Topological, Functional, and Phylogenetic Analysis of the Cell Division Site Selection Protein MinJ of Bacillus subtilis.

The Min system disassembles FtsZ-rings after septation in Bacillus subtilis and is localized to the nascent division plane and cell poles by the protein MinJ. The N-terminal region of MinJ contains transmembrane segments while the C-terminal region contains a PDZ domain, but its membrane topology and functional domains are poorly understood. Here, we empirically tested competing topology models and found that MinJ is a nine-pass transmembrane protein with an extracellular N terminus and a cytoplasmic C terminal PDZ domain. Deletion of the PDZ domain resembled a MinJ null mutation but retained polar localization and interaction with both MinD and DivIVA. Deletion of transmembrane segments 6 and 7 abolished polar localization and interaction with MinD and DivIVA but conferred a hyper-minicell phenotype that was additive in the absence of MinD. Structural prediction by AlphaFold3 suggests that segments 6 and 7 may constitute an unusual transmembrane β-sheet, making MinJ perhaps the first of its kind to have such a secondary structure in the plasma membrane. Finally, phylogenetic analyses and structural comparison showed that MinJ sequence and structure are highly conserved within the Firmicutes and tightly co-conserved with MinD and DivIVA with which it interacts. Our work clarifies the membrane topology and functional organization of MinJ and supports models in which MinJ contributes to bacterial cell division through mechanisms beyond canonical Min system regulation.