The micronutrient zinc has an essential role in physiological and metabolic processes in plants as a cofactor or structural element in 300 catalytic and noncatalytic proteins, but it is very toxic when available in elevated amounts. Plants tightly regulate their internal zinc concentrations in a process called zinc homeostasis. The exceptional zinc hyperaccumulator species Thlaspi caerulescens can accumulate up to 3% of zinc, but also high amounts of nickel and cadmium, without any sign of toxicity. This should have drastic effects on the zinc homeostasis mechanism. We examined in detail the transcription profiles of roots of Arabidopsis thaliana and T. caerulescens plants grown under deficient, sufficient, and excess supply of zinc. A total of 608 zinc-responsive genes with at least a 3-fold difference in expression level were detected in A. thaliana and 352 in T. caerulescens in response to changes in zinc supply. Only 14% of these genes were also zinc responsive in A. thaliana. When comparing A. thaliana with T. caerulescens at each zinc exposure, more than 2,200 genes were significantly differentially expressed (≥5-fold and false discovery rate < 0.05). While a large fraction of these genes are of yet unknown function, many genes with a different expression between A. thaliana and T. caerulescens appear to function in metal homeostasis, in abiotic stress response, and in lignin biosynthesis. The high expression of lignin biosynthesis genes corresponds to the deposition of lignin in the endodermis, of which there are two layers in T. caerulescens roots and only one in A. thaliana.