Defective trafficking and cell death is characteristic of skin disease-associated connexin 31 mutations.
Distinct germline mutations in the gene (GJB3) encoding connexin 31 (Cx31) underlie the skin disease erythrokeratoderma variabilis (EKV) or sensorineural hearing loss with/without peripheral neuropathy. Here we describe a number of functional analyses to investigate the effect of these different disease-associated Cx31 mutants on connexon trafficking and intercellular communication. Immunostaining of a biopsy taken from an EKV patient harbouring the R42P mutation revealed sparse epidermal staining of Cx31, and, when present, it had a perinuclear localization. Transfection and microinjection studies in both keratinocytes and fibroblast cell lines also demonstrated that R42P and four other EKV-associated mutant Cx31 proteins displayed defective trafficking to the plasma membrane. The deafness/neuropathy only mutant 66delD had primarily a cytoplasmic localization, but some protein was visualized at the plasma membrane in a few transfected cells. Both 66delD- and R32W-Cx31/EGFP proteins had significantly impaired dye transfer rates compared to wild-type Cx31/EGFP protein. A striking characteristic feature observed with the dominant skin disease Cx31 mutations was a high incidence of cell death. This was not observed with wild-type, R32W 66delD Cx31 proteins. In conclusion, we have identified some key cellular phenotypic differences with respect to disease-associated Cx31 mutations.
Di
Wei-Li
WL
Centre for Cutaneous Research, Barts and the London School of Medicine and Dentistry, Queen Mary, University of London, Whitechapel, London E1 2AT, UK.
Monypenny
James
J
Common
John E A
JE
Kennedy
Cameron T C
CT
Holland
Katalin A
KA
Leigh
Irene M
IM
Rugg
Elizabeth L
EL
Zicha
Daniel
D
Kelsell
David P
DP
D001706:Biopsy; D002450:Cell Communication / Q000502:physiology; D016923:Cell Death / Q000502:physiology; D002462:Cell Membrane; D017630:Connexins / Q000235:genetics* / Q000378:metabolism; D017931:DNA Primers / Q000737:chemistry; D005347:Fibroblasts / Q000378:metabolism / Q000473:pathology; D049452:Green Fluorescent Proteins; D006801:Humans; D015603:Keratinocytes / Q000378:metabolism / Q000473:pathology; D007642:Keratosis / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D016297:Mutagenesis, Site-Directed; D010523:Peripheral Nervous System Diseases / Q000235:genetics* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection
D017630:Connexins; D017931:DNA Primers; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; C068044:GJB3 protein, human; D049452:Green Fluorescent Proteins