A sudden decrease in ambient temperature induces the expression of a number of genes in poikilothermic organisms. We report here that the cold inducibility of gene expression in Synechocystis sp. PCC 6803 was enhanced by the rigidification of membrane lipids that was engineered by disruption of genes for fatty acid desaturases. DNA microarray analysis revealed that cold-inducible genes could be divided into three groups according to the effects of the rigidification of membrane lipids. The first group included genes whose expression was not induced by cold in wild-type cells but became strongly cold-inducible upon rigidification of membrane lipids. This group included certain heat-shock genes, genes for subunits of the sulfate transport system, and the hik34 gene for a histidine kinase. The second group consisted of genes whose cold inducibility was moderately enhanced by the rigidification of membrane lipids. Most genes in this group encoded proteins of as yet unknown function. The third group consisted of genes whose cold inducibility was unaffected by the rigidification of membrane lipids. This group included genes for an RNA helicase and an RNA-binding protein. DNA microarray analysis also indicated that the rigidification of membrane lipids had no effect on the heat inducibility of gene expression. Hik33, a cold-sensing histidine kinase, regulated the expression of most genes in the second and third groups but of only a small number of genes in the first group, an observation that suggests that the cold-inducible expression of genes in the first group might be regulated by a cold sensor that remains to be identified.