The hig (host inhibition of growth) genes of plasmid Rts1 belong to the plasmid-encoded proteic killer gene family. Compared with other proteic killer genes described so far, hig is unique in that the toxic part (higB) exists upstream of the antidote gene (higA). Here we describe results of the promoter analysis of hig genes together with identification of the proteic gene products of higA and higB. Two promoters were identified in the hig locus; a stronger one, named Phig, is located upstream of higB and a weaker one, PhigA, is upstream of higA within the higB coding region. The Phig activity was negatively regulated by HigA and this regulation was augmented by HigB, whereas PhigA was not subjected to such a regulation.