The riboflavin kinase encoding gene ribR is situated within a 12 genes locus ytmI-ytnM of the Bacillus subtilis chromosome. Here we demonstrate that ribR is transcribed as part of a 10 kb ytmI-ytnM operon. The riboflavin overproduction phenotype of B. subtilis ribC mutant strains, which is a result of the strongly reduced flavokinase activity of the riboflavin kinase/FAD synthetase RibC, was suppressed by ribR expression. Analysis of mutations with an upregulated ribR gene revealed 2 different groups of mutants. One class of mutants contained base substitutions in an 8 nucleotide sequence of the promoter region of the ytmI-ytnM operon. A second class of mutants had single point mutations within the yrzC gene or in the RBS of this gene. Dot-blot analysis of ytmI-ytnM transcription and the results of in trans complementation experiments for the yrzC mutants confirmed a role of the yrzC gene product as a negative regulator for the ytmI-ytnM operon.