Abstract
Laminin-5 is the major adhesion ligand of epithelial cells. Mutations in the three genes (LAMA3, LAMB3, LAMC2) encoding the laminin-5 chains cause junctional epidermolysis bullosa, a clinically and genetically heterogeneous blistering skin disease. Here, we describe a non-Herlitz junctional epidermolysis bullosa patient, compound heterozygote for two novel mutations affecting the LAMC2 gene. The mutation in the paternal allele is a de novo splice site mutation (522-1G→A) that results in in-frame skipping of exon 4 and synthesis of a mutated γ2 polypeptide (γ2Δ4) carrying a 33 amino acid deletion within the N-terminal domain V. The maternal mutation is a one base pair insertion (3511insA) in the 3′ terminal exon of LAMC2 resulting in a frameshift and a premature termination codon. Mutation 3511insA is predicted to lead to the synthesis of a γ2 polypeptide (γ2t) disrupted in its α-helical C-terminal structure and truncated of the last 25 amino acids. Keratinocytes isolated from the patient's skin showed a markedly decreased level of γ2 chain mRNA and secreted scant amounts of laminin-5, which undergoes physiologic proteolytic processing. To investigate the biologic function of the laminin-5 molecules synthesized by the patient, mutant γ2 cDNAs were transiently expressed in γ2-null keratinocytes. Transfection of the γ2Δ4 cDNA resulted in restoration of laminin-5 deposition onto the culture substrate, which demonstrates that the γ2 polypeptides carrying a deletion in domain V, upstream of the γ2 proteolytic cleavage site, are assembled into native laminin-5 that is secreted and extracellularly processed. In contrast, transfection of a mutant cDNA expressing the γ2t chain failed to restore laminin-5 immunoreactivity, which indicates that integrity of the γ2 C-terminal amino acid sequences is required for laminin-5 assembly. These results correlate for the first time a functional alteration in a laminin-5 domain with a mild junctional epidermolysis bullosa phenotype.
| Original language | English |
|---|---|
| Pages (from-to) | 731-739 |
| Number of pages | 9 |
| Journal | Journal of Investigative Dermatology |
| Volume | 117 |
| Issue number | 3 |
| DOIs | |
| Publication status | Published - 2001 |
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Keywords
- Inherited blistering skin diseases
- Laminin γ2 chain
- Molecular genetics
ASJC Scopus subject areas
- Dermatology
Cite this
Novel mutations in the LAMC2 gene in non-Herlitz junctional epidermolysis bullosa : Effects on laminin-5 assembly, secretion, and deposition. / Castiglia, Daniele; Posteraro, Patrizia; Spirito, Flavia; Pinola, Mari; Angelo, Corrado; Puddu, Pietro; Meneguzzi, Guerrino; Zambruno, Giovanna.
In: Journal of Investigative Dermatology, Vol. 117, No. 3, 2001, p. 731-739.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Novel mutations in the LAMC2 gene in non-Herlitz junctional epidermolysis bullosa
T2 - Effects on laminin-5 assembly, secretion, and deposition
AU - Castiglia, Daniele
AU - Posteraro, Patrizia
AU - Spirito, Flavia
AU - Pinola, Mari
AU - Angelo, Corrado
AU - Puddu, Pietro
AU - Meneguzzi, Guerrino
AU - Zambruno, Giovanna
PY - 2001
Y1 - 2001
N2 - Laminin-5 is the major adhesion ligand of epithelial cells. Mutations in the three genes (LAMA3, LAMB3, LAMC2) encoding the laminin-5 chains cause junctional epidermolysis bullosa, a clinically and genetically heterogeneous blistering skin disease. Here, we describe a non-Herlitz junctional epidermolysis bullosa patient, compound heterozygote for two novel mutations affecting the LAMC2 gene. The mutation in the paternal allele is a de novo splice site mutation (522-1G→A) that results in in-frame skipping of exon 4 and synthesis of a mutated γ2 polypeptide (γ2Δ4) carrying a 33 amino acid deletion within the N-terminal domain V. The maternal mutation is a one base pair insertion (3511insA) in the 3′ terminal exon of LAMC2 resulting in a frameshift and a premature termination codon. Mutation 3511insA is predicted to lead to the synthesis of a γ2 polypeptide (γ2t) disrupted in its α-helical C-terminal structure and truncated of the last 25 amino acids. Keratinocytes isolated from the patient's skin showed a markedly decreased level of γ2 chain mRNA and secreted scant amounts of laminin-5, which undergoes physiologic proteolytic processing. To investigate the biologic function of the laminin-5 molecules synthesized by the patient, mutant γ2 cDNAs were transiently expressed in γ2-null keratinocytes. Transfection of the γ2Δ4 cDNA resulted in restoration of laminin-5 deposition onto the culture substrate, which demonstrates that the γ2 polypeptides carrying a deletion in domain V, upstream of the γ2 proteolytic cleavage site, are assembled into native laminin-5 that is secreted and extracellularly processed. In contrast, transfection of a mutant cDNA expressing the γ2t chain failed to restore laminin-5 immunoreactivity, which indicates that integrity of the γ2 C-terminal amino acid sequences is required for laminin-5 assembly. These results correlate for the first time a functional alteration in a laminin-5 domain with a mild junctional epidermolysis bullosa phenotype.
AB - Laminin-5 is the major adhesion ligand of epithelial cells. Mutations in the three genes (LAMA3, LAMB3, LAMC2) encoding the laminin-5 chains cause junctional epidermolysis bullosa, a clinically and genetically heterogeneous blistering skin disease. Here, we describe a non-Herlitz junctional epidermolysis bullosa patient, compound heterozygote for two novel mutations affecting the LAMC2 gene. The mutation in the paternal allele is a de novo splice site mutation (522-1G→A) that results in in-frame skipping of exon 4 and synthesis of a mutated γ2 polypeptide (γ2Δ4) carrying a 33 amino acid deletion within the N-terminal domain V. The maternal mutation is a one base pair insertion (3511insA) in the 3′ terminal exon of LAMC2 resulting in a frameshift and a premature termination codon. Mutation 3511insA is predicted to lead to the synthesis of a γ2 polypeptide (γ2t) disrupted in its α-helical C-terminal structure and truncated of the last 25 amino acids. Keratinocytes isolated from the patient's skin showed a markedly decreased level of γ2 chain mRNA and secreted scant amounts of laminin-5, which undergoes physiologic proteolytic processing. To investigate the biologic function of the laminin-5 molecules synthesized by the patient, mutant γ2 cDNAs were transiently expressed in γ2-null keratinocytes. Transfection of the γ2Δ4 cDNA resulted in restoration of laminin-5 deposition onto the culture substrate, which demonstrates that the γ2 polypeptides carrying a deletion in domain V, upstream of the γ2 proteolytic cleavage site, are assembled into native laminin-5 that is secreted and extracellularly processed. In contrast, transfection of a mutant cDNA expressing the γ2t chain failed to restore laminin-5 immunoreactivity, which indicates that integrity of the γ2 C-terminal amino acid sequences is required for laminin-5 assembly. These results correlate for the first time a functional alteration in a laminin-5 domain with a mild junctional epidermolysis bullosa phenotype.
KW - Inherited blistering skin diseases
KW - Laminin γ2 chain
KW - Molecular genetics
UR - http://www.scopus.com/inward/record.url?scp=0034792542&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0034792542&partnerID=8YFLogxK
U2 - 10.1046/j.0022-202x.2001.01453.x
DO - 10.1046/j.0022-202x.2001.01453.x
M3 - Article
C2 - 11564184
AN - SCOPUS:0034792542
VL - 117
SP - 731
EP - 739
JO - Journal of Investigative Dermatology
JF - Journal of Investigative Dermatology
SN - 0022-202X
IS - 3
ER -