The spaceflight environment presents unique challenges to terrestrial biology, including but not limited to the direct effects of gravity. As we near the end of the Space Shuttle era, there remain fundamental questions about the response and adaptation of plants to orbital spaceflight conditions. We address a key baseline question of whether gene expression changes are induced by the orbital environment, and then we ask whether undifferentiated cells, cells presumably lacking the typical gravity response mechanisms, perceive spaceflight. Arabidopsis seedlings and undifferentiated cultured Arabidopsis cells were launched in April, 2010, as part of the BRIC-16 flight experiment on STS-131. Biologically replicated DNA microarray and averaged RNA digital transcript profiling revealed several hundred genes in seedlings and cell cultures that were significantly affected by launch and spaceflight. The response was moderate in seedlings; only a few genes were induced by more than 7-fold, and the overall intrinsic expression level for most differentially expressed genes was low. In contrast, cell cultures displayed a more dramatic response, with dozens of genes showing this level of differential expression, a list comprised primarily of heat shock-related and stress-related genes. This baseline transcriptome profiling of seedlings and cultured cells confirms the fundamental hypothesis that survival of the spaceflight environment requires adaptive changes that are both governed and displayed by alterations in gene expression. The comparison of intact plants with cultures of undifferentiated cells confirms a second hypothesis: undifferentiated cells can detect spaceflight in the absence of specialized tissue or organized developmental structures known to detect gravity.