Expression of Kingella kingae Type IV Pili Is Regulated by σ ⁵⁴ , PilS, and PilR

Abstract

Kingella kingae is a member of the Neisseriaceae and is being recognized increasingly as an important cause of serious disease in children. Recent work has demonstrated that K. kingae expresses type IV pili that mediate adherence to respiratory epithelial and synovial cells and are selected against during invasive disease. In the current study, we examined the genome of K. kingae strain 269-492 and identified homologs of the rpoN and the pilS and pilR genes that are essential for pilus expression in Pseudomonas aeruginosa but not in the pathogenic Neisseria species. The disruption of either rpoN or pilR in K. kingae resulted in a marked reduction in the level of transcript for the major pilus subunit (pilA1) and eliminated piliation. In contrast, the disruption of pilS resulted in only partial reduction in the level of pilA1 transcript and a partial decrease in piliation. Furthermore, the disruption of pilS in colony variants with high-density piliation resulted in variants with low-density piliation. Mutations in the promoter region of pilA1 and gel shift analysis demonstrated that both 54 and PilR act directly at the pilA1 promoter, with PilR binding to two repetitive elements. These data suggest that the regulation of K. kingae type IV pilus expression is complex and multilayered, influenced by both the genetic state and environmental cues.

Genome-wide screen identifies host colonization determinants in a bacterial gut symbiont

“…Pilin expression is triggered by environmental cues; in S. alvi, this is likely controlled by the two-component pilR/S system, as documented in the related species Kingella kingae (20). Transposon insertions in both pilR and pilS were underrepresented in vivo; although, as in Kingella, nonfunctionalization of the sensor pilS was more tolerated than that of pilR (20).…”

“…Pilin expression is triggered by environmental cues; in S. alvi, this is likely controlled by the two-component pilR/S system, as documented in the related species Kingella kingae (20). Transposon insertions in both pilR and pilS were underrepresented in vivo; although, as in Kingella, nonfunctionalization of the sensor pilS was more tolerated than that of pilR (20). The pilE gene in many bacteria, including commensal Neisseria species (21), is directly regulated by the products of pilR and rpoN (encoding the master transcriptional regulator σ-54), another gene required for gut colonization in our screen.…”

Control of Copper Resistance and Inorganic Sulfur Metabolism by Paralogous Regulators in Staphylococcus aureus

“…The supernatant was removed and the bacterial pellet was air dried. RNA was harvested as previously described using a combination of Tri-Reagent (Sigma) and RNeasy Minikit purification (Qiagen, Valencia, CA) (18) with the exception that after the addition of TRIzol the samples were transferred to bead beater tubes and processed at 6 M/s for 40 s in a bead beater to aid in cell lysis. Random hexamers and Moloney murine leukemia virus reverse transcriptase (Promega, Madison, WI) were used to generate cDNA.…”

“…Random hexamers and Moloney murine leukemia virus reverse transcriptase (Promega, Madison, WI) were used to generate cDNA. Quantitative PCR was performed using the iQ SYBR Green Supermix (Bio-Rad) and the primer pairs as indicated under supplemental Table S3 (18). Quantitation of 16 S ribosomal RNA was used to normalize each sample.…”

Type IV pilins regulate their own expression via direct intramembrane interactions with the sensor kinase PilS

“…pilA transcription in P. aeruginosa and many other species is dependent on the PilS-PilR two-component regulatory system (13)(14)(15)(16). Two-component systems (TCSs) allow bacteria to rapidly detect and adapt to changes in their environment (17).…”