Mimecan is a small leucine-rich proteoglycan that can occur as either keratan sulfate proteoglycan in the cornea or as glycoprotein in many connective tissues. As yet, there is no information on its transcriptional regulation. Recently we demonstrated the presence of eight mimecan mRNA transcripts generated by alternative transcription initiation, alternative polyadenylation, and differential splicing, all of which encode an identical protein. Here we report a conserved consensus p53-binding DNA sequence in the first intron of bovine and human mimecan genes and show that wild-type p53 binds to this sequence in vitro. Co-transfections of Saos-2, HeLa, NIH 3T3, and primary bovine corneal keratocytes with bovine mimecan promoter/luciferase reporter constructs in combination with p53 expression vectors activate the second mimecan promoter through the p53-binding sequence. In addition, we show absence of mimecan expression in different tumors and cancer cell lines, where p53 frequently is inactivated/mutated. Thus, this work provides novel information that links mimecan to the p53 network.