Corrales Lorite, S.

Universitat Autònoma de Barcelona. Facultat de Biociències

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S. Corrales Lorite, Genetics Degree, Facultat de Biociències, Universitat Autònoma de Barcelona, Spain.     Transposable elements (TEs) are DNA sequences capable of mobilizing and multiply within the genome of a host organism in both somatic and germ found in many species. In humans, comprise approximately 45% of the genetic material and, therefore, is not an insignificant fact, since although not part of the coding DNA (1.5%) are the largest source of variability among humans. TEs are divided into class I elements or retrotransposons (LTR as retroviruses and non-LTR as LINE, SINE and SVA) and class II or DNA transposons. This work is a review of TEs in humans, particularly LINE-1 retrotransposons (L1) and their impact on the human genome as well as the most characteristic disease-causing mutations caused directly by these, not in trans by other elements as Alu or SVA. The reason to focus only on the L1 retrotransposon mutagenesis is because they are the most abundant elements in whole human genome (16.9%), in addition to being the only currently active and autonomous elements, unlike Alu or SVA, which are active but non-autonomous.       Schematic representation of the structural organization of a human full-length LINE-1 (6 kb) showing 5’UTR, ORF1, ORF2 and 3’UTR with poli(A) tail.    L1 is transcribed by RNA Pol II, then the mRNA is exported to the cytoplasm, and translation of ORF1p and ORF2p leads to ribonucleoprotein (RNP) formation. Components of the L1 RNP are transported to the nucleus, and retrotransposition occurs by target-site primed reverse transcription (TPRT).  During TPRT, the ORF2p endonuclease activity (EN) nicks genomic DNA, exposing a free 3’-OH that can serve as a primer for RT of the L1 mRNA. The process continues with a second-strand cleavage, second-strand cDNA synthesis, and completion of L1 integration. Resolution of the structure results in TSD. Schematic representation of the various ways that L1 retrotransposition events can affecte the human genome. A Hypothetical wild-type gene locus. B Genomic instability caused by L1 EN activity from ORF2p generating dsDNA breaks corrected by Eni retrotransposition. C Structural variation caused by NAHR, transductions or genomic deletions. D Potential effects on gene expression by initiation by L1 promoter, premature polyadenylation or epigenetic alterations. E Disease-causing mutations by disruption of an exon or disruption of correct splicing pattern. INTRODUCTION RETROTRANSPOSITION CYCLE OF LINE-1 DIFFERENT WAYS TO AFFECT THE HUMAN GENOME   DISEASE-CAUSING MUTATIONS CAUSED BY LINE-1 RETROTRANSPOSITION  Most carachteristic disease-causing mutations by L1 described until 2012.  DMD – Duchenne Muscular Dystrophy  DMD – X-Linked Dilated Cardiomyopathy FVIII – Hemophilia A APC – Colon Cancer PDHX– Pyruvate Dehydrogenase complex deficiency CYBB – Chronic  Granulomatous Disease Identification by PCR of a unique de novo L1 5’-truncated insertion in the 5’UTR muscle exon 1 of DMD gene in three XLDCM patients from two unrelated Japanese families which affected the stability of the muscle form of dystrophin transcripts but not that of the brain or Purkinje cell form.  When the PCR was used to amplify a region of the DMD gene encompassing exon 44 from genomic DNA of two Japanese brothers with DMD, it was found to be 600 bp larger than expected.  LINE-1 retrotransposons and their impact on the  human genome and in disease-causing mutations Sequencing of the mRNA disclosed that the sequence of exon 43 was directly joined to that of exon 45, suggesting an exon 44 skipping from the transcript. CONCLUSIONS Schematic representation of de novo insertions of L1 elements into exon 14 of the FVIII gene in two of 240 unrelated patients with hemophilia A (3.8 kb insertion in family JH-27 and 2.3 kb in family JH-28).  Disruption of the APC gene caused by somatic insertion of a L1 into the last exon in a colon cancer. This is the first report of the disruption of a tumor supressor gene caused by somatic insertion of a TE. Identification of the 750 pb insertion was realized by Southern Blot. Analysis were done in normal (N) and tumoral (T) tissue from same patient. Schematic representation of the two hairpin structure (formed by inverted repeats present immediately 5’ to the upstream breakpoint and 3’ to the downstream breakpoint) responsible of a deletion of 46 kb by template jumping model between intron 2 and 9 caused by a L1 full-length L1 insertion. In this patient, CGD is  caused by the insertion of an 1 kb L1 into intron 5 of the CYBB gene.   Transposable elements comprise nearly 50% of the genome, and within these, LINE-1 elements are the most abundant group (16.9% in the entire human genome) and the only active besides autonomous elements.   LINE-1 retrotransposons can affect the genome causing genomic instability, structural variations, potential effects on gene expression or epigenetics and disease-causing mutations.   Disease-causing mutations comprise various mechanisms, such as exonic disruption (colon cancer and hemophilia A), altered splicing (CGD), exon skipping (DMD), large deletion (PDHc deficiency) or transcript instability (XLDCM).   Transposable elements also provide beneficial aspects on the host genome such as gene duplication, genome rearrangements, gene and epigenetic regulation or providing variability that plays a key role throughout the evolution.   With the improvement of technology, it presents a great opportunity to expand knowledge about transposable elements.  The L1 insertion into the intron sequence introduce new splice sites resulting in a highly heterogeneous splicing pattern. No wild-type cDNA was found in the patient. REFERENCES Yoshida K et al. Insertional mutation by transposable element, L1, in the DMD gene results in X-linked dilated cardiomyopathy. Hum Mol Genet. 1998.   Narita N et al. Insertion of a 5' truncated L1 element into the 3' end of exon 44 of the dystrophin gene resulted in skipping of the exon during splicing in a case of Duchenne muscular dystrophy. J Clin Invest. 1993.   Kazazian HH Jr et al. Haemophilia A resulting from de novo insertion of L1 sequences represents a novel mechanism for mutation in man. Nature. 1988.   Miki Y et al. Disruption of the APC gene by a retrotransposal insertion of L1 sequence in a colon cancer. Cancer Res. 1992.   Miné M et al. A large genomic deletion in the PDHX gene caused by the retrotranspositional insertion of a full-length LINE-1 element. Hum Mutat. 2007.   Meischl C et al. A new exon created by intronic insertion of a rearranged LINE-1 element as the cause of chronic granulomatous disease. Eur J Hum Genet. 2000. 

LINE-1 retrotransposons and their impact on the human genome and in disease-causing mutations

http://ddd.uab.cat/pub/tfg/2013/113796/TFG_sergiocorraleslorite.pdf

LINE-1 retrotransposons and their impact on the human genome and in disease-causing mutations

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