title: Filaggrin-dependent secretion of sphingomyelinase protects against staphylococcal α-toxin-induced keratinocyte death.
abstract: BACKGROUND
  The skin of patients with atopic dermatitis (AD) has defects in keratinocyte differentiation, particularly in expression of the epidermal barrier protein filaggrin. AD skin lesions are often exacerbated by Staphylococcus aureus-mediated secretion of the virulence factor α-toxin. It is unknown whether lack of keratinocyte differentiation predisposes to enhanced lethality from staphylococcal toxins.
  
  
  OBJECTIVE
  We investigated whether keratinocyte differentiation and filaggrin expression protect against cell death induced by staphylococcal α-toxin.
  
  
  METHODS
  Filaggrin-deficient primary keratinocytes were generated through small interfering RNA gene knockdown. RNA expression was determined by using real-time PCR. Cell death was determined by using the lactate dehydrogenase assay. Keratinocyte cell survival in filaggrin-deficient (ft/ft) mouse skin biopsies was determined based on Keratin 5 staining. α-Toxin heptamer formation and acid sphingomyelinase expression were determined by means of immunoblotting.
  
  
  RESULTS
  We found that filaggrin expression, occurring as the result of keratinocyte differentiation, significantly inhibits staphylococcal α-toxin-mediated pathogenicity. Furthermore, filaggrin plays a crucial role in protecting cells by mediating the secretion of sphingomyelinase, an enzyme that reduces the number of α-toxin binding sites on the keratinocyte surface. Finally, we determined that sphingomyelinase enzymatic activity directly prevents α-toxin binding and protects keratinocytes against α-toxin-induced cytotoxicity.
  
  
  CONCLUSIONS
  The current study introduces the novel concept that S aureus α-toxin preferentially targets and destroys filaggrin-deficient keratinocytes. It also provides a mechanism to explain the increased propensity for S aureus-mediated exacerbation of AD skin disease.
authors.affiliation: Division of Pediatric Allergy and Immunology, National Jewish Health, Denver, Colo, USA.
authors.forename: Anne M
authors.initials: AM
authors.lastname: Brauweiler
authors.forename: Lianghua
authors.initials: L
authors.lastname: Bin
authors.forename: Byung Eui
authors.initials: BE
authors.lastname: Kim
authors.forename: Michiko K
authors.initials: MK
authors.lastname: Oyoshi
authors.forename: Raif S
authors.initials: RS
authors.lastname: Geha
authors.forename: Elena
authors.initials: E
authors.lastname: Goleva
authors.forename: Donald Y M
authors.initials: DY
authors.lastname: Leung
mesh_terms: D000818:Animals; D001427:Bacterial Toxins / Q000276:immunology / Q000633:toxicity*; D016923:Cell Death / Q000187:drug effects / Q000276:immunology; D002454:Cell Differentiation / Q000187:drug effects / Q000276:immunology; D002470:Cell Survival / Q000187:drug effects / Q000276:immunology; D002478:Cells, Cultured; D003876:Dermatitis, Atopic / Q000276:immunology / Q000382:microbiology / Q000473:pathology; D000091344:Filaggrin Proteins; D006460:Hemolysin Proteins / Q000276:immunology / Q000633:toxicity*; D006801:Humans; D007381:Intermediate Filament Proteins / Q000096:biosynthesis* / Q000172:deficiency / Q000276:immunology; D015603:Keratinocytes / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D012867:Skin / Q000166:cytology / Q000276:immunology / Q000378:metabolism / Q000382:microbiology; D013108:Sphingomyelin Phosphodiesterase / Q000276:immunology* / Q000378:metabolism*; D013203:Staphylococcal Infections / Q000276:immunology / Q000382:microbiology / Q000473:pathology; D013211:Staphylococcus aureus / Q000378:metabolism
chemical_list: D001427:Bacterial Toxins; C000717249:FLG protein, human; D000091344:Filaggrin Proteins; D006460:Hemolysin Proteins; D007381:Intermediate Filament Proteins; C021091:staphylococcal alpha-toxin; D013108:Sphingomyelin Phosphodiesterase