Surface expression of O-specific lipopolysaccharide in Escherichia coli requires the function of the TolA protein

We investigated the involvement of Tol proteins in the surface expression of lipopolysaccharide (LPS). tolQ, -R, -A and -B mutants of Escherichia coli K-12, which do not form a complete LPS-containing O antigen, were transformed with the O7+ cosmid pJHCV32. The tolA and tolQ mutants showed reduced O7 LPS expression compared with the respective isogenic parent strains. No changes in O7 LPS expression were found in the other tol mutants. The O7-deficient phenotype in the tolQ and tolA mutants was complemented with a plasmid encoding the tolQRA operon, but not with a similar plasmid containing a frameshift mutation inactivating tolA. Therefore, the reduction in O7 LPS was attributed to the lack of a functional tolA gene, caused either by a direct mutation of this gene or by a polar effect on tolA gene expression exerted by the tolQ mutation. Reduced surface expression of O7 LPS was not caused by changes in lipid A-core structure or downregulation of the O7 LPS promoter. However, an abnormal accumulation of radiolabelled mannose was detected in the plasma membrane. As mannose is a sugar unique to the O7 subunit, this result suggested the presence of accumulated O7 LPS biosynthesis intermediates. Attempts to construct a tolA mutant in the E. coli O7 wild-type strain VW187 were unsuccessful, suggesting that this mutation is lethal. In contrast, a polar tolQ mutation affecting tolA expression in VW187 caused slow growth rate and serum sensitivity in addition to reduced O7 LPS production. VW187 tolQ cells showed an elongated morphology and became permeable to the membrane-impermeable dye propidium iodide. All these phenotypes were corrected upon complementation with cloned tol genes but were not restored by complementation with the tolQRA operon containing the frameshift mutation in tolA. Our results demonstrate that the TolA protein plays a critical role in the surface expression of O antigen subunits by an as yet uncharacterized involvement in the processing of O antigen.

Role of tol genes in cloacin DF13 susceptibility of Escherichia coli K-12 strains expressing the cloacin DF13-aerobactin receptor IutA

IutA is the outer membrane protein receptor for ferric aerobactin and the bacteriocin cloacin DF13. Although the same receptor is shared, ferric aerobactin transport across the outer membrane in Escherichia coli is TonB dependent, whereas cloacin DF13 transport is not. We have recently observed that tolQ is required for cloacin DF13 susceptibility (J.A. Thomas and M.A. Valvano, FEMS Microbiol. Lett. 91:107-112, 1992). In this study, we demonstrate that the genes tolQ, tolR, and tolA, but not tolB, tolC, and ompF, are required for the internalization of cloacin DF13 and they are not involved in the transport of ferric aerobactin.

tolQ is required for cloacin DF13 susceptibility in Escherichia coli expressing the aerobactin/cloacin DF13 receptor IutA

We investigated the role of the tolQ gene in the import of cloacin DF13 across the outer membrane of Escherichia coli strains expressing the IutA receptor. The IutA outer-membrane protein is the receptor for the siderophore ferric aerobactin and also binds cloacin DF13, a bacteriocin produced by strains of Enterobacter aerogenes. In this report we present evidence that tolQ is required for the internalization of cloacin DF13 upon binding to IutA but it is not involved in the transport of ferric aerobactin.

Church of St George and St Thomas

Abstract: A vibrant inner city parish needed space for meetings, language classes, children’s play and other support accommodation as well as a clearer link between the interior of the listed church and the space outside.

The project builds itself about the entrance to the church. The form is manipulated such that the intervention recedes from those entering the church, drawing them into the plan before becoming readable as an addition. The resultant poché between this entrance sequence and the fabric of the church is hollowed out to provide the required accommodation. These rooms are insulated and lined in cork to allow for their use separate to the main body of the church. With budget at a premium the construction methodology was developed from an analysis of traditional Irish boat building techniques, which allowed the use of the solid timber to act as the primary structure with no additional material support.

Constructed in solid walnut the intervention reads with the existing brick interior and yet is clearly identifiable as a contemporary addition.

Aims / Objectives Questions

1 To accommodate new space inside an existing protected structure.
2 To form a new threshold between interior and exterior.
3 To develop an affordable means of construction that would be durable and rapid to erect.
4 To make a contemporary addition in sympathy with the qualities of the existing protect structure, in line with best conservation practice and research.
5 Traditional forms of construction as a model for contemporary technologies.

In the Shadows of the Old Town: Thomas Anna and 19th-century Glasgow

In 1858, a volume entitled Midnight Scenes and Social Photographs – being sketches of life in the streets, wynds and dens of the city of Glasgow was published under the pseudonym of ‘Shadow’ by Alexander Brown, a Glaswegian flâneur and reformer. Its frontispiece is an etching which depicts a theatre-like proscenium arch whose curtains have been withdrawn to reveal to the audience all the poverty, destitution and disorder that one was likely to find after dark in the insalubrious quarters of the city. At the extreme left-hand side, partly obscured by the curtain a silhouetted figure stands behind an unwieldy camera perched on a tripod. Distinctly unaffected by the mêlée, an arm is calmly raised and a finger precisely arched in the moment before the shutter is clicked and the scene committed to record. The volume, however, relies exclusively on textual descriptions to evoke the underside of the city and contains no photographs at all. Instead, the use of the word photograph in the title can be understood as a metaphor for detached scientific objectivity, a quality much celebrated by nineteenth-century reformers and investigators of social ills. As it happened, a decade after Shadow disappeared into the labyrinthine back-lands of Old Town Glasgow, he was followed there by a real photographer. In 1868, Thomas Annan was commissioned by the City Improvements Trust to take photographs of the Old Town in its last moments of existence before it was pulled down under a series of legislative acts. But perhaps paradoxically, given Shadow’s faith in the analytical properties of photography, Annan’s work seems to refute much of the material contained in Midnight Scenes and other similar tracts. Instead of the dens, shebeens, labyrinths and rowdy crowds described by Shadow, Annan’s depictions of the Old Town convey a static, calm environment, one which is often sparsely inhabited by a curious but apparently orderly population.

Taking account of the sensational tendencies of many reformists’ texts, this paper investigates the discrepancies between the two representations, focussing in particular on the constraints which operated on Annan during his commission. It argues that Annan’s compositions – which became very influential on other 19th century photographers of everyday life such as John Thomson or Jacob Riis – far from being dispassionate analytical works, emerged as a result of a matrix of factors which included: photographic and artistic precedents; Annan’s own predilections as a photographer; technological limitations; the nature of the commission from the City Improvements Trust and political climate in which it was given; the medieval urban fabric in which he had to operate; and, perhaps, most importantly, the identity of the Old Towns inhabitants themselves.

'That Puissant City': John Milton's Roman Sojourns 1638-1639

John Milton’s sojourns in Rome (1638-9) are attested by his comments in Defensio Secunda, by the minutes of the English College, by Latin encomia which he received from Roman academicians, and, not least, by his Latin letter to Lucas Holstenius (19/29 March 1639), and several Latin poems which he composed in the course of his residency in the capital city: Ad Salsillum, and three Latin epigrams extolling the praises of the virtuosa soprano, Leonora Baroni. Read together, these texts serve to reveal much about Milton’s participation in, and reaction to, the ‘Puissant City’, (History of Britain, Bk 2).

The present monograph presents fresh evidence of Milton's integration into the academic and cultural life of seventeenth-century Rome. It argues that his links with two Roman academies: the Accademia dei Fantastici and Accademia degli Umoristi constitute a sustained participation in an academic community paralleling that of his independently attested performance in Florentine academies (on which I have published extensively). It also investigates his links with Alessandro Cherubini, David Codner, Giovanni Batista Doni, and the Baroni circle hymned in three published anthologies.

Chapter 1: Milton and the Accademia dei Fantastici investigates the cultural climate surrounding Milton's Ad Salsillum by examining two of that academy's publications: the Poesie dei Signori Accademici Fantastici di Roma (Rome, 1637) and the Academia Tenuta da Fantastici a. 12 di Maggio 1655 (Rome, 1655), the latter celebrating the creation of Fabio Chigi as Pope Alexander VII on 5 April 1655. Read in a new light, Milton’s self-fashioning, it is argued, takes its place not only alongside Salzilli’s encomium in Milton's honour, and his Italian sonnets in the 1637 Poesie, but also in relation to other poems in that collection, and the academy's essentially Catholic eulogistic trend. The chapter also provides fresh evidence of Salzilli’s survival of the illness described in Milton’s poem by his epistolary correspondence with Tomaso Stigliani.

Chapter 2: Milton and the Vatican argues for links between Milton’s Latin letter to Holstenius and a range of Holstenius’ published works: his edition of the axioms of the later Pythagoreans gifted by him to Milton, and his published neo-Platonic works. This is achieved by mutual appropriation of Similitudes in a series of Miltonic similes, the anabasis/katabasis motifs in a reworking of the Platonic theory of the transmigration of souls, and allusion to etymological details highlighted in Holstenius’ published editions. The chapter also reveals Milton’s alertness to typographical procedures and, by association, to Holstenius’ recent role (1638) as Director of the press of the Biblioteca Vaticana.

Chapter 3: Milton and the Accademia degli Umoristi argues for Milton’s likely participation in this Roman academy, as suggested by his links with its members. His three Latin epigrams in praise of Leonora Baroni, the only female member of the Umoristi, have hitherto been studied in relation to the 1639 Applausi in her honour. In a new reading, Milton, it is suggested, invokes and interrogates Catholic doctrine before a Catholic audience only to view the whole through the lens of a neo-Platonic Hermeticism (by echoing the phraseology of the sixteenth-century Franciscan Hannibal Rosselli) that refreshingly transcends religious difference. Crucially, the hitherto neglected L’Idea della Veglia (Rome, 1640) includes further encomiastic verse, sonnets to, and by Leonora, and details of the conversazioni hosted by her family at the precise time of Milton’s Roman sojourns. Milton may well have been a participant. The chapter concludes in an assessment of his links with the youthful prodigy Alessandro Cherubini, and of his audience with Francesco Barberini.

Chapter 4: Milton at a Roman Opera analyses the potential impact of ‘Chi Soffre, Speri’, which he attended on 18/28 February 1639, mounted by Francesco Barberini to inaugurate the recently completed theatre of the Palazzo Barberini. A detailed analysis of the opera's libretto, music, and theatricality casts a backward glance to Milton's Comus, and a forward glance to Paradise Lost. It also assesses Milton’s musical interests at this time, as attested by his links with Doni, and his purchase of works by Monteverdi and others.

Chapter 5: Milton’s English Connections in Rome develops the work of Miller and Chaney by investigating Milton’s co-diners at the English College in Rome on 30 October 1638, and by analysing his links between David Codner (alias Matteo Selvaggio), and the family of Jane Savage, Marchioness of Winchester, lamented by Milton in 1631. It also assesses his potential relations with the Englishman Thomas Gawen, who ‘accidentally sometimes fell into the company of John Milton’ (Antony Wood).