Background Altered function of laminin-332 (α3β3γ2) consequent to mutations in the LAMA3, LAMB3 and LAMC2 genes causes junctional epidermolysis bullosa non-Herlitz (JEB-nH). JEB-nH patients suffer from skin blistering and have an increased risk of developing aggressive skin carcinomas in adulthood. Laminin-332 is proteolytically processed and its extracellular mature form lacks the α3 chain C-terminal globules 4 and 5 (LG45). The LG45 tandem has cell adhesion and protumorigenic properties. However, mutations that affect this domain are very rare and their functional effects in patients have not been explored to date. Objective To characterize molecularly an adult patient with JEB-nH and altered laminin-332 expression presenting multiple skin carcinomas, and to analyse LG45-mediated biological functions using keratinocytes from the patient. Methods A mutational search in laminin-332 genes was performed by hetero-duplex analysis. LAMA3 mRNA and laminin-332 protein levels in patient keratinocytes were investigated by real-time reverse transcriptase polymerase chain reaction and radioimmunoprecipitation assay, respectively. Keratinocyte migration was examined by scratch and Boyden chamber assays. Results We identified a homozygous LAMA3 mutation, p.Leu1648TrpfsX32, which truncates the last 45 amino acids of the carboxyl terminal LG5 subdomain. Gene expression studies revealed that the mutant transcripts were stable and even increased, precursor laminin-332 molecules were retained intracellularly and the amount of mature extracellular heterotrimers was reduced to about 50%. Finally, the patient's keratinocytes migrated faster than normal keratinocytes. Conclusions Structural disruption of LG5 highlights the critical functions of the LG45 proteolytic region in precursor laminin-332 secretion and keratinocyte adhesion and migration. Perturbation of LG45 function might explain the non-aggressive behaviour of carcinomas in this patient. What's already known about this topic? Exceptionally, homozygous null mutations in laminin-332 genes cause junctional epidermolysis bullosa-non Herlitz (JEB-nH). The C-terminal globules 4 and 5 (LG45) of laminin-332 influence in vitro cell adhesion and promote in vivo squamous cell carcinoma (SCC) progression. What does this study add? A naturally occurring frame-shift mutation in the penultimate LAMA3 exon results in stable mRNA and synthesis of a truncated α3 chain, which causes intracellular protein retention. Secreted and deposited laminin-332 with truncated LG5 globule induces keratinocyte migration. Perturbation of LG45 specific functions is critical for JEB-nH pathogenesis and might influence SCC progression in JEB-nH adults.
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