Zinc (Zn2+) is an important trace metal ion that has been shown to regulate the expression of several (virulence) genes in streptococci. Previously, we analyzed the genome-wide response of S. pneumoniae to Zn2+-stress. In this work, we have performed a transcriptomic analysis to identify genes that are differentially expressed under intracellular Zn2+ limitation. This revealed a number of genes that are highly upregulated in the absence of extracellular Zn2+, amongst which the genes belonging to the regulon of the Zn2+-responsive repressor AdcR, like adcBCA, encoding a Zn2+-dependent ABC-uptake system, adcAII, encoding a Zn2+-binding lipoprotein, and also virulence genes belonging to the Pht family (phtA, phtB, phtD and phtE). Using transcriptome analysis, lacZ-reporter studies, in vitro DNA binding experiments, and in silico operator predictions, we show that AdcR directly represses the promoters of adcRCBA, adcAII-phtD, phtA, phtB and phtE in the presence of Zn2+. AdcR can also function as an activator, since in the presence of Zn2+ it directly induces expression of adh that encodes a Zn2+-containing alcohol dehydrogenase. In conclusion, the genome-wide transcriptional response of S. pneumoniae to Zn2+ limitation was established, which is mainly mediated via direct regulation by the Zn2+-dependent regulator AdcR.