c Flagella are surface appendages that are important for bacterial motility and invasion of host cells. Two flagellin subunits in Salmonella enterica serovar Typhimurium, FliC and FljB, are alternatively expressed by a site-specific DNA inversion mechanism called flagellar phase variation. Although this inversion mechanism is understood at the molecular level, the key factor controlling the expression of the two flagellin subunits has not been determined. In this study, we found that a putative acyl carrier protein, IacP, affects flagellar phase variation in S. Typhimurium strain UK-1 under Salmonella pathogenicity island 1 (SPI1)-inducing conditions. Liquid chromatography-mass spectrometry analysis of the secreted proteins from S. Typhimurium determined that the amount of FljB secreted was significantly higher in the iacP mutant strain, a finding confirmed by Western blot analysis. Northern blotting, quantitative PCR, and microarray data showed that the level of FljB in the iacP mutant strain was regulated at the transcriptional level, although the transcription and expression of the fliC gene were independent of IacP. FljB production was abolished by the deletion of the Hin DNA invertase but could be restored by the introduction of a plasmid carrying the hin gene. We also found that in the iacP mutant strain, the orientation of the invertible H segment is in the FljB-expressing phase. Furthermore, electron microscopy observations indicated that the iacP mutant strain had more flagella per cell than the wildtype strain. These results suggest that IacP is associated with flagellar phase switching under SPI1-inducing conditions. F lagella play crucial roles in bacterial motility, chemotactic behavior, and host cell invasion as a virulence determinant (1). Salmonella enterica serovar Typhimurium has 5 to 10 flagella emanating from the bacterial surface. The individual flagellum is a complex structure composed of three basic parts: a basal body, a hook, and a filament (22). The flagellar filament consists of thousands of flagellin protein monomers, which are polymerized into a helical structure that can be 10 m long. S. Typhimurium possesses two genetically distinct flagellin subunits, FliC (phase 1 flagellin) and FljB (phase 2 flagellin), which are alternatively expressed via a site-specific DNA inversion process (19,30). When the 1-kb upstream region of the fljB gene (H segment) is inverted by the DNA invertase Hin, the promoter for the fljB gene is properly oriented for the transcription of the fljB-fljA operon. Because FljA is responsible for the posttranscriptional and posttranslational inhibition of FliC expression (2, 38), only FljB flagellin is produced. Although the molecular mechanism of the Hin-mediated DNA inversion mechanism has been studied intensively (6, 24, 28), the question of which cellular signals control the flagellinswitching mechanism and the frequency of DNA inversion remains unclear.Salmonella pathogenicity island 1 (SPI1) encodes a type III secretion system (T3SS) that is required for the ...