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Enteroinvasive Escherichia coli virulence-plasmid-carried apyrase (apy) and ospB genes are organized as a bicistronic operon and are subject to differential expression^a,b

Dipartimento di Scienze Biomediche, Sezione di Microbiologia, Università G. D’Annunzio, 66100 Chieti, Italy¹
Dipartimento di Biologia, Università Roma Tre, 00146 Rome, Italy²
Istituto Superiore di Sanità, 00161 Rome, Italy³
Dipartimento di Genitica e Biologia Molecolare⁴, Dipartimento di Scienze di Sanità Pubblica, Sezione di Microbiologia⁵ and Dipartimento di Biologia Cellulare e dello Sviluppo, Sezione di Scienze Microbiologiche⁶, Università di Roma La Sapienza, 00185 Rome, Italy

Author for correspondence: Mauro Nicoletti. Tel: +39 06 4991 4657. Fax: +39 06 4991 4626. e-mail: mauro.nicoletti{at}uniroma1.it

In Shigella flexneri and enteroinvasive Escherichia coli (EIEC) the expression of the virulence-plasmid(pINV)-carried potential pathogenesis-associated apy gene, which encodes apyrase (ATP diphosphohydrolase), is regulated by the same regulators that govern the expression of virulence genes. To understand the transcriptional organization of the apy gene, the authors sequenced an 8023 bp PstI fragment of the pINV of EIEC strain HN280, which encompasses apy as well as its adjacent genes. The PstI fragment displays 99% identity with the corresponding fragment of pWR100, the pINV of S. flexneri strain M90T, and contains four genes. One of these genes, ospB, encodes a secreted protein of unknown activity and is located immediately upstream of apy. Analyses of sequence, Northern hybridization, RT-PCR and primer extension data and transcriptional fusions indicated that ospB and apy are co-transcribed as a 2 kb bicistronic, temperature-regulated mRNA from an upstream promoter that precedes ospB. The 2 kb mRNA is post-transcriptionally processed in the intercistronic ospB–apy region, leading to the considerable accumulation of a more stable 1 kb apy-specific mRNA (half-life of 2·2±0·3 min, versus 27±4 s for the 2 kb transcript). Upon temperature induction, peak expression of the ospB–apy operon occurs when bacteria enter into the late phases of bacterial growth, where the apy-specific transcript was found to be much more prevalent if compared to the ospB–apy transcript.

Keywords: ATP diphosphohydrolase, mRNA processing, polycistronic mRNA

Abbreviations: EIEC, enteroinvasive Escherichia coli; pINV, virulence plasmid; RT, reverse transcriptase

^a This paper is dedicated to the memory of our dear friends and colleagues Giuseppe Carruba and Franco Tatò, who died prematurely.

^b The GenBank accession number for the sequence reported in this paper is AJ315184.

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