We have addressed the distribution and abundance of 75 transcription factor (TF) families in complete genomes from 90 different bacterial and archaeal species. We found that the proportion of TFs increases with genome size. The deficit of TFs in some genomes might be compensated by the presence of proteins organizing and compacting DNA, such as histone-like proteins. Nine families are represented in all the bacteria and archaea we analyzed, whereas 17 families are specific to bacteria, providing evidence for regulon specialization at an early stage of evolution between the bacterial and archeal lineages. Ten of the 17 families identified in bacteria belong exclusively to the proteobacteria defining a specific signature for this taxonomical group. In bacteria, 10 families are lost mostly in intracellular pathogens and endosymbionts, while 9 families seem to have been horizontally transferred to archaea. The winged helix-turn-helix (HTH) is by far the most abundant structure (motif) in prokaryotes, and might have been the earliest HTH motif to appear as shown by its distribution and abundance in both bacterial and archaeal cellular domains. Horizontal gene transfer and lineage-specific gene losses suggest a progressive elimination of TFs in the course of archaeal and bacterial evolution. This analysis provides a framework for discussing the selective forces directing the evolution of the transcriptional machinery in prokaryotes.