Athila4 of Arabidopsis and Calypso of soybean define a lineage of endogenous plant retroviruses.

Retrotransposons and retroviruses (collectively referred to as retroelements) replicate by a common mechanism of reverse transcription (for review, see Coffin et al. 1997). Retroelement genomes are delimited by direct long terminal repeats (LTRs), and they encode gag and pol genes, whose products form a particulate replication intermediate wherein reverse transcription takes place. The primary distinguishing feature between the retrotransposons and retroviruses is that the latter have a third gene called envelope (env). env encodes a transmembrane protein that associates with the cell membrane. The replication intermediate buds from the cell as a membrane-bound virion, and Env extends from the virion surface and interacts with cellular receptors to mediate infection. Phylogenetic relationships based on reverse transcriptase amino acid sequences identify six distinct lineages of retroelements (Xiong and Eickbush 1990; Malik 2000). One of these—the vertebrate retroviruses—encodes env genes and is infectious. The five remaining groups are comprised mostly of retrotransposons and include the well-studied Ty1-copia (Pseudoviridae) and Ty3-gypsy (Metaviridae) elements (van Regenmortel et al. 2000), the so-called DIRS1 and BEL groups, and the caulimoviruses (Malik et al. 2000). With the exception of the caulimoviruses and the sparsely populated DIRS1 group, some members of each lineage encode open reading frames (ORFs) with env-like features—most notably transmembrane domains. These include a large number of invertebrate Ty3-gypsy elements (e.g., gypsy, 17.6, 297, and ZAM from Drosophila melanogaster; TOM from Drosophila ananassae; TED from Trichoplusia ni; Yoyo from Ceratitis capitata; for review, see Lerat and Capy 1999), two Ty1-copia elements from plants (i.e., SIRE-1 from Glycine max [soybean] and Endovir from A. thaliana; Laten et al. 1998; Kapitonov and Jurka 1999; Peterson-Burch et al. 2000), and several BEL group elements (e.g., Tas from Ascaris lumbricoides and Cer7 from Caenorhabditis elegans; Felder et al. 1994; Bowen and McDonald 1999). Analyses of env-like genes from the various retroelement groups suggests that env was independently acquired from viruses multiple times during evolution. The env-like ORFs of several insect Ty3-gypsy elements are closely related to env of the bacculoviruses, and for some Cer elements, the env-like gene is related to env of the phleboviruses (Malik et al. 2000). Despite the widespread presence of env-like ORFs and their similarity to known viral env genes, gypsy of D. melanogaster is the only known retroelement outside of the retroviruses for which Env is known to play a role in infection (Kim et al. 1994; Song et al. 1994). In our analysis of the A. thaliana genome sequence, we determined that Athila—a degenerate, centromere-associated retroelement (Pelissier et al. 1995, 1996; Copenhaver et al. 1999)—is a Ty3-gypsy group retrotransposon with an env-like ORF (Wright and Voytas 1998). A related element was also described in Pisum sativum (pea) called Cyclops-2 (Chavanne et al. 1998). Because Cyclops-2 was less degenerate than Athila and prevalent in related legumes, we sought potential functional homologs in soybean. The soybean elements, called Calypso, encode an env-like gene that shares 29% amino acid identity to the corresponding gene of Cyclops-2 (Peterson-Burch et al. 2000). This suggests that the env-like ORF has evolved under functional constraint and likely plays a role in the life cycle of these elements. For simplicity, we refer to Athila and related retroelements as endogenous retroviruses, with the understanding that the biological role of their env-like genes remains to be determined. The sequence degeneracy of the endogenous plant retroviruses described to date has frustrated attempts to define their structural features. However, further characterization of the soybean Calypso elements and completion of the A. thaliana genome sequence has enabled us to construct consensus elements that likely approximate functional elements. Here we report a detailed description of these endogenous retroviruses and provide evidence of their widespread distribution in higher plants.