Sulfur mustard (SM) is an alkylating agent that has been used as a chemical warfare agent in numerous conflicts around the world. The lung is a primary target of SM exposure. To understand the mechanism of SM-induced lung injury we analyzed global changes in gene expression in a rat lung SM exposure model. Male Sprague Dawley rats were injected in the femoral vein with liquid SM, which circulates directly to the pulmonary vein and to the lung for reoxygenation of blood. This results in a deep lung SM exposure. Rats were exposed to 1 mg, 3 mg or 6 mg of SM and lung tissue was harvested at 0.5, 1, 3, 6 and 24 hours post-injection. RNA was extracted from the lung and used as starting material for the probing of oligonucleotide microarrays. The gene expression data were analyzed using principal component analysis and two-way analysis of variance to identify the genes most significantly changed as a result of variation over time and dose. These genes were rank ordered by p-value and categorized based on molecular function and biological process. Previously, inflammation has been shown to play a role in SM injury and our data corroborate these findings. However, the most significant changes in gene expression reflect alterations in the cell cycle and vesicular trafficking. In particular, the changes in cell cycle reflect a potential cell cycle block at the G1/S transition due to activation of the p53 pathway. Although our data do not determine directly the activation of p53, many of the genes that are significantly upregulated in a dose dependent fashion have been shown by other investigators to be p53 responsive genes. Thus, SM exposure appears to induce a robust p53 response in the lung.