Localization and interactions of Teichoic Acid synthetic enzymes in Bacillus subtilis

Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford, OX1 3RE, UK; Génétique Microbienne. Institut National de la Recherche Agronomique, 78352 Jouy-en-Josas Cedex, France; Institute for Cell and Molecular Biosciences, Faculty of Medical Sciences, Newcastle University,, Framlington Place, Newcastle, NE2 4HH, UK; Molecular Microbiology, Institute for Molecular Cell Biology, VU University Amsterdam, De Boelelaan 1085, 1081 HV Amsterdam, the Netherlands

^* To whom correspondence should be addressed. Email: jeff.errington{at}ncl.ac.uk.

	Abstract

The thick wall of Gram-positive bacteria is a polymer meshwork composed predominantly of peptidoglycan (PG) and teichoic acids, both of which have a critical function in maintenance of the structural integrity and the shape of the cell. In Bacillus subtilis 168 the major teichoic acid is covalently coupled to PG and is known as wall teichoic acid (WTA). Recently, PG insertion/degradation over the lateral wall has been shown to occur in a helical pattern. However, the spatial organization of WTA assembly and its relationship with cell shape and PG assembly are largely unknown. We have characterized the localization of GFP fusions to proteins involved in several steps of WTA synthesis in B. subtilis: TagB/F/G/H/O. All of these localized similarly to the inner side of the cytoplasmic membrane, in a pattern strikingly similar to that displayed by probes of nascent PG. Helix-like localization patterns are often attributable to the morphogenic cytoskeletal proteins of the MreB family. However, localization of the Tag proteins did not appear to be substantially affected by single disruption of any of the three MreB homologs of B. subtilis. Bacterial and yeast two-hybrid experiments revealed a complex network of interactions involving TagA/B/E/F/G/H/O, and the cell shape determinants MreC and MreD (encoded by the mreBCD operon and presumably involved in the spatial organization of PG synthesis). Taken together, our results suggest that, in B. subtilis at least, the synthesis and export of WTA precursors are mediated by a large multi-enzyme complex that may be associated with the PG-synthesizing machinery.