Biology subjects

Casjens, S. R.

Publications and source records attributed to Casjens, S. R..

4 recordsLinked to original sources

bioRxiv2026

Active removal of inhibitory components drives the flagellarType III Secretion Specificity Switch

Type III secretion (T3S) systems assemble bacterial nanomachines, including the flagellum and virulence-associated injectisomes, by exporting distinct classes of substrates in a defined temporal order. In both systems, completion of an early assembly intermediate triggers an irreversible switch from early to late substrate secretion. In the flagellar system, this switch is controlled by the secreted molecular ruler FliK acting on the core T3S component FlhB, but the molecular mechanism governing this transition has remained unclear. Here we show that removal of two components, Fluke and the cleaved C-terminal domain of FlhB (FlhBCCD), locks the secretion apparatus in a constitutive late secretion state. In these mutants, secretion specificity no longer requires completion of the hook-basal body or the FliK ruler, indicating that Fluke and FlhBCCD function to maintain the apparatus in early secretion mode. Consistent with this model, synchronized flagellar gene expression experiments reveal that FlhBCCD is retained during early assembly and is lost coincident with hook-basal body completion and activation of {sigma}28-dependent late gene expression of flagellin and chemosensory genes. Structural modeling of the FliK C-terminal switch domain and FlhBCCD supports a mechanism in which secretion of FliK promotes destabilization and ejection of FlhBCCD from the secretion apparatus. Disruption of a folded region within FliK switch domain uncouples secretion from switching, indicating that the timing of FliK unfolding during secretion is critical for activation of the specificity switch. These findings show that secretion specificity switching is driven by FliK-dependent removal of inhibitory components, rather than passive sensing of assembly completion. Significance StatementType III secretion (T3S) systems build complex bacterial nanomachines, including the flagellum and virulence-associated injectisomes, by exporting distinct classes of substrates in a defined temporal order. How these systems switch secretion specificity during assembly has remained a long-standing question. We demonstrate that the flagellar T3S specificity switch requires removal of two inhibitory components that actively maintain the apparatus in an early secretion state. Their FliK-dependent ejection irreversibly triggers the transition to late substrate export, revealing that secretion switching is controlled by active inhibitory regulation rather than passive sensing of assembly completion. These results define a molecular mechanism for hierarchical control in bacterial secretion systems and provide insights likely relevant to other T3S nanomachines.

microbiology↗

bioRxiv2023

Bacteriophage P22 SieA mediated superinfection exclusion

Many temperate phages encode prophage-expressed functions that interfere with superinfection of the host bacterium by external phages. Salmonella phage P22 has four such systems that are expressed from the prophage in a lysogen that are encoded by the c2 (repressor), gtrABC, sieA, and sieB genes. Here we report that the P22-encoded SieA protein is the only phage protein required for exclusion by the SieA system, and that it is an inner membrane protein that blocks DNA injection by P22 and its relatives, but has no effect on infection by other tailed phage types. The P22 virion injects its DNA through the host cell membranes and periplasm via a conduit assembled from three "ejection proteins" after their release from the virion. Phage P22 mutants were isolated that overcome the SieA block, and they have amino acid changes in the C-terminal regions of the gene 16 and 20 encoded ejection proteins. Three different single amino acid changes in these proteins are required to obtain nearly full resistance to SieA. Hybrid P22 phages that have phage HK620 ejection protein genes are also partially resistant to SieA. There are three sequence types of extant phage-encoded SieA proteins that are less than 30% identical to one another, yet comparison of two of these types found no differences in target specificity. Our data are consistent with a model in which the inner membrane protein SieA interferes with the assembly or function of the periplasmic gp20 and membrane-bound gp16 DNA delivery conduit. HIGHLIGHTSO_LIPhage P22 SieA protein blocks DNA injection by P22-like phages C_LIO_LISieA is an inner membrane protein C_LIO_LIHybrid P22 phages with phage HK620 ejection proteins partially escape SieA exclusion C_LIO_LISieA escape mutants of P22 alter the gp16 and 20 proteins that form the DNA ejection tube. C_LI

biochemistry↗

bioRxiv2022

Detection of Borrelia garinii in the USA

Borrelia garinii, is a cause of Lyme disease in Europe and Asia. For the first time, we report it in the southeastern United States in rodents. Whole genome sequencing and phylogenetic analysis revealed that USA-located B. garinii is part of a clade consisting primarily of few European and majority of Far Eastern strains. Continued surveillance of wildlife hosts and ticks is necessary to assess the ecological status and public health risks of B. garinii in southeastern US.

microbiology↗

bioRxiv2018

How phage L capsid decoration protein distinguishes between nearly identical binding sites on an icosahedral virion

The major coat proteins of dsDNA tailed phages and herpesviruses form capsids by a mechanism that includes active packaging of the dsDNA genome into a precursor procapsid, followed by expansion and stabilization of the capsid. These viruses have evolved diverse strategies to fortify their capsids, such as non-covalent binding of auxiliary "decoration" (Dec) proteins. The Dec protein from the P22-like phage L has a highly unusual binding strategy that precisely distinguishes between nearly identical three-fold and quasi-three-fold sites of the icosahedral capsid. Cryo-electron microscopy and three-dimensional image reconstruction were employed to determine the structure of native phage L particles. NMR was used to determine the structure/dynamics of Dec in solution. Lastly, the NMR structure and the cryo-EM density envelope were combined to build a model of the capsid-bound Dec trimer. Key regions that modulate the binding interface were verified by site-directed mutagenesis.

biophysics↗