Structural basis of self-assembly in the lipid-binding domain of mycobacterial polar growth factor Wag31

Choukate, Komal; Chaudhuri, Barnali N.

doi:10.1107/s2052252520006053

Cited by 13 publications

(12 citation statements)

References 53 publications

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“…While the N-terminal domain is mainly conserved across the two subgroups, it has been previously shown that the conservation does not extend to the residues that are directly responsible for the interaction with the membrane. This function is, in fact, attributed to the phenylalanine 17 residue in the B. subtilis DivIVA protein (PDB code 2wuj; [ 22 ]) while it has been instead tentatively assigned to the structurally rigid P16P17I18 motif in Mycobacterium tuberculosis [ 53 ].…”

Section: Resultsmentioning

confidence: 99%

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Subcellular Dynamics of a Conserved Bacterial Polar Scaffold Protein

Giacomelli

Feddersen

Feng

et al. 2022

Genes

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In order to survive, bacterial cells rely on precise spatiotemporal organization and coordination of essential processes such as cell growth, chromosome segregation, and cell division. Given the general lack of organelles, most bacteria are forced to depend on alternative localization mechanisms, such as, for example, geometrical cues. DivIVA proteins are widely distributed in mainly Gram-positive bacteria and were shown to bind the membrane, typically in regions of strong negative curvature, such as the cell poles and division septa. Here, they have been shown to be involved in a multitude of processes: from apical cell growth and chromosome segregation in actinobacteria to sporulation and inhibition of division re-initiation in firmicutes. Structural analyses revealed that DivIVA proteins can form oligomeric assemblies that constitute a scaffold for recruitment of other proteins. However, it remained unclear whether interaction with partner proteins influences DivIVA dynamics. Using structured illumination microscopy (SIM), single-particle tracking (SPT) microscopy, and fluorescent recovery after photobleaching (FRAP) experiments, we show that DivIVA from Corynebacterium glutamicum is mobilized by its binding partner ParB. In contrast, we show that the interaction between Bacillus subtilis DivIVA and its partner protein MinJ reduces DivIVA mobility. Furthermore, we show that the loss of the rod-shape leads to an increase in DivIVA dynamics in both organisms. Taken together, our study reveals the modulation of the polar scaffold protein by protein interactors and cell morphology. We reason that this leads to a very simple, yet robust way for actinobacteria to maintain polar growth and their rod-shape. In B. subtilis, however, the DivIVA protein is tailored towards a more dynamic function that allows quick relocalization from poles to septa upon division.

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Section: Resultsmentioning

confidence: 99%

“…We first modeled the N-terminal region of DivIVA cgb ( Figure 1 A) with Wag31 (Rv2145c) as a template (PDB code 6lfa; [ 53 ]) and could confirm that the PPI membrane interface domain three-dimensional arrangement is conserved between C. glutamicum and M. tuberculosis .…”

Section: Resultsmentioning

confidence: 99%

Subcellular Dynamics of a Conserved Bacterial Polar Scaffold Protein

Giacomelli

Feddersen

Feng

et al. 2022

Genes

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“…2B). This leucine forms an apparent hydrophobic interaction with L2 of the other monomer in the crystal structure of the N-terminus (34). The Wag31 L34A protein is largely functional (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…S1). This part of the protein dimerizes and has been structurally characterized (29, 34). In Bacillus subtilis , F17 is involved in interaction with the membrane at the cell pole (29).…”

Section: Resultsmentioning

confidence: 99%

Polar growth protein Wag31 undergoes changes in homo-oligomeric network topology, and has distinct functions at both cell poles and the septum

Arejan

Patel

Quintanilla

et al. 2022

Preprint

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Mycobacterial cell elongation occurs at the cell poles; however, it is not clear how cell wall insertion is restricted to the pole and organized. Wag31 is a pole-localized cytoplasmic protein that is essential for polar growth, but its molecular function has not been described. Wag31 homo-oligomerizes in a network at the poles, but it is not known how the structure of this network affects Wag31 function. In this study we used a protein fragment complementation assay to identify Wag31 residues involved in homo-oligomeric interactions, and found that amino acids all along the length of the protein mediate these interactions. We then used both N-terminal and C-terminal splitGFP fusions to probe Wag31 network topology at different sites in the cell, and found that Wag31 C-terminal-C-terminal interactions predominate at the septa, while C-terminal-C-terminal and C-terminal-N-terminal interactions are found equally at the poles. This suggests the Wag31 network is formed through an ordered series of associations. We then dissected Wag31’s functional roles by phenotyping a series of wag31 alanine mutants; these data show that Wag31 has separate functions in not only new and old pole elongation, but also inhibition of both septation and new pole elongation. This work establishes new functions for Wag31, and indicates that changes in Wag31 homo-oligomeric network topology may contribute to cell wall regulation in mycobacteria.

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“…The N-terminus of Wag31 is highly conserved and it has been structurally characterized (35, 36). In Bacillus subtilis , F17 is involved in interaction with the membrane at the cell pole (35).…”

Section: Resultsmentioning

confidence: 99%