The β-galactoside-binding protein galectin-3 (gal-3) has pleitropic biological functions and has been implicated in cell growth, differentiation, adhesion, RNA processing, apoptosis, and malignant transformation. To investigate the pattern of inactivation of the gal-3 gene (LGALS3) in colorectal cancers (CRC), we studied a series of Tunisian patients with CRC to identify abnormal methylation in LGALS3 promoter using a methylation-specific PCR. We also examined the gal-3 gene expression by reverse transcription-PCR and the expression of gal-3 protein by immunohistochemistry. Analysis of DNA methylation in nonmucinous colorectal carcinomas expressing gal-3 protein showed an unmethylated profile of LGALS3 promoter, whereas gal-3 was aberrantly methylated in mucinous colorectal carcinomas. Complete loss of the gal-3 expression both at mRNA and the protein level was associated with the gal-3 methylation in the mucinous colorectal carcinomas. Our results show that methylation of the gal-3 promoter could be an important mechanism in the regulation of the expression of this gene in mucinous CRCs.