Outer membrane lipoprotein RlpA is a novel periplasmic interaction partner of the cell division protein FtsK in Escherichia coli

Alison M Berezuk,Jonathan R Krieger,Cezar M Khursigara,Elyse J Roach,Mara C Goodyear,Sabrina Glavota

doi:10.1038/s41598-018-30979-5

Alison M Berezuk, Jonathan R Krieger + Show 4 more

Open Access

https://doi.org/10.1038/s41598-018-30979-5

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Abstract

In Escherichia coli, formation of new cells is mediated by the elongasome and divisome that govern cell elongation and septation, respectively. Proper transition between these events is essential to ensure viable progeny are produced; however, the components of each complex responsible for transmission of the cell signal to shift from elongation to septation are unclear. Recently, a region within the N-terminal domain of the essential divisome protein FtsK (FtsKN) was identified that points to a key role for FtsK as a checkpoint of cell envelope remodeling during division. Here, we used site-specific in vivo UV cross-linking to probe the periplasmic loops of FtsKN for protein interaction partners critical for FtsKN function. Mass spectrometry analysis of five unique FtsKN periplasmic cross-links revealed a network of potential FtsKN interactors, one of which included the septal peptidoglycan binding protein rare lipoprotein A (RlpA). This protein was further verified as a novel interaction partner of FtsKN by an in vitro pull-down assay. Deletion of rlpA from an FtsK temperature-sensitive E. coli strain partially restored cell growth and largely suppressed cellular filamentation compared to the wild-type strain. This suggests that interaction with RlpA may be critical in suppressing septation until proper assembly of the divisome.

Highlights

Through the combined action of two large macromolecular protein complexes, cells undergo two broad morphological changes during division
FtsK is a large, multi-spanning membrane protein that is composed of 1329 amino acids and has a molecular weight of approximately 147 kDa20,21. It belongs to the large, well-conserved SpoIIIE family of proteins that facilitates double-stranded DNA translocation via its C-terminal domain during division and sporulation in E. coli and Bacillus subtilis, respectively[22,23]
To identify FtsKN periplasmic interaction partners, a site-specific in vivo UV cross-linking approach was used. This technique has been successfully used in E. coli to probe various protein interaction surfaces, including mapping of the SecA dimer interface and its interaction with the Sec translocon[38,39], transmembrane translocation by the Tat-pathway[40], capsular polysaccharide export by Wza[41] and formation of the essential divisome sub-complex FtsQ/B/L42

Summary

Introduction

Through the combined action of two large macromolecular protein complexes, cells undergo two broad morphological changes during division. Many parallels exist between the functions and general organization of each complex[2,3], it is still unclear how the elongasome and divisome interact to integrate essential information regarding the growth state of the cell It is unknown how the switch from dispersed lateral peptidoglycan synthesis to concentrated synthesis at the new cell poles and invagination of the cell envelope during septation occurs. Co-localization and protein interaction analysis of several elongasome and divisome proteins indicates that these complexes are simultaneously present at mid-cell for approximately 40% of the cell division cycle, and that the peptidoglycan synthesis enzymes from each complex physically interact prior to visible cell constriction[19]. While sufficient biochemical evidence suggests that FtsKN interacts with the Z-ring during division[27,31,32,33], interaction between FtsK and proteins involved in peptidoglycan synthesis (e.g., FtsI) has not been unequivocally verified[27,31,32]

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