The unstable melC operon of Streptomyces antibioticus is codeleted with a Tn4811-homologous locus

Abstract

The meiC operon ofStreptomyces antibioticus is unstable, undergoing frequent spontaneous deletions. All the AIelC mutants analyzed also lost 2-kb V1 DNA, which contained two open reading frames (ORFs) (2,17,20,21). The two operons share sequence homology and gene arrangement. In both cases, the operon consists of the structural genes for apotyrosinase (melC2) and a trans-acting protein (melCI) necessary for copper transfer, activation, and secretion of tyrosinase (10, 23). The meiC sequences in other melaninproducing Streptomyces species also appear to be conserved (29). The meiC operon has been located on the chromosome of S. glaucescens by genetic mapping (13) and on the chromosomes of Streptomyces reticuli and Streptomyces scabies by hybridization (29).The melanin pigmentation (Mel') phenotype in Streptomyces species is unstable (29). Spontaneous melaninless (Mel-) mutants arise at frequencies of the order of i0'. The mutations are primarily deletions of the meiC sequence (4,16,28,29). Other Mel-variants result from mutations in other loci (9, 13). Although the cloned S. antibioticus meiC operon has been a widely used marker (21), it is structurally unstable on recombinant vectors, undergoing deletions and insertions (for examples, see references 9, 24, and 30). We previously isolated a 5.4-kb transposon, Tn4811, of Streptomyces lividans from within the meiC operon (9). Tn4811 contained five typical Streptomyces coding sequences (9). Open reading frame 1 (ORF1), ORF2, and ORF3 were proposed to be involved in transposition and regulatory functions. ORF4 and ORF5 appeared to form a unit divergently transcribed from a shared promoter region. The ORF4 product resembled a group of oxidoreductases, and the ORF5 product resembled the AraC family of prokaryotic transcription activators. Fifteen other Streptomyces species were probed for Tn4811 homologs. Only S. antibioti-TABLE 1. Bacterial strains and plasmids used in this study Strain(s) or plasmid

Extracellular and Intracellular Polyphenol Oxidases Cause Opposite Effects on Sensitivity of Streptomyces to Phenolics: A Case of Double-Edged Sword

“…Such instability of melC in S. avermitilis is at about the same level as in several other species studied [e.g. ], [7], [8], [9,10]. The seven Mel − derivatives still possessing melC are presumably defective in other genetic elements required for expression of tyrosinase, as previously described in other species [21], [22], [23].…”

“…B19 is a ΔmelC mutant of S. avermitilis isolated in this study. YU5 is a ΔmelC mutant of S. antibioticus isolated previously [10] . M145/pIJ702-117 is M145 harbouring pIJ702-117, a high-copy-number plasmid containing the melC operon from S. antibioticus under a strong promoter [27] .…”

“…The melC operon has been mapped to the linear chromosomes of many Streptomyces species. It is highly unstable, being prone to spontaneous deletion along with neighboring sequences [7] , [8] , [9] , [10] . This reflects a terminal location of this operon on the linear chromosomes [10] .…”

Arac/XylS family of transcriptional regulators.

“…Members of the AraC/XylS family are widely distributed in diverse prokaryote genera ( Table 1). The GϩC content of genes encoding AraC/XylS family members vary from 28% for E. coli rns (36) to at least 67% for Streptomyces araL (43,265). Most of the genes encoding members of this family are in the genomes of the gamma subdivision of the proteobacteria (purple bacteria) (194).…”

High-frequency transposition of IS1373, the insertion sequence delimiting the amplifiable element AUD2 of Streptomyces lividans

“…Both consist of two large, terminal inverted repeats (IRs) and contain one copy of Tn4811 (11). A Tn4811-homologous locus is also codeleted with the unstable melC operon of S. antibioticus (25). Nevertheless, in S. lividans, when both copies of the transposon are artificially deleted with the chromosomal ends, the level of genetic instability is not reduced but on the contrary is enhanced after concomitant circularization of the chromosome (24).…”