Prx1 (MHox) and Prx2 (S8) are non-clustered homeobox genes that are expressed in a complex, mostly mesenchyme-specific pattern throughout embryogenesis. The expression pattern and gene-targeted mice previously revealed a major role for Prx1 in skeletogenesis. In addition, specific and high expression of both Prx genes was reported in the developing cardiovascular system, predominantly in prospective connective tissues of the heart and in the great arteries and veins. We examined embryos of previously generated gene-targeted mice. Prx2-/- mutants were viable and did not show cardiovascular malformations. Intracardiac morphology of Prxl-/- and Prx1/Prx2-combined null mutants also appeared normal throughout development. However, the Prx1-/- and Prx1/Prx2 double-null mutants showed a vascular abnormality with an abnormal positioning and awkward curvature of the aortic arch in addition to a misdirected and elongated ductus arteriosus, and in two of seven combined mutants, an anomalous retro-oesophageal right subclavian artery. Generally, all great arteries appeared to run somewhat tortuously through the surrounding mesenchyme. The vascular histology and vessel wall thickness were normal in all mutants. Prx1-/- and Prx double-gene-targeted mice revealed similar spectra of vascular anomalies, but double mutants appeared to be more seriously affected. The current findings suggest that other genes may compensate for the loss of Prx in the heart, but, in contrast, our data support a role for Prx in the development of vascular and perivascular matrix.