Differential susceptibility of Cx26 mutations associated with epidermal dysplasias to peptidoglycan derived from Staphylococcus aureus and Staphylococcus epidermidis

Abstract

Mutations in Connexin26 (Cx26) give rise to a spectrum of dominantly inherited hyperproliferating skin disorders, the severest being keratitis-ichthyosis-deafness (KID) syndrome, an inflammatory skin disorder, with patients prone to opportunistic infections. We compared the effects of peptidoglycan (PGN) extracted from the skin commensal Staphylococcus epidermidis and the opportunistic pathogen Staphylococcus aureus on interleukin-6 and connexin expression in HaCaT cells (a keratinocyte cell line) and connexin channel activity in HaCaT and HeLa (connexin deficient) cells transfected to express KID and non-KID Cx26 mutations. In both cell types, PGN from S. aureus induced hemichannel activity in cells expressing KID mutants as monitored by ATP release assays following 15-min challenge, while that from S. epidermidis evoked a response in HeLa cells. In KID mutant expressing cells, ATP release was significantly higher than in cells transfected with wild-type Cx26. No ATP release was observed in non-KID mutant transfected cells or in the presence of carbenoxolone, a connexin channel blocker. PGN isolated from S. aureus but not S. epidermidis induced interleukin-6 and Cx26 expression in HaCaT cells following 6-h challenge. Challenge by PGN from S. aureus evoked a greater interleukin-6 response in cells expressing KID mutants than in cells expressing wtCx26 or non-KID mutants. This response returned to basal levels if acute KID hemichannel signalling was blocked prior to PGN challenge. Thus, KID mutants form channels that can be triggered by the pro-inflammatory mediator PGN from opportunistic pathogens but not skin commensals, providing further insight into the genotype-phenotype relationship of Cx26 disorders.

The human Cx26-D50A and Cx26-A88V mutations causing keratitis-ichthyosis-deafness syndrome display increased hemichannel activity

“…The fact that distinct pathologies can arise from different mutations within the same gene has suggested that unique channel activities contribute to deafness and skin disease (17,28,45,49). It has been previously shown by several laboratories that functional expression of the KID mutations Cx26-G12R, Cx26-N14K, Cx26-A40V, Cx26-G45E, and Cx26-D50N all resulted in enhanced hemichannel activity (9,13,21,23,31,37,41). Here we show that two additional KID-causing mutations, Cx26-D50A and Cx26-A88V, also produced greatly increased hemichannel activity in both cRNA-injected Xenopus oocytes and transiently transfected mammalian cells.…”

“…Injection of Cx26-A40V cRNA into Xenopus oocytes resulted in the induction of large membrane currents followed by cell death, suggesting that the mutation increased hemichannel activity (31). Subsequent evaluation of additional KID syndrome mutations has indicated that altered hemichannel activity may be a general pathological mechanism for this syndromic Cx26 disorder (9,13,21,37,41).…”

Connexins and pannexins in the integumentary system: the skin and appendages

“…Connexins have been described to have many channel and nonchannel functions and in the epidermis are involved in the regulation of the differentiation programme, control of innate immunity [33] and wound healing processes [11,34] amongst many other functions. The literature reveals essential roles of gap junctions in cell proliferation, migration and differentiation of keratinocytes and dermal fibroblasts [25,35,36].…”

Pseudomonas aeruginosa–Induced Apoptosis in Airway Epithelial Cells Is Mediated by Gap Junctional Communication in a JNK-Dependent Manner

“…Gap junctions ensure the direct cell-to-cell transfer of ions, sugars, nucleotides, short peptides, and second messengers (8). Bacterial contact, bacterial internalization, and/or bacterial cytotoxins have been found to modulate hemichannel activity and gap junctional communication in the host epithelium, resulting in imbalanced homeostasis (7,9,10). It was recently reported that gap junction channels favor the cell-to-cell spread of an inflammatory response dominated by IL-8 production upon Shigella flexneri internalization in an intestinal epithelial cell line (11).…”