Aldolase C mRNA is detected by Northern blot in all fetal tissues in rat; it is very abundant in the adult brain and undetectable in the other adult tissues. However, reverse transcriptase polymerase chain reaction amplification indicates that this gene is not totally repressed in these tissues. A DNase-I hypersensitivity site located in a 115-base pair proximal promoter fragment is detectable in the brain as well as in other adult tissues. Two MspI/HpaII restriction sites located at -3800 and -450 base pairs are demethylated in the brain and totally or partially methylated in other tissues. In transgenic mice, a 12.5-kilobase genomic fragment is strongly and tissue specifically expressed in different lines, with conservation of a methylation pattern similar to that of the endogenous gene. A chloramphenicol acetyltransferase gene directed by either 800 or 115 base pairs of aldolase C 5'-flanking sequences is tissue specifically expressed in transgenic mice, but the level of expression is very low. This level is greatly increased when the transgene consists of a chloramphenicol acetyltransferase hybrid gene directed by 5.5 kilobases of aldolase C 5'-flanking sequences. We propose therefore that the chromatin structure around the aldolase C promoter is accessible in fetal tissues, then remains open in the adult brain, where the gene is very active, as well as in tissues in which it is practically inactive. The specificity of expression in the brain is conferred by a short 115-base pair proximal promoter fragment that needs more upstream sequences to be fully active.