Identification and sequence analysis of a silent gene (ushA0) in Salmonella typhimurium

The evolution of new or improved enzyme specificities in prokaryotes has been proposed to involve gene duplication, followed by silencing of one of the duplicates at the transcriptional or translational level. Such "silent gene intermediates" are distinct from "cryptic" genes, which are proposed to have a different role in evolution. We describe the identification in Salmonella typhimurium of a silent gene (ushA0) using the active (homologous) ushA gene (encoding UDP-sugar hydrolase) from Escherichia coli as a probe. The ushA0 gene has been cloned and, in the multicopy state, very weak expression can be detected; the gene product was shown to be immunologically and functionally related to the enzyme from E. coli. The sequence of the ushA0 gene was found to be highly homologous to the previously determined sequence of the ushA gene, and the respective promoter and ribosomal-binding sites are also very similar. However, a presumed strong rho-independent terminator in the ushA gene is absent from ushA0; although a weak stem-and-loop structure is present in the 3' region of ushA0, its structure is atypical of rho-independent terminators. The sequence analysis also revealed an insertion-sequence like sequence at the 3' end of ushA0 with a convergent open reading frame terminating 116 base-pairs from the ushA0 stop codon. A deletion of the 5' region of the open reading frame results in increased expression of ushA0, indicating that convergent transcription plays some role in the silencing of ushA0. S. typhimurium contains a UDP-sugar hydrolase, biochemically and genetically distinct from that in E. coli, encoded by the ushB gene. Our results indicate that ushB is not strongly sequence-related to ushA0, and its gene product is not immunologically related to the ushA gene product. ushB is hence a functional duplicate of ushA and provides a rationale for the silencing of ushA0. This situation, and the DNA sequence comparison of ushA and ushA0, strongly suggests that rather than being a cryptic gene, ushA0 has been silenced recently during the evolution of S. typhimurium.