SVA - structural and functional characterization of a composite primate specific retrotransposon
Final Report Abstract
To draw conclusions on the assembly process that led to the current organization of SVA elements and on their transcriptional regulation, we initiated our study by assessing differences in structures of the 116 SVA elements located on human chromosome 19. The comprehensive analysis of chromosome 19 uncovered 116 SVA elements that could be assorted into seven types of structural variants. Novel variants identified were 3’-truncated elements and a significant number of SVA elements with 5’-flanking sequence transductions. Our genome-wide analysis uncovered 220 5’-transducing SVA elements that originated from 93 diverse genomic source loci. The 93 resultant 5’ transduction groups are composed of one to 84 members. The organization of these elements demonstrates that SVA elements can recruit external heterologous promoters for their own transcriptional regulation, which in turn might affect mobilization frequencies. In a number of cases, 5’-transduced sequences are constituted by spliced cellular mRNAs. Therefore, SVA-mediated 5’ transduction represents a further mechanism of retrotransposon-mediated exon shuffling. SVA-driven transduction of 5’-flanking DNA is likely to be one mechanism of genome evolution via increasing genome plasticity and facilitating new combinations of coding and regulatory sequences. Acquisition of new 5’ sequences may also be a common theme in SVA evolution, with the objective of identifying new genomic components whose incorporation increases the efficiency of SVA mobilization. Next, we set out to investigate if the acquirement of the MAST2 CpG-island played a role in the success of the SVAF1 subfamily, and if transcriptional activity of the MAST2 gene correlates with the expression of functional mammalian non-LTR retrotransposons. We could confirm that human MAST2 transcription peaks in testicular and heart tissues. MAST2 was cotranscribed with the SVAF1 subfamily in human tissues. We found that in human testicles, MAST2 transcription correlates also with the transcription pattern of non-LTR retrotransposon families L1, Alu and SVA, and with the expression of L1 ORF1 protein. We showed that the 324 bp-long MAST2 sequence that is part of SVAF1 subfamily members acts as a positive transcriptional regulator in the human testicular germ cell tumor cell line Tera-1. We conclude that the acquirement of the MAST2 CpG-island might play a positive role in the success of the SVAF1 subfamily. MAST2 sequences at the 5’ end of SVAF1 elements act as positive transcriptional regulator in human germ cells. We observed that in 16 tissue samples representing seven different human tissues, MAST2 was cotranscribed with the members of the SVAF1 subfamily. Methylation status of the MAST2-derived sequences of SVAF1 elements reversely correlates with transcriptional activity of MAST2. Finally, in various testicular tissue samples we observed transcriptional correlation of MAST2 with human retrotransposon families L1, Alu and SVA. Thirdly, we set out to test the hypothesis that the L1 protein machinery is mobilizing SVA elements in trans by establishing an SVA retrotransposition reporter assay in cell culture. We compared the rate of processed pseudogene formation with the trans-mobilization frequencies of two human-specific SVA elements that were identified as potential source elements. We found that SVA RNAs transcribed from the retrotransposition reporter plasmids are trans-mobilized 12- to 300-fold more efficiently than RNA-Pol II transcripts expressed from a pseudogene formation reporter plasmid in HeLa-HA cells. Furthermore, we demonstrate that the hexameric (CCCTCT)n repeat/Alu-like region at the 50-end of canonical SVA elements and the 30-transduced AluSp sequence of an SVA source element have different effects on SVA retrotransposition frequencies in cell culture. Marked SVA de novo insertions were predominantly full-length and exhibited all structural features of L1-mediated retrotransposition that are observed in pre-existing genomic SVA insertions.
Publications
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(2007). APOBEC3 proteins: major players in intracellular defence against LINE-1-mediated retrotransposition. Biochem Soc Trans. 35: 637-42
Schumann, G.G.
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(2009). 5’-Transducing SVA retrotransposon groups spread efficiently throughout the human genome. Genome Res. 19: 1992-2008
Damert, A., Raiz, J., Horn, A.V., Löwer, J., Wang, H, Xing, J., Batzer, M.A., Löwer, R. and Schumann, G.G.
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(2010). Trex-1 deficiency in rheumatoid arthritis synovial fibroblasts. Arthritis Rheum 62: 2673-2679
Neidhart, M., Karouzakis, E., Schumann, G.G., Gay R.E., Gay S.
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(2010). Unique functions of the repetitive transcriptome. Int. Rev. Cell. Mol. Biol. 285: 115-188
Schumann, G.G., Gogvadze, E.V., Osanai-Futahashi, M.,Kuroki, A., Münk, C., Fujiwara, H., Ivics, Z., and Buzdin, A.A.
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(2012). The non-autonomous retrotransposon SVA is trans-mobilized by the human LINE-1 protein machinery. Nucleic Acids Res. 40:1666-83
Raiz, J., Damert, A., Chira, S., Held, U., Klawitter, S., Hamdorf, M., Löwer, J. Strätling, W.H., Löwer, R. and Schumann, G.G.
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(2012). Transcriptional regulation of human-specific SVAF1 retrotransposons by cis-regulatory MAST2 sequences. Gene. 15 May (epub ahead of print)
Zabolotneva, A.A., Bantysh, O., Suntsova, M.V., Efimova, N., Malakhova, G.V, Schumann, G.G., Gayfullin, N. Buzdin, A.A.
