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Multiple splice variants within the bovine silver homologue (SILV) gene affecting coat color in cattle indicate a function additional to fibril formation in melanophores.

Kuehn C, Weikard R - BMC Genomics (2007)

Bottom Line: Thus, the bovine SILV gene can serve as a model for the investigation of the putative additional functions of SILV.Furthermore, the splice variants of the bovine SILV gene represent a comprehensive natural model to refine the knowledge about functional domains in the SILV protein.Our study exemplifies that the extent of alternative splicing is presumably much higher than previously estimated and that alternatively spliced transcripts presumably can generate molecules of deviating function compared to their constitutive counterpart.

View Article: PubMed Central - HTML - PubMed

Affiliation: Research Institute for the Biology of Farm Animals (FBN), Res, Unit Molecular Biology, Wilhelm-Stahl-Allee 2, D-18196 Dummerstorf, Germany. kuehn@fbn-dummerstorf.de

ABSTRACT

Background: The silver homologue(SILV) gene plays a major role in melanosome development. SILV is a target for studies concerning melanoma diagnostics and therapy in humans as well as on skin and coat color pigmentation in many species ranging from zebra fish to mammals. However, the precise functional cellular mechanisms, in which SILV is involved, are still not completely understood. While there are many studies addressing SILV function upon a eumelaneic pigment background, there is a substantial lack of information regarding the further relevance of SILV, e.g. for phaeomelanosome development.

Results: In contrast to previous results in other species reporting SILV expression exclusively in pigmented tissues, our experiments provide evidence that the bovine SILV gene is expressed in a variety of tissues independent of pigmentation. Our data show that the bovine SILV gene generates an unexpectedly large number of different transcripts occurring in skin as well as in non-pigmented tissues, e.g. liver or mammary gland. The alternative splice sites are generated by internal splicing and primarily remove complete exons. Alternative splicing predominantly affects the repeat domain of the protein, which has a functional key role in fibril formation during eumelanosome development.

Conclusion: The expression of the bovine SILV gene independent of pigmentation suggests SILV functions exceeding melanosome development in cattle. This hypothesis is further supported by transcript variants lacking functional key elements of the SILV protein relevant for eumelanosome development. Thus, the bovine SILV gene can serve as a model for the investigation of the putative additional functions of SILV. Furthermore, the splice variants of the bovine SILV gene represent a comprehensive natural model to refine the knowledge about functional domains in the SILV protein. Our study exemplifies that the extent of alternative splicing is presumably much higher than previously estimated and that alternatively spliced transcripts presumably can generate molecules of deviating function compared to their constitutive counterpart.

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Structure of the bovine SILV mRNA including alternative transcripts. Position of exons (open boxes) and introns (solid line) in the SILV genomic sequence are indicated. Protein domains according to [4,31] are shown: SIG: signal peptide, NTD: N-terminal domain, PKD: polycystic kidney domain, RPT: repeat domain, KRG: Kringle-like domain, TM: transmembrane domain, CTD: C-terminal domain, G1, G2, G3: undefined domains. E: SILV exon.* Numbers in the SILV protein diagram represent the first amino acid of the respective domain. ** Numbers for the SILV transcripts represent the first nucleotide for the respective exon or the last constitutive nucleotide of an alternative transcript. fm: frame shift mutation. Length [in bp] of the alternative c SILV transcripts is indicated in red (left), the missing part relative to the constitutive SILV transcript (nucleotides and amino acids) is indicated in green (right).
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Figure 1: Structure of the bovine SILV mRNA including alternative transcripts. Position of exons (open boxes) and introns (solid line) in the SILV genomic sequence are indicated. Protein domains according to [4,31] are shown: SIG: signal peptide, NTD: N-terminal domain, PKD: polycystic kidney domain, RPT: repeat domain, KRG: Kringle-like domain, TM: transmembrane domain, CTD: C-terminal domain, G1, G2, G3: undefined domains. E: SILV exon.* Numbers in the SILV protein diagram represent the first amino acid of the respective domain. ** Numbers for the SILV transcripts represent the first nucleotide for the respective exon or the last constitutive nucleotide of an alternative transcript. fm: frame shift mutation. Length [in bp] of the alternative c SILV transcripts is indicated in red (left), the missing part relative to the constitutive SILV transcript (nucleotides and amino acids) is indicated in green (right).

Mentions: Analysis of several 5'RACE clones from total RNA of eumelaneic, non-dilute (black) skin indicated a sharp peak of transcription start sites (TSS) at position -28 bp to the A of the translation start ATG of the SILV gene (GenBank: EF065525). No further promoters were detected. The SILV transcription start obtained in this study adds an additional 7 bp to the previously deposited bovine SILV cDNA sequence [14]. The cDNA sequence generated by this experiment based on CAP carrying full length mRNA confirmed previous findings about the structure of the bovine SILV gene comprising 2046 bp organized in 11 exons (Figure 1) and encoding 649 amino acids. Aligning the obtained SILV cDNA sequence with the bovine genomic contig NCBI: NW_001495046 [22]) indicated that the bovine SILV gene spans a total of 8107 bp with introns sizes ranging from 108 to 2220 bp. A TTATA motif representing the putative TATA box of the SILV promoter is located 30 bp upstream of the transcription initiation site.


Multiple splice variants within the bovine silver homologue (SILV) gene affecting coat color in cattle indicate a function additional to fibril formation in melanophores.

Kuehn C, Weikard R - BMC Genomics (2007)

Structure of the bovine SILV mRNA including alternative transcripts. Position of exons (open boxes) and introns (solid line) in the SILV genomic sequence are indicated. Protein domains according to [4,31] are shown: SIG: signal peptide, NTD: N-terminal domain, PKD: polycystic kidney domain, RPT: repeat domain, KRG: Kringle-like domain, TM: transmembrane domain, CTD: C-terminal domain, G1, G2, G3: undefined domains. E: SILV exon.* Numbers in the SILV protein diagram represent the first amino acid of the respective domain. ** Numbers for the SILV transcripts represent the first nucleotide for the respective exon or the last constitutive nucleotide of an alternative transcript. fm: frame shift mutation. Length [in bp] of the alternative c SILV transcripts is indicated in red (left), the missing part relative to the constitutive SILV transcript (nucleotides and amino acids) is indicated in green (right).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2099443&req=5

Figure 1: Structure of the bovine SILV mRNA including alternative transcripts. Position of exons (open boxes) and introns (solid line) in the SILV genomic sequence are indicated. Protein domains according to [4,31] are shown: SIG: signal peptide, NTD: N-terminal domain, PKD: polycystic kidney domain, RPT: repeat domain, KRG: Kringle-like domain, TM: transmembrane domain, CTD: C-terminal domain, G1, G2, G3: undefined domains. E: SILV exon.* Numbers in the SILV protein diagram represent the first amino acid of the respective domain. ** Numbers for the SILV transcripts represent the first nucleotide for the respective exon or the last constitutive nucleotide of an alternative transcript. fm: frame shift mutation. Length [in bp] of the alternative c SILV transcripts is indicated in red (left), the missing part relative to the constitutive SILV transcript (nucleotides and amino acids) is indicated in green (right).
Mentions: Analysis of several 5'RACE clones from total RNA of eumelaneic, non-dilute (black) skin indicated a sharp peak of transcription start sites (TSS) at position -28 bp to the A of the translation start ATG of the SILV gene (GenBank: EF065525). No further promoters were detected. The SILV transcription start obtained in this study adds an additional 7 bp to the previously deposited bovine SILV cDNA sequence [14]. The cDNA sequence generated by this experiment based on CAP carrying full length mRNA confirmed previous findings about the structure of the bovine SILV gene comprising 2046 bp organized in 11 exons (Figure 1) and encoding 649 amino acids. Aligning the obtained SILV cDNA sequence with the bovine genomic contig NCBI: NW_001495046 [22]) indicated that the bovine SILV gene spans a total of 8107 bp with introns sizes ranging from 108 to 2220 bp. A TTATA motif representing the putative TATA box of the SILV promoter is located 30 bp upstream of the transcription initiation site.

Bottom Line: Thus, the bovine SILV gene can serve as a model for the investigation of the putative additional functions of SILV.Furthermore, the splice variants of the bovine SILV gene represent a comprehensive natural model to refine the knowledge about functional domains in the SILV protein.Our study exemplifies that the extent of alternative splicing is presumably much higher than previously estimated and that alternatively spliced transcripts presumably can generate molecules of deviating function compared to their constitutive counterpart.

View Article: PubMed Central - HTML - PubMed

Affiliation: Research Institute for the Biology of Farm Animals (FBN), Res, Unit Molecular Biology, Wilhelm-Stahl-Allee 2, D-18196 Dummerstorf, Germany. kuehn@fbn-dummerstorf.de

ABSTRACT

Background: The silver homologue(SILV) gene plays a major role in melanosome development. SILV is a target for studies concerning melanoma diagnostics and therapy in humans as well as on skin and coat color pigmentation in many species ranging from zebra fish to mammals. However, the precise functional cellular mechanisms, in which SILV is involved, are still not completely understood. While there are many studies addressing SILV function upon a eumelaneic pigment background, there is a substantial lack of information regarding the further relevance of SILV, e.g. for phaeomelanosome development.

Results: In contrast to previous results in other species reporting SILV expression exclusively in pigmented tissues, our experiments provide evidence that the bovine SILV gene is expressed in a variety of tissues independent of pigmentation. Our data show that the bovine SILV gene generates an unexpectedly large number of different transcripts occurring in skin as well as in non-pigmented tissues, e.g. liver or mammary gland. The alternative splice sites are generated by internal splicing and primarily remove complete exons. Alternative splicing predominantly affects the repeat domain of the protein, which has a functional key role in fibril formation during eumelanosome development.

Conclusion: The expression of the bovine SILV gene independent of pigmentation suggests SILV functions exceeding melanosome development in cattle. This hypothesis is further supported by transcript variants lacking functional key elements of the SILV protein relevant for eumelanosome development. Thus, the bovine SILV gene can serve as a model for the investigation of the putative additional functions of SILV. Furthermore, the splice variants of the bovine SILV gene represent a comprehensive natural model to refine the knowledge about functional domains in the SILV protein. Our study exemplifies that the extent of alternative splicing is presumably much higher than previously estimated and that alternatively spliced transcripts presumably can generate molecules of deviating function compared to their constitutive counterpart.

Show MeSH
Related in: MedlinePlus