Down syndrome occurs due to triplication of human chromosome 21, and results in many phenotypic defects. Here we focus on the role of adhesions in this disorder, which are involved in tissue development, cell migration, and memory formation. Previous work demonstrates that fibroblasts from individuals with Down syndrome have increased focal adhesions. We also found that the area, perimeter and motility of Down syndrome fibroblasts is altered. Here we show that RACK1, a scaffolding protein and member of the focal adhesion complex, is overexpressed in Down syndrome fibroblasts and can be knocked down with shRNA in Down syndrome fibroblasts to levels comparable to apparently healthy control cells. Examining the role of RACK1 in Down syndrome is relevant for providing insight into therapeutic and clinical treatments.