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An intronic alteration of the fibroblast growth factor 10 gene causing ALSG-(aplasia of lacrimal and salivary glands) syndrome.

Scheckenbach K, Balz V, Wagenmann M, Hoffmann TK - BMC Med. Genet. (2008)

Bottom Line: The alteration derogates the regular splice acceptor site and leads to the use of a new splice acceptor site 127 bp upstream of exon 3.Furthermore, no diseased member of the family displayed additional abnormalities that are indicative for the clinically overlapping lacrimo-auriculo-dento-digital syndrome (LADD).This family-based approach revealed an intronic variation of the FGF10 gene causing ALSG-syndrome.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Otorhinolaryngology, Heinrich-Heine-University, Düsseldorf, Germany. scheckenbach@med.uni-duesseldorf.de

ABSTRACT

Background: A combined aplasia, hypoplasia or atresia of lacrimal points and salivary glands is rarely diagnosed. Those patients suffer from epiphora, xerostomia and severe dental caries. This phenotype represents the autosomal-dominant aplasia of lacrimal and salivary glands syndrome (ALSG). Recently, aberrations of the Fibroblast Growth Factor 10 (FGF10) gene have been identified to be causative for this disorder.

Methods: We performed a sequence analysis of the FGF10 gene of a patient with ALSG-syndrome and his also affected brother as well as 193 controls. The FGF10 transcript was analyzed using RNA extracted from primary fibroblasts of the patient's mucosa.

Results: We detected a novel heterozygous sequence variation in intron 2 (c.430-1, G > A) causing the ALSG syndrome. The alteration derogates the regular splice acceptor site and leads to the use of a new splice acceptor site 127 bp upstream of exon 3. The aberration was detected in the genomic DNA derived from two affected brothers, but not in 193 control individuals. Furthermore, no diseased member of the family displayed additional abnormalities that are indicative for the clinically overlapping lacrimo-auriculo-dento-digital syndrome (LADD).

Conclusion: This family-based approach revealed an intronic variation of the FGF10 gene causing ALSG-syndrome. Our results expand the mutational and clinical spectrum of the ALSG syndrome.

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Detection and analysis of an aberrant transcript. 4a: Separation of FGF10 RT-PCR amplificates using sensitive silver-stained polyacrylamid gels reveals additional fragments for the patient's samples but not for control samples. Upper panel, Amplification of the FGF10 transcript using oligonucleotides designed to amplify exon 1 to exon 3 sequences; Lower panel, Amplification of the FGF10 transcript using oligonucleotides designed to amplify exon 2 to exon 3 sequences. C: control; P: patient. 4b: Sequence analysis of the excised and re-amplified additional exon 1/exon 3 fragment. The c.430-1, G > A aberration results in the use of an alternative splice acceptor site 127 bp upstream of exon 3. 4c: Putative effect of the 127 bp insertion on protein translation.
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Figure 4: Detection and analysis of an aberrant transcript. 4a: Separation of FGF10 RT-PCR amplificates using sensitive silver-stained polyacrylamid gels reveals additional fragments for the patient's samples but not for control samples. Upper panel, Amplification of the FGF10 transcript using oligonucleotides designed to amplify exon 1 to exon 3 sequences; Lower panel, Amplification of the FGF10 transcript using oligonucleotides designed to amplify exon 2 to exon 3 sequences. C: control; P: patient. 4b: Sequence analysis of the excised and re-amplified additional exon 1/exon 3 fragment. The c.430-1, G > A aberration results in the use of an alternative splice acceptor site 127 bp upstream of exon 3. 4c: Putative effect of the 127 bp insertion on protein translation.

Mentions: In case that one of the potential acceptor sites is used, RT-PCR amplificates of the affected and the wild type transcript deviate in molecular size. However, FGF10 mRNA amplificates from the patient did not show an anomalous molecular weight by size separation using conventional agarose gel electrophoresis. In contrast, using the more sensitive separation and detection system of silver-stained polyacrylamid gels, we were able to identify aberrant migrating fragments in RT-PCR reactions using two different primer combinations for the patient's samples but not for control samples (Fig. 4a). Re-amplification and sequence analysis of the excised DNA fragments revealed the use of an alternative acceptor splice site upstream of exon 3 that results in the insertion of additional 127 bp into the FGF10 transcript (Fig. 4b). However, this acceptor splice site has not been foretold by any of the splice site prediction algorithms. The insertion leads to a preliminary translation stop codon after the insertion of three novel amino acids following amino acid 143 (Fig. 4c).


An intronic alteration of the fibroblast growth factor 10 gene causing ALSG-(aplasia of lacrimal and salivary glands) syndrome.

Scheckenbach K, Balz V, Wagenmann M, Hoffmann TK - BMC Med. Genet. (2008)

Detection and analysis of an aberrant transcript. 4a: Separation of FGF10 RT-PCR amplificates using sensitive silver-stained polyacrylamid gels reveals additional fragments for the patient's samples but not for control samples. Upper panel, Amplification of the FGF10 transcript using oligonucleotides designed to amplify exon 1 to exon 3 sequences; Lower panel, Amplification of the FGF10 transcript using oligonucleotides designed to amplify exon 2 to exon 3 sequences. C: control; P: patient. 4b: Sequence analysis of the excised and re-amplified additional exon 1/exon 3 fragment. The c.430-1, G > A aberration results in the use of an alternative splice acceptor site 127 bp upstream of exon 3. 4c: Putative effect of the 127 bp insertion on protein translation.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2626586&req=5

Figure 4: Detection and analysis of an aberrant transcript. 4a: Separation of FGF10 RT-PCR amplificates using sensitive silver-stained polyacrylamid gels reveals additional fragments for the patient's samples but not for control samples. Upper panel, Amplification of the FGF10 transcript using oligonucleotides designed to amplify exon 1 to exon 3 sequences; Lower panel, Amplification of the FGF10 transcript using oligonucleotides designed to amplify exon 2 to exon 3 sequences. C: control; P: patient. 4b: Sequence analysis of the excised and re-amplified additional exon 1/exon 3 fragment. The c.430-1, G > A aberration results in the use of an alternative splice acceptor site 127 bp upstream of exon 3. 4c: Putative effect of the 127 bp insertion on protein translation.
Mentions: In case that one of the potential acceptor sites is used, RT-PCR amplificates of the affected and the wild type transcript deviate in molecular size. However, FGF10 mRNA amplificates from the patient did not show an anomalous molecular weight by size separation using conventional agarose gel electrophoresis. In contrast, using the more sensitive separation and detection system of silver-stained polyacrylamid gels, we were able to identify aberrant migrating fragments in RT-PCR reactions using two different primer combinations for the patient's samples but not for control samples (Fig. 4a). Re-amplification and sequence analysis of the excised DNA fragments revealed the use of an alternative acceptor splice site upstream of exon 3 that results in the insertion of additional 127 bp into the FGF10 transcript (Fig. 4b). However, this acceptor splice site has not been foretold by any of the splice site prediction algorithms. The insertion leads to a preliminary translation stop codon after the insertion of three novel amino acids following amino acid 143 (Fig. 4c).

Bottom Line: The alteration derogates the regular splice acceptor site and leads to the use of a new splice acceptor site 127 bp upstream of exon 3.Furthermore, no diseased member of the family displayed additional abnormalities that are indicative for the clinically overlapping lacrimo-auriculo-dento-digital syndrome (LADD).This family-based approach revealed an intronic variation of the FGF10 gene causing ALSG-syndrome.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Otorhinolaryngology, Heinrich-Heine-University, Düsseldorf, Germany. scheckenbach@med.uni-duesseldorf.de

ABSTRACT

Background: A combined aplasia, hypoplasia or atresia of lacrimal points and salivary glands is rarely diagnosed. Those patients suffer from epiphora, xerostomia and severe dental caries. This phenotype represents the autosomal-dominant aplasia of lacrimal and salivary glands syndrome (ALSG). Recently, aberrations of the Fibroblast Growth Factor 10 (FGF10) gene have been identified to be causative for this disorder.

Methods: We performed a sequence analysis of the FGF10 gene of a patient with ALSG-syndrome and his also affected brother as well as 193 controls. The FGF10 transcript was analyzed using RNA extracted from primary fibroblasts of the patient's mucosa.

Results: We detected a novel heterozygous sequence variation in intron 2 (c.430-1, G > A) causing the ALSG syndrome. The alteration derogates the regular splice acceptor site and leads to the use of a new splice acceptor site 127 bp upstream of exon 3. The aberration was detected in the genomic DNA derived from two affected brothers, but not in 193 control individuals. Furthermore, no diseased member of the family displayed additional abnormalities that are indicative for the clinically overlapping lacrimo-auriculo-dento-digital syndrome (LADD).

Conclusion: This family-based approach revealed an intronic variation of the FGF10 gene causing ALSG-syndrome. Our results expand the mutational and clinical spectrum of the ALSG syndrome.

Show MeSH
Related in: MedlinePlus