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Cited by 107Chromosomal and Genetic VariationsGenomic variations and chromosomal abnormalitiesGenomics and Chromatin Dynamics

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Many LINE1 elements contribute to the transcriptome of human somatic cells

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Rangwala, Sanjida H, Zhang, Lili, Kazazian, Haig H·2009·Genome Biology
DOI 10.1186/gb-2009-10-9-r100PMC PMC2768975
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Abstract

BACKGROUND: While LINE1 (L1) retroelements comprise nearly 20% of the human genome, the majority are thought to have been rendered transcriptionally inactive, due to either mutation or epigenetic suppression. How many L1 elements 'escape' these forms of repression and contribute to the transcriptome of human somatic cells? We have cloned out expressed sequence tags corresponding to the 5' and 3' flanks of L1 elements in order to characterize the population of elements that are being actively transcribed. We also examined expression of a select number of elements in different individuals. RESULTS: We isolated expressed sequence tags from human lymphoblastoid cell lines corresponding to 692 distinct L1 element sites, including 410 full-length elements. Four of the expression tagged sites corresponding to full-length elements from the human specific L1Hs subfamily were examined in European-American individuals and found to be differentially expressed in different family members. CONCLUSIONS: A large number of different L1 element sites are expressed in human somatic tissues, and this expression varies among different individuals. Paradoxically, few elements were tagged at high frequency, indicating that the majority of expressed L1s are transcribed at low levels. Based on our preliminary expression studies of a limited number of elements in a single family, we predict a significant degree of inter-individual transcript-level polymorphism in this class of sequence.

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Figure 1 Description of data collection method and overview of results. (a) Diagram of expression tag capture. L1 elements are often naturally transcribed with non-L1 sequences at the 5' and/or 3' end. A 3' RACE adaptor/oligo(dT) primer and L1 specific primer can be used to capture expressed sequence from the 3' end. Similarly, a 5' RACE adaptor and L1 specific primer can capture 5' start sites that occur in non-L1 sequence. PCR is subsequently used to amplify the signal from the expressed tags. (b) Examples of 3' expression tags. Sequences start at the end of the L1 and terminate in 12 adenosines derived from the 3' RACE adaptor. (c) Histogram depicting the distribution of polyadenylation positions of 3' expression tags, relative to the end of the L1. (d) Histogram depicting distribution of 5' start site positions upstream of the 5' end of full-length L1 elements. Negative start sites occur in the 5' UTR downstream of the consensus 5' end of the element. Nt, nucleotides.
Figure 1 Description of data collection method and overview of results. (a) Diagram of expression tag capture. L1 elements are often naturally transcribed with non-L1 sequences at the 5' and/or 3' end. A 3' RACE adaptor/oligo(dT) primer and L1 specific primer can be used to capture expressed sequence from the 3' end. Similarly, a 5' RACE adaptor and L1 specific primer can capture 5' start sites that occur in non-L1 sequence. PCR is subsequently used to amplify the signal from the expressed tags. (b) Examples of 3' expression tags. Sequences start at the end of the L1 and terminate in 12 adenosines derived from the 3' RACE adaptor. (c) Histogram depicting the distribution of polyadenylation positions of 3' expression tags, relative to the end of the L1. (d) Histogram depicting distribution of 5' start site positions upstream of the 5' end of full-length L1 elements. Negative start sites occur in the 5' UTR downstream of the consensus 5' end of the element. Nt, nucleotides.CC BY · PubMed Central
Figure 2 Characterization of L1 at 4p15.32. (a) Diagram of L1 at 4p15.32 and the surrounding region. The arrow designates the L1 transcript. Blue boxes indicate exons of the CD38 gene, with exon number designated. Oligonucleotides CD38-a and CD38-b are indicated. Unmarked triangles indicate the positions of oligonucleotides used in L1 TaqMan qPCR assay. (b) Alignments of L1 at 4p15.32 3' end and related sequences. 'chr 4 short tag' - the major 3' expression tag cloned from this site. 'chr4 long tag' - longer 3' expression tag and 3' RACE sequence cloned from this site. 'chr6 transduction' - paralogous, transduced sequence downstream of L1 on chromosome 6. 'U35' - similar distinct 3' expression tag that cannot be mapped to the human reference genome. 3' end target site duplications are highlighted in blue. Single nucleotide differences in the chromosome 6 sequence are highlighted in dark red. (c) Diagram of the pedigree of the CEPH/UTAH individuals used in this study. (d) Relative expression of the L1 at 4p15.32 in lymphoblastoid cell lines from CEPH individuals. Expression is in arbitrary units normalized to HPRT1. Error bars indicate ± standard deviation from three replicates. (e) Expression of CEPH individuals of the L1 at 4p15.32 compared to flanking exons of CD38, normalized to HPRT1. Expression is plotted on a logarithmic scale so that levels for both amplicons can be clearly visualized. Error bars represent ± standard deviations from three replicates. All data are representative of at least two biological replicates.
Figure 2 Characterization of L1 at 4p15.32. (a) Diagram of L1 at 4p15.32 and the surrounding region. The arrow designates the L1 transcript. Blue boxes indicate exons of the CD38 gene, with exon number designated. Oligonucleotides CD38-a and CD38-b are indicated. Unmarked triangles indicate the positions of oligonucleotides used in L1 TaqMan qPCR assay. (b) Alignments of L1 at 4p15.32 3' end and related sequences. 'chr 4 short tag' - the major 3' expression tag cloned from this site. 'chr4 long tag' - longer 3' expression tag and 3' RACE sequence cloned from this site. 'chr6 transduction' - paralogous, transduced sequence downstream of L1 on chromosome 6. 'U35' - similar distinct 3' expression tag that cannot be mapped to the human reference genome. 3' end target site duplications are highlighted in blue. Single nucleotide differences in the chromosome 6 sequence are highlighted in dark red. (c) Diagram of the pedigree of the CEPH/UTAH individuals used in this study. (d) Relative expression of the L1 at 4p15.32 in lymphoblastoid cell lines from CEPH individuals. Expression is in arbitrary units normalized to HPRT1. Error bars indicate ± standard deviation from three replicates. (e) Expression of CEPH individuals of the L1 at 4p15.32 compared to flanking exons of CD38, normalized to HPRT1. Expression is plotted on a logarithmic scale so that levels for both amplicons can be clearly visualized. Error bars represent ± standard deviations from three replicates. All data are representative of at least two biological replicates.CC BY · PubMed Central
Figure 3 Characterization of L1 at 13q14.2. (a) Diagram of L1 at 13q14.2 and the surrounding region. The arrow designates the L1 transcript. Triangles at F and R indicate positions of oligonucleotides 13q14.2F and 13q14.2R. Blue boxes indicate exons of the RB1 gene, with exon number designated. Oligonucleotides RB1-1 and RB1-2 are indicated. The sequence of the 3' expression tag is provided. (b) Relative expression of the L1 at 13q14.2 in lymphoblastoid cell lines from CEPH individuals. Expression is in arbitrary units normalized to HPRT1. Error bars indicate ± standard deviation from three replicates. (c) Expression of CEPH individuals of the L1 at 13q14.2 compared to flanking exons from RB1, normalized to HPRT1. Expression is plotted on a logarithmic scale so that levels for both amplicons can be clearly visualized. Error bars represent ± standard deviations from three replicates. All data are representative of at least two biological replicates.
Figure 3 Characterization of L1 at 13q14.2. (a) Diagram of L1 at 13q14.2 and the surrounding region. The arrow designates the L1 transcript. Triangles at F and R indicate positions of oligonucleotides 13q14.2F and 13q14.2R. Blue boxes indicate exons of the RB1 gene, with exon number designated. Oligonucleotides RB1-1 and RB1-2 are indicated. The sequence of the 3' expression tag is provided. (b) Relative expression of the L1 at 13q14.2 in lymphoblastoid cell lines from CEPH individuals. Expression is in arbitrary units normalized to HPRT1. Error bars indicate ± standard deviation from three replicates. (c) Expression of CEPH individuals of the L1 at 13q14.2 compared to flanking exons from RB1, normalized to HPRT1. Expression is plotted on a logarithmic scale so that levels for both amplicons can be clearly visualized. Error bars represent ± standard deviations from three replicates. All data are representative of at least two biological replicates.CC BY · PubMed Central
Figure 4 Characterization of L1 at 6p22.2. (a) Diagram of L1 at 6p22.2 and the surrounding region. The arrow designates the L1 transcript. Triangles at F and R indicate positions of oligonucleotides 6p22F and 6p22R. Blue boxes indicate exons of the FAM65B gene, with exon number designated. Oligonucleotides FAM65B-1 and FAM65B-2 are indicated. The sequence of the 3' expression tag is provided. (b) Expression of CEPH individuals GM10861 and GM11994 of the L1 at 6P22.2 compared to flanking exons from FAM65B, normalized to HPRT1. Expression is plotted on a logarithmic scale so that levels for both amplicons can be clearly visualized. Error bars represent ± standard deviations from three replicates. Data are representative of two experimental replicates.
Figure 4 Characterization of L1 at 6p22.2. (a) Diagram of L1 at 6p22.2 and the surrounding region. The arrow designates the L1 transcript. Triangles at F and R indicate positions of oligonucleotides 6p22F and 6p22R. Blue boxes indicate exons of the FAM65B gene, with exon number designated. Oligonucleotides FAM65B-1 and FAM65B-2 are indicated. The sequence of the 3' expression tag is provided. (b) Expression of CEPH individuals GM10861 and GM11994 of the L1 at 6P22.2 compared to flanking exons from FAM65B, normalized to HPRT1. Expression is plotted on a logarithmic scale so that levels for both amplicons can be clearly visualized. Error bars represent ± standard deviations from three replicates. Data are representative of two experimental replicates.CC BY · PubMed Central
Figure 5 Characterization of L1 at 1p22.3. (a) Diagram of L1 at 1p22.3 and the surrounding region. The arrow designates the L1 transcript. Triangles at F and R indicate positions of oligonucleotides 1p22.3F and 1p22.3R. Blue boxes indicate exons of nearest genes SH3GLB1 and CLCA, with exon number designated. The sequence of the 3' expression tag is provided. (b) Relative expression of the L1 at 1p22.3 in lymphoblastoid cell lines from CEPH individuals. Expression is in arbitrary units normalized to HPRT1. Error bars indicate ± standard deviation from three replicates. All data are representative of at least two biological replicates. (c) Bisulfite sequencing of amplicon surrounding the start of transcription of the L1 at 1p22.3 in GM10860, GM11992 and GM11993. Each circle represents a single CpG, while each row is an individual sequence clone. The position of the start of transcription as determined by 5' RACE is indicated. Filled circles indicate lack of bisulfite conversion - that is, cytosine methylation. Grey circles indicate residues that could not be assigned due to sequence polymorphism or poor sequence quality. The CpG at position -120 is unmethylated in the majority of clones from all three individuals.
Figure 5 Characterization of L1 at 1p22.3. (a) Diagram of L1 at 1p22.3 and the surrounding region. The arrow designates the L1 transcript. Triangles at F and R indicate positions of oligonucleotides 1p22.3F and 1p22.3R. Blue boxes indicate exons of nearest genes SH3GLB1 and CLCA, with exon number designated. The sequence of the 3' expression tag is provided. (b) Relative expression of the L1 at 1p22.3 in lymphoblastoid cell lines from CEPH individuals. Expression is in arbitrary units normalized to HPRT1. Error bars indicate ± standard deviation from three replicates. All data are representative of at least two biological replicates. (c) Bisulfite sequencing of amplicon surrounding the start of transcription of the L1 at 1p22.3 in GM10860, GM11992 and GM11993. Each circle represents a single CpG, while each row is an individual sequence clone. The position of the start of transcription as determined by 5' RACE is indicated. Filled circles indicate lack of bisulfite conversion - that is, cytosine methylation. Grey circles indicate residues that could not be assigned due to sequence polymorphism or poor sequence quality. The CpG at position -120 is unmethylated in the majority of clones from all three individuals.CC BY · PubMed Central

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Metadata

Authors
Rangwala, S. H., Zhang, L., & Kazazian, H. H.
Affiliations
University of Pennsylvania (US)
Corresponding author
Kazazian, Haig H
ORCID
Venue
Genome Biology
Publisher
Springer Science and Business Media LLC
Published
2009-09-22
Volume
10
Issue
9
Pages
R100-R100
ISSN
  • 1465-6906 (ISSN-L)
  • 1474-760X
DOI
10.1186/gb-2009-10-9-r100
PMC
PMC2768975
OpenAlex
W2107950128
Cited by
102
References
70
Citations by year
2026: 4 · 2025: 6 · 2024: 7 · 2023: 3 · 2022: 6 · 2021: 6 · 2020: 5 · 2019: 8 · 2018: 1 · 2017: 4 · 2016: 9 · 2015: 2 · 2014: 6 · 2013: 9 · 2012: 10
FWCI
10.52
Type
journal article
Language
en
Open access
Licence
Locations
  • Genome biology (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
Funders
  • University of Pennsylvania
  • National Institutes of Health
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Chromosomal and Genetic Variations
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Genomic variations and chromosomal abnormalities
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Chromatin Dynamics
Keywords
somatic cells, expressed sequence tags, lymphoblastoid cell lines, epigenetic silencing, human genome
MeSH terms
Binding Sites, Cell Line, Chromosomes, Human, Pair 1, Chromosomes, Human, Pair 13, Chromosomes, Human, Pair 4, Chromosomes, Human, Pair 6, Gene Expression Regulation, Humans, Lymphocytes, Transcription, Genetic, Genome, Human, Sequence Analysis, DNA, DNA, Complementary, Long Interspersed Nucleotide Elements, Reverse Transcriptase Polymerase Chain Reaction, Expressed Sequence Tags, Gene Expression Profiling

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