L1 As An Arbiter Of Genome Diversification

The sheer mass of Lis in mammalian genomes provides molecular scaffolds that sometimes can be coopted by the host as regulatory sequences for gene expression. For example, the polyadenylation signal of the mouse thymidylate synthetase resides within a Li (105). Similarly, cis-acting sequences that function in the regulation of the human apolipoprotein (A) gene, the human C1D gene, and the humanfactor IX gene are derived from Lis (106-108). The over-representation of Lis on the X-chromosome has led to speculation that they might act as booster elements that function in X-inactivation (109), though Boissinot et al. (110) have offered simpler interpretations to explain this phenomenon. Clearly, the analysis of whole genome sequences in conjunction with comparative genomic and functional studies should accelerate the discovery of retrotransposon sequences that function in host gene expression.

Li also can serve as an agent of genomic diversification by facilitating the movement of non-L1 DNA sequences to new chromosomal locations (Fig. 6). For example, Lis can retrotranspose sequences from their 5' and 3' flanks to new genomic sites in cis by a process called "L1-mediated transduction." Ll-mediated 5' transduction occurs when transcription is initiated from a cellular promoter that resides upstream of a retrotransposition-competent L1 (1). Ll-mediated 3' transduction occurs when the L1 poly (A) signal is bypassed in favor of a stronger downstream poly (A) signal present in flanking genomic DNA (14,37). Retro-transposition of these transcripts allows the "duplication" of genomic DNA to a new chromosomal location. Because of the high frequency of 5' truncation, Ll-mediated 5' transduction is relatively infrequent and only a few examples have been reported in silico and in vitro (1,9,29,38). By comparison, 15-20% of Ta-subfamily Lls present in the HGWD contain 3' transductions, and it has been proposed that L1-mediated transduction is responsible for as much as 1% of human genomic DNA (1,111,112).

The L1-encoded proteins also can mobilize non-autonomous retrotransposons (e.g., Alu and perhaps SVA elements), retrotransposition-defective L1s, cellular mRNAs, and partial mRNA transcripts (Fig. 6) (7-9,113-115). The high copy number of Alu elements indicates that they have evolved a means to efficiently compete with L1 RNA for the L1 RT, perhaps by associating with ribosomes and commandeering the L1 RT during its translation (116). Interestingly, only ORF2p is required for Alu retrotransposition (8), whereas both ORF1p and ORF2p seemingly are required for processed pseudogene formation (7,9). Thus, other proteins in Alu RNPs ( SRP9 and SRP14) may compensate for ORF1p during retrotransposition (see Chapter 1) (117).

Besides mobilizing RNA polymerase II transcripts in trans to generate processed pseudogenes, in silico analyses indicate that template switching between L1 RNA and some RNA polymerase III-derived transcripts can lead to the formation of chimeric pseudogenes (e.g., U6/L1, U3/L1 and 7SL/L1; Fig. 6). Indeed, these data suggest that a cohort of small uracil-rich nuclear RNAs can compete for the L1 retrotransposition machinery during TPRT and demonstrate that non-coding RNAs can be duplicated by retrotransposition (118-120).

The mobilization of sequences by L1-mediated retrotransposition represents a potentially powerful mechanism to generate diversity in randomly mating sexual populations because: (1) the process does not depend on homologous DNA sequences; (2) the relative genomic locations of the "shuffled" sequences are not important; (3) the original donor sequence remains unchanged because the process occurs via an RNA intermediate; and (4) it provides a mechanism to exchange limited amounts of information between different chromosomes (37,121).

Other than simply increasing the amount of "junk DNA" in the genome, L1-mediated retrotransposition occasionally can result in exon shuffling and the formation of new genes. For example, in mice the Cdyl gene produces two transcripts, one that is ubiquitously expressed and one that is testis specific. The human homologs of this gene (CDYL and CDY) are present on chromosome 13 and the Y chromosome, respectively. CDY is a processed pseudogene that is derived from CDYL and arose during primate evolution (122). This duplication resulted in a partitioning of gene expression because CDYL is expressed ubiquitously, whereas CDY exhibits testis specific expression. Similarly, L1-mediated retrotransposition of a cyclophilin A mRNA into the TRIM5 locus in owl monkeys led to the formation of a functional chimeric protein that acts to experimentally restrict HIV infection (123).

Finally, L1 poly (A) tails tend to be unstable genetically and probably serve as seeding grounds for the generation of microsatellite repeats. Consistent with this hypothesis, microsatellites are more often found near the 3' ends of older L1 elements when compared to younger "human-specific" L1s (124). A similar scenario has been observed for Alu elements (125) and is discussed further in Chapters 1 and 3.

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