RETROTRANSPOSONS SHAPED NON-REDUNDANT CLLU1 GENE MODULES
(Abstract release date: 05/19/16)
EHA Library. Papamichos S. 06/09/16; 132591; E1042
Dr. Spyros Papamichos
Contributions
Contributions
Abstract
Abstract: E1042
Type: Eposter Presentation
Background
CLLU1 was originally identified as a gene differentiating IgVH-unmutated (IgVH-UM) and IgVH-mutated (IgVH-M) chronic lymphocytic leukemia (CLL) and is now appreciated to represent a potent predictor of overall survival of CLL patients. Whilst CLLU1 is located in a genomic region that is highly expressed in both germinal center B cells and CLL cells, indicating an open-chromatin structure, its transcription is highly restricted. CLLU1 expression is markedly high in IgVH-UM CLL, low in IgVH-M CLL and negligible in normal B-cell subpopulations or any other hematological specimens tested. Alternative splicing of the CLLU1 gene generates 7 transcript variants. Previous studies revealed a powerful linear relation between the expression levels of all the variants, signifying that transcription is orchestrated by a common promoter located upstream of exon 1. Accordingly, it was deduced that (epi)genetic aberrations, occurring during leukemogenesis, could trigger the inordinate activation of this promoter. DNA methylation microarray analysis identified CLLU1 to present lower methylation levels in IgVH-UM CLL cases compared to IgVH-M CLL cases and to normal B-cells. It was shown that the larger proportion of methylated sites occupied CLLU1 gene body, while a strong correlation between CLLU1 body hypomethylation and gene expression was also established.Transposable elements (TEs) carry a markedly high density of CG dinucleotides and remain heavily methylated in normal cells. Aberrations in DNA methylation marks could allow for TEs to exert a significant impact on host gene expression. For example, long terminal repeats (LTRs) of endogenous retroviruses (ERVs) contain transcription regulatory elements that, in the sense orientation, are functional and could initiate transcription of adjacent human genes.
Aims
Document that non-redundant CLLU1 gene modules were evolutionary provided by TEs. Provide a rationale for the intriguing transcriptional profile of the gene.
Methods
CLLU1 genomic sequence was downloaded from the NCBI Gene database and scanned for the presence of integrated TEs by RepeatMasker. EMBOSS Cpgplot was used to identify genomic regions of unusual CG composition located within CLLU1 body. PROMO Version 3.0.2 software, employing version 8.3 of TRANSFAC database, was used to evaluate transcription factor (TF) binding affinity of the region spanning 150 nucleotides (nts) upstream and downstream of CLLU1 transcriptional start site (TSS), assumed to include important transcription regulatory elements. CLLU1 locus syntenic alignments of numerous mammals were downloaded from the UCSC Genome Browser Database. Phylogenetic and molecular evolutionary analyses were conducted using MEGA version 6.
Results
CLLU1 exon 1 is located within a region reported from RepeatMasker to match a sense orientation MLT2B1 LTR, of the ERV-L family. The LTR extends 109 nts upstream of CLLU1 TSS. CLLU1 start codon was evolutionarily provided by a MIR element. CLLU1 polyadenylation signal was also provided by a MIR element. CLLU1 body contains CG-rich genomic content. EMBOSS Cpgplot analysis identified a CpG island in the gene body, which, however, is an AluY element of unusually high CG composition than a bona fide CpG island. Of note is the high Observed/Expected CG ratio within and adjacent to CLLU1 coding DNA sequence. The region spanning 150 nts upstream and downstream of CLLU1 TSS includes a plethora of high-affinity TF binding sites. Analysis identified multiple binding sites for TFs known to play a major role in CLL pathogenesis, namely PAX5, the NF-κB subunit RELA, IRF1 and ELK1. Binding motifs for known oncoproteins (JUN, MYB) as well as for XBP1, a potent TF factor in B cells, were also detected. Of note, XBP1 is known to immensely enhance the transcription of HTLV-I exogenous retrovirus. Phylogenetic analysis revealed that a 4 bp genomic deletion allowed the generation of an ELK1 binding site exclusively in the Hominid genome. Because the DNA sequence adjacent to the site presents null variation between Hominids and also because this genomic region includes no common single nucleotide polymorphisms in human, we assume the occurrence of a selective sweep fostering the creation of the motif.
Conclusion
Session topic: E-poster
Keyword(s): Chronic lymphocytic leukemia, Hypomethylation, Retrovirus, Transcriptional regulation
Type: Eposter Presentation
Background
CLLU1 was originally identified as a gene differentiating IgVH-unmutated (IgVH-UM) and IgVH-mutated (IgVH-M) chronic lymphocytic leukemia (CLL) and is now appreciated to represent a potent predictor of overall survival of CLL patients. Whilst CLLU1 is located in a genomic region that is highly expressed in both germinal center B cells and CLL cells, indicating an open-chromatin structure, its transcription is highly restricted. CLLU1 expression is markedly high in IgVH-UM CLL, low in IgVH-M CLL and negligible in normal B-cell subpopulations or any other hematological specimens tested. Alternative splicing of the CLLU1 gene generates 7 transcript variants. Previous studies revealed a powerful linear relation between the expression levels of all the variants, signifying that transcription is orchestrated by a common promoter located upstream of exon 1. Accordingly, it was deduced that (epi)genetic aberrations, occurring during leukemogenesis, could trigger the inordinate activation of this promoter. DNA methylation microarray analysis identified CLLU1 to present lower methylation levels in IgVH-UM CLL cases compared to IgVH-M CLL cases and to normal B-cells. It was shown that the larger proportion of methylated sites occupied CLLU1 gene body, while a strong correlation between CLLU1 body hypomethylation and gene expression was also established.Transposable elements (TEs) carry a markedly high density of CG dinucleotides and remain heavily methylated in normal cells. Aberrations in DNA methylation marks could allow for TEs to exert a significant impact on host gene expression. For example, long terminal repeats (LTRs) of endogenous retroviruses (ERVs) contain transcription regulatory elements that, in the sense orientation, are functional and could initiate transcription of adjacent human genes.
Aims
Document that non-redundant CLLU1 gene modules were evolutionary provided by TEs. Provide a rationale for the intriguing transcriptional profile of the gene.
Methods
CLLU1 genomic sequence was downloaded from the NCBI Gene database and scanned for the presence of integrated TEs by RepeatMasker. EMBOSS Cpgplot was used to identify genomic regions of unusual CG composition located within CLLU1 body. PROMO Version 3.0.2 software, employing version 8.3 of TRANSFAC database, was used to evaluate transcription factor (TF) binding affinity of the region spanning 150 nucleotides (nts) upstream and downstream of CLLU1 transcriptional start site (TSS), assumed to include important transcription regulatory elements. CLLU1 locus syntenic alignments of numerous mammals were downloaded from the UCSC Genome Browser Database. Phylogenetic and molecular evolutionary analyses were conducted using MEGA version 6.
Results
CLLU1 exon 1 is located within a region reported from RepeatMasker to match a sense orientation MLT2B1 LTR, of the ERV-L family. The LTR extends 109 nts upstream of CLLU1 TSS. CLLU1 start codon was evolutionarily provided by a MIR element. CLLU1 polyadenylation signal was also provided by a MIR element. CLLU1 body contains CG-rich genomic content. EMBOSS Cpgplot analysis identified a CpG island in the gene body, which, however, is an AluY element of unusually high CG composition than a bona fide CpG island. Of note is the high Observed/Expected CG ratio within and adjacent to CLLU1 coding DNA sequence. The region spanning 150 nts upstream and downstream of CLLU1 TSS includes a plethora of high-affinity TF binding sites. Analysis identified multiple binding sites for TFs known to play a major role in CLL pathogenesis, namely PAX5, the NF-κB subunit RELA, IRF1 and ELK1. Binding motifs for known oncoproteins (JUN, MYB) as well as for XBP1, a potent TF factor in B cells, were also detected. Of note, XBP1 is known to immensely enhance the transcription of HTLV-I exogenous retrovirus. Phylogenetic analysis revealed that a 4 bp genomic deletion allowed the generation of an ELK1 binding site exclusively in the Hominid genome. Because the DNA sequence adjacent to the site presents null variation between Hominids and also because this genomic region includes no common single nucleotide polymorphisms in human, we assume the occurrence of a selective sweep fostering the creation of the motif.
Conclusion
- Non-redundant CLLU1 gene modules were provided by TEs thereby remain heavily methylated in normal cells and become susceptible to increased hypomethylation during leukemogenesis. This finding provides a mechanistic link between high CLLU1 expression and poor clinical outcome prognostication since CLLU1 body hypomethylation occurs predominantly in IgVH-UM CLL cases.
- In the Eμ-TCL1 transgenic mouse model, whilst TCL1 mice are expected to develop an IgVH-UM CLL-like disease phenotype at around 11 month, hypomethylation of TEs occurs as early as in 7-months-old mice. In addition, DNA methylation levels remain relatively stable over time among IgVH-UM CLL cases, not affected by treatment. The above, combined with our findings, provide a rationale for CLLU1 expression representing an inherent and constant feature of a particular CLL clone.
Session topic: E-poster
Keyword(s): Chronic lymphocytic leukemia, Hypomethylation, Retrovirus, Transcriptional regulation
Abstract: E1042
Type: Eposter Presentation
Background
CLLU1 was originally identified as a gene differentiating IgVH-unmutated (IgVH-UM) and IgVH-mutated (IgVH-M) chronic lymphocytic leukemia (CLL) and is now appreciated to represent a potent predictor of overall survival of CLL patients. Whilst CLLU1 is located in a genomic region that is highly expressed in both germinal center B cells and CLL cells, indicating an open-chromatin structure, its transcription is highly restricted. CLLU1 expression is markedly high in IgVH-UM CLL, low in IgVH-M CLL and negligible in normal B-cell subpopulations or any other hematological specimens tested. Alternative splicing of the CLLU1 gene generates 7 transcript variants. Previous studies revealed a powerful linear relation between the expression levels of all the variants, signifying that transcription is orchestrated by a common promoter located upstream of exon 1. Accordingly, it was deduced that (epi)genetic aberrations, occurring during leukemogenesis, could trigger the inordinate activation of this promoter. DNA methylation microarray analysis identified CLLU1 to present lower methylation levels in IgVH-UM CLL cases compared to IgVH-M CLL cases and to normal B-cells. It was shown that the larger proportion of methylated sites occupied CLLU1 gene body, while a strong correlation between CLLU1 body hypomethylation and gene expression was also established.Transposable elements (TEs) carry a markedly high density of CG dinucleotides and remain heavily methylated in normal cells. Aberrations in DNA methylation marks could allow for TEs to exert a significant impact on host gene expression. For example, long terminal repeats (LTRs) of endogenous retroviruses (ERVs) contain transcription regulatory elements that, in the sense orientation, are functional and could initiate transcription of adjacent human genes.
Aims
Document that non-redundant CLLU1 gene modules were evolutionary provided by TEs. Provide a rationale for the intriguing transcriptional profile of the gene.
Methods
CLLU1 genomic sequence was downloaded from the NCBI Gene database and scanned for the presence of integrated TEs by RepeatMasker. EMBOSS Cpgplot was used to identify genomic regions of unusual CG composition located within CLLU1 body. PROMO Version 3.0.2 software, employing version 8.3 of TRANSFAC database, was used to evaluate transcription factor (TF) binding affinity of the region spanning 150 nucleotides (nts) upstream and downstream of CLLU1 transcriptional start site (TSS), assumed to include important transcription regulatory elements. CLLU1 locus syntenic alignments of numerous mammals were downloaded from the UCSC Genome Browser Database. Phylogenetic and molecular evolutionary analyses were conducted using MEGA version 6.
Results
CLLU1 exon 1 is located within a region reported from RepeatMasker to match a sense orientation MLT2B1 LTR, of the ERV-L family. The LTR extends 109 nts upstream of CLLU1 TSS. CLLU1 start codon was evolutionarily provided by a MIR element. CLLU1 polyadenylation signal was also provided by a MIR element. CLLU1 body contains CG-rich genomic content. EMBOSS Cpgplot analysis identified a CpG island in the gene body, which, however, is an AluY element of unusually high CG composition than a bona fide CpG island. Of note is the high Observed/Expected CG ratio within and adjacent to CLLU1 coding DNA sequence. The region spanning 150 nts upstream and downstream of CLLU1 TSS includes a plethora of high-affinity TF binding sites. Analysis identified multiple binding sites for TFs known to play a major role in CLL pathogenesis, namely PAX5, the NF-κB subunit RELA, IRF1 and ELK1. Binding motifs for known oncoproteins (JUN, MYB) as well as for XBP1, a potent TF factor in B cells, were also detected. Of note, XBP1 is known to immensely enhance the transcription of HTLV-I exogenous retrovirus. Phylogenetic analysis revealed that a 4 bp genomic deletion allowed the generation of an ELK1 binding site exclusively in the Hominid genome. Because the DNA sequence adjacent to the site presents null variation between Hominids and also because this genomic region includes no common single nucleotide polymorphisms in human, we assume the occurrence of a selective sweep fostering the creation of the motif.
Conclusion
Session topic: E-poster
Keyword(s): Chronic lymphocytic leukemia, Hypomethylation, Retrovirus, Transcriptional regulation
Type: Eposter Presentation
Background
CLLU1 was originally identified as a gene differentiating IgVH-unmutated (IgVH-UM) and IgVH-mutated (IgVH-M) chronic lymphocytic leukemia (CLL) and is now appreciated to represent a potent predictor of overall survival of CLL patients. Whilst CLLU1 is located in a genomic region that is highly expressed in both germinal center B cells and CLL cells, indicating an open-chromatin structure, its transcription is highly restricted. CLLU1 expression is markedly high in IgVH-UM CLL, low in IgVH-M CLL and negligible in normal B-cell subpopulations or any other hematological specimens tested. Alternative splicing of the CLLU1 gene generates 7 transcript variants. Previous studies revealed a powerful linear relation between the expression levels of all the variants, signifying that transcription is orchestrated by a common promoter located upstream of exon 1. Accordingly, it was deduced that (epi)genetic aberrations, occurring during leukemogenesis, could trigger the inordinate activation of this promoter. DNA methylation microarray analysis identified CLLU1 to present lower methylation levels in IgVH-UM CLL cases compared to IgVH-M CLL cases and to normal B-cells. It was shown that the larger proportion of methylated sites occupied CLLU1 gene body, while a strong correlation between CLLU1 body hypomethylation and gene expression was also established.Transposable elements (TEs) carry a markedly high density of CG dinucleotides and remain heavily methylated in normal cells. Aberrations in DNA methylation marks could allow for TEs to exert a significant impact on host gene expression. For example, long terminal repeats (LTRs) of endogenous retroviruses (ERVs) contain transcription regulatory elements that, in the sense orientation, are functional and could initiate transcription of adjacent human genes.
Aims
Document that non-redundant CLLU1 gene modules were evolutionary provided by TEs. Provide a rationale for the intriguing transcriptional profile of the gene.
Methods
CLLU1 genomic sequence was downloaded from the NCBI Gene database and scanned for the presence of integrated TEs by RepeatMasker. EMBOSS Cpgplot was used to identify genomic regions of unusual CG composition located within CLLU1 body. PROMO Version 3.0.2 software, employing version 8.3 of TRANSFAC database, was used to evaluate transcription factor (TF) binding affinity of the region spanning 150 nucleotides (nts) upstream and downstream of CLLU1 transcriptional start site (TSS), assumed to include important transcription regulatory elements. CLLU1 locus syntenic alignments of numerous mammals were downloaded from the UCSC Genome Browser Database. Phylogenetic and molecular evolutionary analyses were conducted using MEGA version 6.
Results
CLLU1 exon 1 is located within a region reported from RepeatMasker to match a sense orientation MLT2B1 LTR, of the ERV-L family. The LTR extends 109 nts upstream of CLLU1 TSS. CLLU1 start codon was evolutionarily provided by a MIR element. CLLU1 polyadenylation signal was also provided by a MIR element. CLLU1 body contains CG-rich genomic content. EMBOSS Cpgplot analysis identified a CpG island in the gene body, which, however, is an AluY element of unusually high CG composition than a bona fide CpG island. Of note is the high Observed/Expected CG ratio within and adjacent to CLLU1 coding DNA sequence. The region spanning 150 nts upstream and downstream of CLLU1 TSS includes a plethora of high-affinity TF binding sites. Analysis identified multiple binding sites for TFs known to play a major role in CLL pathogenesis, namely PAX5, the NF-κB subunit RELA, IRF1 and ELK1. Binding motifs for known oncoproteins (JUN, MYB) as well as for XBP1, a potent TF factor in B cells, were also detected. Of note, XBP1 is known to immensely enhance the transcription of HTLV-I exogenous retrovirus. Phylogenetic analysis revealed that a 4 bp genomic deletion allowed the generation of an ELK1 binding site exclusively in the Hominid genome. Because the DNA sequence adjacent to the site presents null variation between Hominids and also because this genomic region includes no common single nucleotide polymorphisms in human, we assume the occurrence of a selective sweep fostering the creation of the motif.
Conclusion
- Non-redundant CLLU1 gene modules were provided by TEs thereby remain heavily methylated in normal cells and become susceptible to increased hypomethylation during leukemogenesis. This finding provides a mechanistic link between high CLLU1 expression and poor clinical outcome prognostication since CLLU1 body hypomethylation occurs predominantly in IgVH-UM CLL cases.
- In the Eμ-TCL1 transgenic mouse model, whilst TCL1 mice are expected to develop an IgVH-UM CLL-like disease phenotype at around 11 month, hypomethylation of TEs occurs as early as in 7-months-old mice. In addition, DNA methylation levels remain relatively stable over time among IgVH-UM CLL cases, not affected by treatment. The above, combined with our findings, provide a rationale for CLLU1 expression representing an inherent and constant feature of a particular CLL clone.
Session topic: E-poster
Keyword(s): Chronic lymphocytic leukemia, Hypomethylation, Retrovirus, Transcriptional regulation
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