human homologs.