Dataset Information

Regulation of cathelicidin gene expression: induction by lipopolysaccharide, interleukin-6, retinoic acid, and Salmonella enterica serovar typhimurium infection.

ABSTRACT: Cathelicidins are a family of antimicrobial peptides prominent in the host defense mechanisms of several mammalian species. In addition to their antimicrobial activities, these peptides have been implicated in wound healing, angiogenesis, and other innate immune mechanisms. To investigate the regulatory mechanisms of cathelicidin gene expression, we conducted in vitro experiments evaluating the bone marrow cell expression of two porcine cathelicidins, PR-39 and protegrin, and cloned and evaluated the promoter sequence of PR-39. In addition, we evaluated in vivo kinetics of cathelicidin gene expression in pigs during an infection with Salmonella enterica serovar Typhimurium. Lipopolysaccharide (LPS) increased PR-39 and protegrin mRNA expression, which was ameliorated by polymyxin B. Concentrations of PR-39 in supernatants from bone marrow cell cultures were increased 10-fold after LPS stimulation. Similarly, interleukin-6 (IL-6) and all-trans retinoic acid (RA) markedly induced cathelicidin gene expression. To verify the transcriptional activation of the PR-39 gene by these agents, we made a PR-39 promoter-luciferase construct containing the full-length PR-39 promoter driving luciferase gene expression and transiently transfected PK-15 epithelial cells. RA and IL-6 increased luciferase activity in PK-15 cells transfected with the PR-39 promoter-luciferase reporter. Similarly, Salmonella-challenged pigs showed increased expression of PR-39 and protegrin mRNA in bone marrow cells at 6 and 24 h postchallenge. Taken together, these findings show that bacterial products (LPS), IL-6, RA, and Salmonella infection enhance the expression of the cathelicidins, PR-39 and protegrin, in bone marrow progenitor cells, and we suggest that extrinsic modulation of this innate host defense mechanism may be possible.

SUBMITTER: Wu H

PROVIDER: S-EPMC101505 | biostudies-literature | 2000 Oct

REPOSITORIES: biostudies-literature

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Regulation of cathelicidin gene expression: induction by lipopolysaccharide, interleukin-6, retinoic acid, and Salmonella enterica serovar typhimurium infection.

Wu H H Zhang G G Minton J E JE Ross C R CR Blecha F F

Infection and immunity 20001001 10

Cathelicidins are a family of antimicrobial peptides prominent in the host defense mechanisms of several mammalian species. In addition to their antimicrobial activities, these peptides have been implicated in wound healing, angiogenesis, and other innate immune mechanisms. To investigate the regulatory mechanisms of cathelicidin gene expression, we conducted in vitro experiments evaluating the bone marrow cell expression of two porcine cathelicidins, PR-39 and protegrin, and cloned and evaluate ...[more]

PMID: 10992453

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Project description:Penetration of intestinal epithelial cells by Salmonella enterica serovar Typhimurium requires the expression of invasion genes, found in Salmonella pathogenicity island 1 (SPI1), that encode components of a type III secretion apparatus. These genes are controlled in a complex manner by regulators within SPI1, including HilA and InvF, and those outside SPI1, such as the two-component regulators PhoP/PhoQ and BarA/SirA. We report here that epithelial cell invasion requires the serovar Typhimurium homologue of Escherichia coli csrA, which encodes a regulator that alters the stability of specific mRNA targets. A deletion mutant of csrA was unable to efficiently invade cultured epithelial cells and showed reduced expression of four tested SPI1 genes, hilA, invF, sipC, and prgH. Overexpression of csrA from an induced araBAD promoter also negatively affected the expression of these genes, indicating that CsrA can act as both a positive and a negative regulator of SPI1 genes and suggesting that the bacterium must tightly control the level or activity of CsrA to achieve maximal invasion. We found that CsrA affected hilA, a regulator of the other three genes we tested, probably by controlling one or more genetic elements that regulate hilA. We also found that both the loss and the overexpression of csrA reduced the expression of two regulators of hilA, hilC and hilD, suggesting that csrA exerts its control of hilA through one or both of these regulators. We further found, however, that CsrA could affect the expression of both invF and sipC independent of its effects on hilA. One additional striking phenotype of the csrA mutant, not observed in a comparable E. coli mutant, was its slow growth. Phenotypic revertants that had normal growth rates, while maintaining the csrA mutation, were common. These suppressed strains, however, did not recover the ability to invade cultured cells, indicating that the csrA-mediated loss of invasion cannot be attributed simply to poor growth and that the growth and invasion deficits of the csrA mutant arise from effects of CsrA on different targets.

Project description:Salmonella enterica serovar Typhimurium (S Typhimurium) relies upon the inner membrane protein PbgA to enhance outer membrane (OM) integrity and promote virulence in mice. The PbgA transmembrane domain (residues 1 to 190) is essential for viability, while the periplasmic domain (residues 191 to 586) is dispensable. Residues within the basic region (residues 191 to 245) bind acidic phosphates on polar phospholipids, like for cardiolipins, and are necessary for salmonella OM integrity. S Typhimurium bacteria increase their OM cardiolipin concentrations during activation of the PhoPQ regulators. The mechanism involves PbgA's periplasmic globular region (residues 245 to 586), but the biological role of increasing cardiolipins on the surface is not understood. Nonsynonymous polymorphisms in three essential lipopolysaccharide (LPS) synthesis regulators, lapB (also known as yciM), ftsH, and lpxC, variably suppressed the defects in OM integrity, rifampin resistance, survival in macrophages, and systemic colonization of mice in the pbgAΔ191-586 mutant (in which the PbgA periplasmic domain from residues 191 to 586 is deleted). Compared to the OMs of the wild-type salmonellae, the OMs of the pbgA mutants had increased levels of lipid A-core molecules, cardiolipins, and phosphatidylethanolamines and decreased levels of specific phospholipids with cyclopropanated fatty acids. Complementation and substitution mutations in LapB and LpxC generally restored the phospholipid and LPS assembly defects for the pbgA mutants. During bacteremia, mice infected with the pbgA mutants survived and cleared the bacteria, while animals infected with wild-type salmonellae succumbed within 1 week. Remarkably, wild-type mice survived asymptomatically with pbgA-lpxC salmonellae in their livers and spleens for months, but Toll-like receptor 4-deficient animals succumbed to these infections within roughly 1 week. In summary, S Typhimurium uses PbgA to influence LPS assembly during stress in order to survive, adapt, and proliferate within the host environment.

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Dataset Information

Regulation of cathelicidin gene expression: induction by lipopolysaccharide, interleukin-6, retinoic acid, and Salmonella enterica serovar typhimurium infection.

Publications

Regulation of cathelicidin gene expression: induction by lipopolysaccharide, interleukin-6, retinoic acid, and Salmonella enterica serovar typhimurium infection.

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