The FlhA linker mediates flagellar protein export switching during flagellar assembly

The flagellar protein export apparatus switches substrate specificity from hook-type to filament-type upon hook assembly completion, thereby initiating filament assembly at the hook tip. The C-terminal cytoplasmic domain of FlhA (FlhAC) serves as a docking platform for flagellar chaperones in complex with their cognate filament-type substrates. Interactions of the flexible linker of FlhA (FlhAL) with its nearest FlhAC subunit in the FlhAC ring is required for the substrate specificity switching. To address how FlhAL brings the order to flagellar assembly, we analyzed the flhA(E351A/W354A/D356A) ΔflgM mutant and found that this triple mutation in FlhAL increased the secretion level of hook protein by 5-fold, thereby increasing hook length. The crystal structure of FlhAC(E351A/D356A) showed that FlhAL bound to the chaperone-binding site of its neighboring subunit. We propose that the interaction of FlhAL with the chaperon-binding site of FlhAC suppresses filament-type protein export and facilitates hook-type protein export during hook assembly.

Monitoring of Particulate Matter Emissions from 3D Printing Activity in the Home Setting

Consumer-level 3D printers are becoming increasingly prevalent in home settings. However, research shows that printing with these desktop 3D printers can impact indoor air quality (IAQ). This study examined particulate matter (PM) emissions generated by 3D printers in an indoor domestic setting. Print filament type, brand, and color were investigated and shown to all have significant impacts on the PM emission profiles over time. For example, emission rates were observed to vary by up to 150-fold, depending on the brand of a specific filament being used. Various printer settings (e.g., fan speed, infill density, extruder temperature) were also investigated. This study identifies that high levels of PM are triggered by the filament heating process and that accessible, user-controlled print settings can be used to modulate the PM emission from the 3D printing process. Considering these findings, a low-cost home IAQ sensor was evaluated as a potential means to enable a home user to monitor PM emissions from their 3D printing activities. This sensing approach was demonstrated to detect the timepoint where the onset of PM emission from a 3D print occurs. Therefore, these low-cost sensors could serve to inform the user when PM levels in the home become elevated significantly on account of this activity and furthermore, can indicate the time at which PM levels return to baseline after the printing process and/or after adding ventilation. By deploying such sensors at home, domestic users of 3D printers can assess the impact of filament type, color, and brand that they utilize on PM emissions, as well as be informed of how their selected print settings can impact their PM exposure levels.

Revised subunit order of mammalian septin complexes explains their in-vitro polymerization properties

Septins are conserved GTP-binding cytoskeletal proteins that polymerize into filaments by end-to-end joining of heterooligomeric complexes. In human cells, both hexamers and octamers exist, and crystallography studies predicted the order of the hexamers to be SEPT7-SEPT6-SEPT2-SEPT2-SEPT6-SEPT7, while octamers are thought to have the same core, but with SEPT9 at the ends. However, based on this septin organization, octamers and hexamers would not be expected to co-polymerize due to incompatible ends. Here we isolated hexamers and octamers of specific composition from human cells and show that hexamers and octamers polymerize individually and, surprisingly, with each other. Binding of the BD3 domain of Borg3 results in distinctive clustering of each filament type. Moreover, we show that the organization of hexameric and octameric complexes is inverted compared to its original prediction. This revised septin organization is congruent with the organization and behavior of yeast septins suggesting that their properties are more conserved than was previously thought.

The FlhA linker mediates flagellar protein export switching during flagellar assembly

The flagellar protein export apparatus switches export specificity from hook-type to filament-type upon completion of hook assembly, thereby initiating filament assembly at the hook tip. The C-terminal cytoplasmic domain of FlhA (FlhAC) forms a homo-nonameric ring structure that serves as a docking platform for flagellar export chaperones in complex with their cognate filament-type substrates. Interactions of the flexible linker of FlhA (FlhAL) with its nearest FlhAC subunit in the ring allow the chaperones to bind to FlhAC to facilitate filament-type protein export, but it remains unclear how it occurs. Here, we report that FlhAL acts as a switch that brings the order to flagellar assembly. The crystal structure of FlhAC(E351A/D356A) showed that Trp-354 in FlhAL bound to the chaperone-binding site of its neighboring subunit. We propose that FlhAL binds to the chaperon-binding site of FlhAC to suppress the interaction between FlhAC and the chaperones until hook assembly is completed.

Revised subunit order of mammalian septin complexes explains their in vitro polymerization properties

ABSTRACTSeptins are conserved GTP-binding cytoskeletal proteins that polymerize into filaments by end-to-end joining of heterooligomeric complexes. In human cells, both hexamers and octamers exist, and crystallography studies predicted the order of the hexamers to be SEPT7-SEPT6-SEPT2-SEPT2-SEPT6-SEPT7, while octamers are thought to have the same core, but with SEPT9 at the ends. However, based on this septin organization, octamers and hexamers would not be expected to co-polymerize due to incompatible ends. Here we isolated hexamers and octamers of specific composition from human cells and show that hexamers and octamers polymerize individually and, surprisingly, with each other. Binding of Borg3 results in distinctive clustering of each filament type. Moreover, we show that the organization of hexameric and octameric complexes is inverted compared to its original prediction. This revised septin organization is congruent with the organization and behavior of yeast septins suggesting that their properties are more conserved than was previously thought.