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First description of the complete human xylosyltransferase-I promoter region.

Faust I, Böker KO, Lichtenberg C, Kuhn J, Knabbe C, Hendig D - BMC Genet. (2014)

Bottom Line: Construct associated changes in XYLT1 promoter activity were detected for several sequence variants, whereas serum XT activity was only marginally affected.Our findings describe for the first time the entire XYLT1 promoter sequence and provide new insights into transcriptional regulation of XT-I.Future studies should analyze the impact of regulatory XYLT1 promoter variations on XT-associated diseases.

View Article: PubMed Central - PubMed

Affiliation: Institut für Laboratoriums- und Transfusionsmedizin, Herz- und Diabeteszentrum Nordrhein-Westfalen, Universitätsklinik der Ruhr-Universität Bochum, Bad Oeynhausen, Germany. ifaust@hdz-nrw.de.

ABSTRACT

Background: Human xylosyltransferase-I (XT-I) catalyzes the rate-limiting step in proteoglycan glycosylation. An increase in XYLT1 mRNA expression and serum XT activity is associated with diseases characterized by abnormal extracellular matrix accumulation like, for instance, fibrosis. Nevertheless, physiological and pathological mechanisms of transcriptional XT regulation remain elusive.

Results: To elucidate whether promoter variations might affect the naturally occurring variability in serum XT activity, a complete sequence analysis of the XYLT1 promoter was performed in genomic DNA of healthy blood donors. Based on promoter amplification by a specialized PCR technique, sequence analysis revealed a fragment of 238 bp, termed XYLT1 238*, which has never been described in the human XYLT1 reference sequence so far. In silico characterization of this unconsidered fragment depicted an evolutionary conservation between sequences of Homo sapiens and Pan troglodytes (chimpanzee) or Mus musculus (mouse), respectively. Promoter activity studies indicated that XYLT1 238* harbors various transcription factor binding sites affecting basal XYLT1 expression and inducibility by transforming growth factor-β1, the key fibrotic mediator. A microsatellite and two single nucleotide variants (SNV), c.-403C>T and c.-1088C>A, were identified and genotyped in 100 healthy blood donors. Construct associated changes in XYLT1 promoter activity were detected for several sequence variants, whereas serum XT activity was only marginally affected.

Conclusions: Our findings describe for the first time the entire XYLT1 promoter sequence and provide new insights into transcriptional regulation of XT-I. Future studies should analyze the impact of regulatory XYLT1 promoter variations on XT-associated diseases.

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Related in: MedlinePlus

Sequence alignment of human genomicXYLT1promoter sequences(c.-610 to c.+122). The sequence examined in this study (XYLT1complete, [GenBank Accession Number KM079589]) was compared with the appropriate XYLT1 promoter region of the current human reference sequence [GenBank Accession Number NG_015843.1] and a human shotgun sequence [GenBank Accession Number NW_001838365.2]. Numbers indicate the nucleotide position up- or downstream of the translation initiation site ATG (red color). Matching nucleotides are shaded in yellow, whereas XYLT1238* (c.-363 to c.-126) is marked in bold.
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Fig1: Sequence alignment of human genomicXYLT1promoter sequences(c.-610 to c.+122). The sequence examined in this study (XYLT1complete, [GenBank Accession Number KM079589]) was compared with the appropriate XYLT1 promoter region of the current human reference sequence [GenBank Accession Number NG_015843.1] and a human shotgun sequence [GenBank Accession Number NW_001838365.2]. Numbers indicate the nucleotide position up- or downstream of the translation initiation site ATG (red color). Matching nucleotides are shaded in yellow, whereas XYLT1238* (c.-363 to c.-126) is marked in bold.

Mentions: The identification of this hitherto undescribed XYLT1 promoter fragment, termed XYLT1238*, was verified by sequencing the genomic DNA of 100 healthy blood donors. All PCR products spanned approximately 700 bp, so none of the amplificates displayed the calculated fragment length of 494 bp, referring to the current XYLT1 reference sequence [GenBank Accession Number NG_015843.1]. A sequence alignment of the reference sequence, as well as the consensus sequence of the XYLT1 promoter sequence XYLT1complete, identified in this study, confirmed the absence of XYLT1238* in the published reference sequence (Figure 1). In the following, base numbering depends on promoter sequence XYLT1complete, whereby the first nucleotide of the translation initiation start site was defined as c.+1 and promoter nucleotides were numbered backwards. XYLT1238* was located at position c.-363 to c.-126. By screening data bases for complementary human DNA reference sequences, in silico analysis revealed the deposition of only one shotgun sequence [GenBank Accession Number NW_001838365.2] conforming to XYLT1238*. However, due to a sequence cut off at position c.-162, XYLT1238* is merely partially listed (Figure 1).Figure 1


First description of the complete human xylosyltransferase-I promoter region.

Faust I, Böker KO, Lichtenberg C, Kuhn J, Knabbe C, Hendig D - BMC Genet. (2014)

Sequence alignment of human genomicXYLT1promoter sequences(c.-610 to c.+122). The sequence examined in this study (XYLT1complete, [GenBank Accession Number KM079589]) was compared with the appropriate XYLT1 promoter region of the current human reference sequence [GenBank Accession Number NG_015843.1] and a human shotgun sequence [GenBank Accession Number NW_001838365.2]. Numbers indicate the nucleotide position up- or downstream of the translation initiation site ATG (red color). Matching nucleotides are shaded in yellow, whereas XYLT1238* (c.-363 to c.-126) is marked in bold.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4264549&req=5

Fig1: Sequence alignment of human genomicXYLT1promoter sequences(c.-610 to c.+122). The sequence examined in this study (XYLT1complete, [GenBank Accession Number KM079589]) was compared with the appropriate XYLT1 promoter region of the current human reference sequence [GenBank Accession Number NG_015843.1] and a human shotgun sequence [GenBank Accession Number NW_001838365.2]. Numbers indicate the nucleotide position up- or downstream of the translation initiation site ATG (red color). Matching nucleotides are shaded in yellow, whereas XYLT1238* (c.-363 to c.-126) is marked in bold.
Mentions: The identification of this hitherto undescribed XYLT1 promoter fragment, termed XYLT1238*, was verified by sequencing the genomic DNA of 100 healthy blood donors. All PCR products spanned approximately 700 bp, so none of the amplificates displayed the calculated fragment length of 494 bp, referring to the current XYLT1 reference sequence [GenBank Accession Number NG_015843.1]. A sequence alignment of the reference sequence, as well as the consensus sequence of the XYLT1 promoter sequence XYLT1complete, identified in this study, confirmed the absence of XYLT1238* in the published reference sequence (Figure 1). In the following, base numbering depends on promoter sequence XYLT1complete, whereby the first nucleotide of the translation initiation start site was defined as c.+1 and promoter nucleotides were numbered backwards. XYLT1238* was located at position c.-363 to c.-126. By screening data bases for complementary human DNA reference sequences, in silico analysis revealed the deposition of only one shotgun sequence [GenBank Accession Number NW_001838365.2] conforming to XYLT1238*. However, due to a sequence cut off at position c.-162, XYLT1238* is merely partially listed (Figure 1).Figure 1

Bottom Line: Construct associated changes in XYLT1 promoter activity were detected for several sequence variants, whereas serum XT activity was only marginally affected.Our findings describe for the first time the entire XYLT1 promoter sequence and provide new insights into transcriptional regulation of XT-I.Future studies should analyze the impact of regulatory XYLT1 promoter variations on XT-associated diseases.

View Article: PubMed Central - PubMed

Affiliation: Institut für Laboratoriums- und Transfusionsmedizin, Herz- und Diabeteszentrum Nordrhein-Westfalen, Universitätsklinik der Ruhr-Universität Bochum, Bad Oeynhausen, Germany. ifaust@hdz-nrw.de.

ABSTRACT

Background: Human xylosyltransferase-I (XT-I) catalyzes the rate-limiting step in proteoglycan glycosylation. An increase in XYLT1 mRNA expression and serum XT activity is associated with diseases characterized by abnormal extracellular matrix accumulation like, for instance, fibrosis. Nevertheless, physiological and pathological mechanisms of transcriptional XT regulation remain elusive.

Results: To elucidate whether promoter variations might affect the naturally occurring variability in serum XT activity, a complete sequence analysis of the XYLT1 promoter was performed in genomic DNA of healthy blood donors. Based on promoter amplification by a specialized PCR technique, sequence analysis revealed a fragment of 238 bp, termed XYLT1 238*, which has never been described in the human XYLT1 reference sequence so far. In silico characterization of this unconsidered fragment depicted an evolutionary conservation between sequences of Homo sapiens and Pan troglodytes (chimpanzee) or Mus musculus (mouse), respectively. Promoter activity studies indicated that XYLT1 238* harbors various transcription factor binding sites affecting basal XYLT1 expression and inducibility by transforming growth factor-β1, the key fibrotic mediator. A microsatellite and two single nucleotide variants (SNV), c.-403C>T and c.-1088C>A, were identified and genotyped in 100 healthy blood donors. Construct associated changes in XYLT1 promoter activity were detected for several sequence variants, whereas serum XT activity was only marginally affected.

Conclusions: Our findings describe for the first time the entire XYLT1 promoter sequence and provide new insights into transcriptional regulation of XT-I. Future studies should analyze the impact of regulatory XYLT1 promoter variations on XT-associated diseases.

Show MeSH
Related in: MedlinePlus