Plasmids pJM1 and pColV-K30 Harbor Iron Uptake Genes That Are Essential for Bacterial Virulence

Jorge H. Crosa

doi:10.1128/9781555817732.ch24

Chapter 24 : Plasmids pJM1 and pColV-K30 Harbor Iron Uptake Genes That Are Essential for Bacterial Virulence

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Affiliations: 1: Department of Molecular Microbiology and Immunology, Oregon Health and Science University, Portland, OR 97201

Content Type: Monograph

Publication Year: 2004

Category: Microbial Genetics and Molecular Biology

Book DOI: 10.1128/9781555817732

Chapter DOI: 10.1128/9781555817732.ch24

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Abstract:

This chapter describes the analysis of pJM1 plasmid and pCoIV-K30 plasmid-mediated iron uptake systems. The iron transport-biosynthesis operon (ITBO) and other anguibactin biosynthetic genes located downstream are bracketed by the highly related ISV-A1 and ISV-A2 insertion sequences. The peptide siderophore anguibactin is synthesized via a nonribosomal peptide synthetase mechanism, an RNA-independent template chain growth process with an assembly line organization of different catalytic and carrier protein domains whose placement and function determine the number and sequence of the amino and carboxylic acids incorporated into the peptide product. Anguibactin can be thought as derived from the precursors dihydroxy benzoic acid (DHBA), histidine, and cysteine. The pJM1 plasmid-mediated proteins AngR, AngM, and AngN play a role in subsequent biosynthetic steps although AngR, in addition to its biosynthetic function, is also essential for regulation of iron transport gene expression. By deletion analysis the TAFr region has been narrowed down to two small subregions that could harbor this TAFr factor. At high iron concentrations iron represses the ITBO mRNA levels and concomitantly induces the synthesis of another antisense RNA (RNAα) that might constitute another novel component of the bacterial iron regulatory circuit. The genetic determinants for the aerobactin iron uptake system of plasmid pColV-K30 were cloned as recombinant plasmid pABN1.

Citation: Crosa J. 2004. Plasmids pJM1 and pColV-K30 Harbor Iron Uptake Genes That Are Essential for Bacterial Virulence, p 493-506. In Funnell B, Phillips G (ed), Plasmid Biology. ASM Press, Washington, DC. doi: 10.1128/9781555817732.ch24

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Plasmids pJM1 and pColV-K30 Harbor Iron Uptake Genes That Are Essential for Bacterial Virulence

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Figure 1

Structure of the siderophores anguibactin, aerobactin, and enterobactin.

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Figure 1

Structure of the siderophores anguibactin, aerobactin, and enterobactin.

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Figure 2

Schematic representation of the pJM1 plasmid. angM, -R, -T, -N, and -B are biosynthetic genes. angR is also a regulatory gene; together with the TAFr regulator they controlled expression of the fatD, -C, -B, -A, angR, angT operon named the ITBO. fatD, -C, -B, and -A are the ferric anguibactin transport genes.

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Figure 2

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Figure 3

Sites of action of Fur on the ITBO promoter. Top and bottom strands represent the nontemplate and template strand, respectively. The protected nucleotides in the DNase I footprint are denoted with an asterisk, and the hypersensitive sites are indicated by vertical arrows. The horizontal arrow indicates the transcription start site (+1) and the direction of transcription in the ITBO. Numbers are nucleotides relative to the transcription start site.

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Figure 3

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Figure 4

Schematic representation of the current understanding of the regulation of transport and biosynthesis genes in V. anguillarum. See text for details.

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Figure 4

Schematic representation of the current understanding of the regulation of transport and biosynthesis genes in V. anguillarum. See text for details.

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Figure 5

Schematic representation of the pColV-K30 plasmid. The aerobactin system region harbors the biosynthetic genes for aerobactin, iucA, -B, -C, and -D and the ferric-aerobactin transport gene iutA. Also shown are the regions encoding the genes for colicinV production (colV), conjugative transfer (tra), replication (REPI and REPII), and the various locations for the insertion element IS1 ( 69 ).

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Figure 5

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Figure 6

The aerobactin biosynthesis transport operon, iucA encodes a 63-kDa protein and iucC a 62-kDa protein that are involved in the synthetase reaction from N epsilon-acetyl-N epsilon-hydroxylysine and citrate; iucB encodes a 33-kDa polypeptide with the activity epsilon-hydroxylysine:acetyl coenzyme A-epsilon transacetylase and iucD encodes a 53-kDa polypeptide a lysine oxygenase, respectively, while iutA encodes the 74-kDa outer membrane protein receptor for ferric aerobactin.

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Figure 6

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Figure 7

Comparison of the Fur-binding sites for the aerobactin operon in pColV-K30 (A) and in the chromosome of E. coli K1 (B). The arrows depict sequences that are inverted and repeated in locations around the sequences. Overlapping arrows indicate their occurrence in both the sense and antisense strands.

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Figure 7

References

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