COVIRO-298; NO. OF PAGES 11

Retrotransposon replication in plantsAlan H Schulman1,2

Available online at www.sciencedirect.com

ScienceDirect

Retrotransposons comprise the bulk of large plant genomes,

replicating via an RNA intermediate whereby the original,

integrated element remains in place. Of the two main orders,

the LTR retrotransposons considerably outnumber the LINEs.

LINEs integrate into target sites simultaneously with the RNA

transcript being copied into cDNA by target-primed reverse

transcription. LTR retrotransposon replication is basically

equivalent to the intracellular phase of retroviral life cycles. The

envelope gene giving extracellular mobility to retroviruses is in

fact widespread in plants and their retrotransposons.

Evolutionary analyses of the retrotransposons and retroviruses

suggest that both form an ancient monophyletic group. The

particular adaptations of LTR retrotransposons to plant life

cycles enabling their success remain to be clarified.

Addresses1 Institute of Biotechnology, Viikki Biocenter, University of Helsinki, P.O.

Box 65, Helsinki FIN-00014, Finland2 Biotechnology and Food Research, MTT Agrifood Research Finland,

Jokioinen FIN-31600, Finland

Corresponding author: Schulman, Alan H (alan.schulman@helsinki.fi)

Current Opinion in Virology 2013, 3:xx–yy

This review comes from a themed issue on Virus replication in

animals and plants

Edited by Ben Berkhout and Kuan-Teh Jeang

1879-6257/$ – see front matter, # 2013 Elsevier B.V. All rights

reserved.

http://dx.doi.org/10.1016/j.coviro.2013.08.009

IntroductionThe sequencing and annotation of increasingly large

plant genomes over the past five years has made clear

that they are comprised mostly of transposable elements

(TEs) rather than the genes directing metabolism,

environmental response, and morphogenesis. TE families

with many thousands of members account for 80% or

more of the total DNA of large genomes such as barley,

wheat or maize [1–3]. Even small genomes such as that of

B. distachyon have hundreds of different TE families

containing several to hundreds of members [4��]. While

many genome projects see TEs only as barriers to assem-

bly and inconveniences to gene annotation, their ubiquity

and abundance makes examining their means of replica-

tion and consequences for the genome worthwhile [5�].The manifold consequences of their replication for the

genome and plant include genome size growth [6,7�],insertional mutagenesis [8,9], imposition of epigenetic

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silencing on neighbouring genes [10,11], duplication and

transposition of genes or gene segments [12,13�,14], and

function as promoters and regulatory signals [10,15�,16]

The TEs fall into two main groups [17]. Class I contains

the retrotransposons, which first replicate via an RNA

intermediate and then propagate daughter copies in a

‘copy-and-paste’ process, whereas the Class II transpo-

sons do not replicate as part of their transpositional

process, but rather move as DNA usually in a ‘cut-and-

paste’ process. Although the Class I TEs are united by a

life cycle involving the copying of a genomic RNA into

dsDNA by reverse transcriptase (RT), the five orders of

Class I [17] otherwise differ dramatically in their means of

replication. The LTR (Long Terminal Repeat) retro-

transposon order is the most abundant in most plant

genomes [18]. Among the Class I TEs not containing

LTRs, the LINEs (Long Interspersed Nuclear Elements,

Figure 1) are the best studied due to their importance in

mammals [19,20].

Retrotransposons amount to small genomes within the

organismal genome, containing sufficient information to

replicate using the transcription and translation machin-

ery of their ‘host’ to thrive. The viral research com-

munity thereby has come to regard the retrotransposons

as intracellular viruses and named them accordingly

[21,22�]. Hence, the two major superfamilies of LTR

retrotransposons [17], Gypsy and Copia, respectively

belong to the families Metaviridae and Pseudoviridae.

From the perspective of molecular evolution, however,

the retroviruses (family Retroviridae of the virus

nomenclature) are offshoots of retrotransposons [17],

both being derived from cellular replication machinery.

Both viewpoints shed light on understanding of retro-

transposon replication.

Replication of LINEsWith their simple structure (Figure 1), the LINEs are

thought to be some 600 MY old and the most ancient of

eukaryotic retrotransposons, originating in the Pre-cam-

brian [23]. The basic forms specify only RT and endo-

nuclease activities. Use of RT for replication may be a

vestige of the transition from primordial RNA genomes to

the DNA genomes of current biota [24]. The LINEs

occur virtually ubiquitously in the plants [25,26]; all

appear to belong to the clade formed by the LINE-1

(also called L1) element of mammals, the best studied

LINE [27]. LINEs are generally a minor component in

plant genomes compared to animal genomes, LTR retro-

transposons being the major component in plants. How-

ever, in sugar beet (Beta vulgaris) the L1 group is

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Current Opinion in Virology 2013, 3:1–11

2 Virus replication in animals and plants

COVIRO-298; NO. OF PAGES 11

Figure 1

5′ UTR 3′ UTR (A)nape ORF1

LINE(a)

(b) LTR retrotransposon

(A)n

SINE

gag ap in Copia

Gypsygag ap rt-rh in

LTR LTR PBS PPT TRIM

LTR

LTRPBS PPT

env

rt-rh

non-coding LARD

ap in BARE2rt-rh(gag)

LTR

rh-rh

Current Opinion in Virology

Main groups of retrotransposons. (a) Autonomous and non-autonomous non-LTR of the LINE order and the non-autonomous SINE order. A grey bar

indicates a non-coding domain. (b) Autonomous and non-autonomous LTR retrotransposons. An LTR retrotransposon comprises: the long terminal

repeats (LTRs); the primer binding site (PBS), which is the (�)-strand priming site for reverse transcription; the polypurine tract (PPT), which is the (+)-

strand priming site for reverse transcription. The PBS and PPT are part of the internal domain, which in autonomous elements includes the protein-

coding ORFs. Below, the major superfamilies Gypsy and Copia. The position of the envelope (env) domain in those Gypsy and Copia clades that

contain it is shown. Bottom, the non-autonomous retrotransposons. BARE2 is an example of a group having a specific deletion that generates a non-

autonomous subfamily (here, of BARE1). LARD elements have a long internal domain with conserved structure but lacking coding capacity. TRIM

elements have virtually no internal domain except for the PBS and PPT signals.

abundant; in lily (Lilium speciosum), it comprises 4% of the

genome and 250 000 copies [28], which is still short of the

517 000 LINE-1 copies that occupy 17% of the human

genome [29].

Replication of LINEs in plants has not been well studied.

Given the similarity of LINE-1 to plant elements and the

detailed analysis of LINE-1 in mammals, LINE-1 repli-

cation in mammals serves as the model for LINEs in

plants. LINE-1 contains an internal Pol II promoter

within the 50 untranslated region (50 UTR) as well as

two open reading frames, ORF1 and ORF2 [30,31]. Being

internal, the LINE promoter propagates with the rest of

the element and is not lost during replication via RNA.

The LTR retrotransposons, generally lacking internal

promoters as detailed below, have a very different and

arguably more complicated way of solving this problem.

In contrast, the transposase of the directly transposing

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‘cut-and-paste’ Class II transposons merely cuts outside

of the Pol II promoter, mobilizing it together with the

transposase ORF. Once transcribed and transported to

the cytoplasm, ORF1 and ORF2 LINEs are translated

(Figure 2). ORF1 encodes the 40 kDa protein ORF1p,

which is essential for replication and functions as a nucleic

acid chaperone and RNA-binding protein [32]. ORF2

encodes a 150 kDa protein, ORF2p, possessing both

endonuclease and RT activities [30].

LINE RT belongs to a large family of enzymes including

the RTs of LTR retrotransposons, retroviruses, and

pararetroviruses, the PLE elements, as well as telo-

merases, all being descended from a common ancestor

[33��]. Consistent with a common origin, LINE RT can

prime reverse transcription from oligonucleotides that

mimic telomere ends [34]. Moreover, primitive LINEs

without the endonuclease function may have been able

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Retrotransposon replication in plants Schulman 3

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Figure 2

TTTTTTAAAAA

AAAAA

AAAAA

(1)

(2)

(3)

(4)

(5)

(6)

(7)

5′ UTR 3′ UTR (A)nape ORF1

rh-rh

TTTTTTAAAAA

TTTTTTAAAAA

TTTTTTAAAAA

Current Opinion in Virology

Replication mechanism of a LINE. The element contains ORF1, specifying an RNA-binding protein, and an open reading frame encoding an apurinic

endonuclease (APE) and the reverse transcriptase (RT) - RNAseH (RH) complex. The LINE is transcribed (Step 1), translated (Step 2; only the RT is

shown), assembled into a ribonucleoprotein particle (Step 3), and transported into the nucleus (not shown). The APE nicks the target site, at which

point the RNA anneals (Step 4). The free 30 hydroxyl group of the nicked target is used to prime reverse transcription by target-primed reverse

transcription (Step 5). The other strand of the target DNA is also nicked, and the second strand of the LINE is synthesized by the RT, generating a target

site duplication (Step 6). The process is completed by strand exchange and the new copy is now inserted at the target site (Step 7). The process is

reviewed elsewhere in detail [20,32].

to replicate as telomeres [35]. Furthermore, mobile orga-

nellar and prokaryotic group II introns, thought to be the

ancestors of sliceosomal introns, specify an RT structu-

rally and functionally similar to that of LINE RT [36].

The ORF1p and ORF2p proteins preferentially associate

with the RNAs from which they are translated, forming

ribonucleoprotein (RNP) particles in the cytoplasm

[37��]. The ORF1p may function analogously to GAG

capsid protein of LTR retrotransposons, described below

[32]. To complete the life cycle, the RNP must be moved

back to the nucleus. The details of that process remain

controversial; active RNP transport across the nuclear

membrane may be required or, alternatively, passive

uptake occurs following disruption of the nuclear envel-

ope during cell division [38,39].

Once at the chromosome, integration of LINE elements

into the genome proceeds via target-primed reverse tran-

scription (TPRT), in which the RNA is converted into

cDNA and the cDNA concomitantly inserted into the

genome [32,40]. The endonuclease domain of ORF2p

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nicks a target site in the genome, generally TTTT j AA.

The TTTT 30 OH anneals to the polyA tail of LINE

RNA and is then extended by the RT activity of ORF2p,

copying the LINE RNA in the process (Figure 2). The

second-strand cDNA is then copied from the first strand

by a similar process involving nicking of the opposite host

strand, annealing of the nicked 30 end of the genomic

DNA to the first-strand cDNA, and then extension by

RT. Completion of the LINE replication cycle is essen-

tially double-strand break repair via non-homologous end

joining, an error-prone process involving enzymes not

encoded by the TE itself [41].

Replication of SINEsPlant SINEs (Short INterspersed Elements), like those in

other organisms, constitute a diverse and polyphyletic set

of non-autonomous elements, which can be mobilized

and propagated by the enzymes of LINEs [19,42,43]. The

SINEs include various tRNA, rRNA, and other polymer-

ase III transcripts [42,44] ranging from 75 to 662 bp [43].

SINEs derived from tRNA have the tRNA sequence at

their 50 end and homology at their 30 ends to a LINE from

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4 Virus replication in animals and plants

COVIRO-298; NO. OF PAGES 11

the same genome, which is thought to provide binding

sites for LINE-encoded proteins. The 30 tails are often

AT-rich, mirroring their origins as reverse-transcribed

transcripts. Found in up to a million copies in mammalian

genomes [44], they are widespread in plants [43,45], and

tend to number in the hundreds to low thousands (SINE

Base, http://sines.eimb.ru/), the SINEs appearing to be

most prevalent in tobacco and other Solanaceae [42].

Replication of SINE elements has been best examined

for Alu, a 7SL-derived element and the most prevalent of

the SINEs in mammalian genomes [31,46]. Following

transcription by Pol III, Alu RNA is localized to RNP

complexes [47], together with the LINE-1 proteins. The

endonuclease and RT activities of ORF2p cooperate to

replicate Alu in a way analogous to LINEs. The poly(A)

tail needed for this process is already present in the

integrated SINE DNA copy. In the plants, replication

of S1, a tRNA-related SINE of Arabidopsis, has been

most closely examined [48]. The processing of its RNA

appears to be very close to that of mammalian 7SL-related

SINEs such as Alu [48].

Replication of LTR retrotransposonsThe structure and replication of plant LTR retrotranspo-

sons is reminiscent of the related Gypsy and Copiaelements of fungi and animals as well as of the retro-

viruses and endogenous retroviruses. The 50 LTR directs

transcription and contains a Pol II promoter. The 30 LTR

provides the signals for RNA termination and polyade-

nylation. LTRs carry response elements directing the

conditions under which retrotransposon replication can

commence. Plant LTR promoters are activated by a

variety of biotic and abiotic stresses as well tissue culture

and plant hormone treatment [49–53].

Transcription by Pol II begins within the 50 LTR, down-

stream from the promoter, and terminates within the 30

LTR before its 30 end. Transcription has been examined

in detail for BARE1 of barley [54] and Tnt1 of tobacco

[55,56]. In barley, multiple pools of polyadenylated and

non-polyadenylated RNAs are produced, respectively for

translation and reverse transcription [57��]. The tran-

scripts of LTR retrotransposons must serve both trans-

lation and reverse transcription. Transcription yields

RNA lacking complete LTRs at either end, because

the promoter and terminator are within the 50 and 30

LTRs respectively. While this does not hinder trans-

lation, the restoration of both LTRs is needed to produce

an integrationally and replicationally competent cDNA.

This is resolved by the complex reverse transcription

mechanism described below.

Following transcription, the retrotransposon RNA is

transported to the cytoplasm (Figure 3). For BARE and

most other plant retrotransposons of the Copia super-

family, translation is effected from a single ORF encoding

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Current Opinion in Virology 2013, 3:1–11

both the Gag capsid protein and the polyprotein [58–60].

The polyprotein contains an aspartic proteinase (AP),

integrase (IN), RT, and RNaseH (RH). Antibodies to

BARE1 Gag can detect, in barley and other grasses, both

the polyprotein and the subsequent endoproteolytic clea-

vage products generated by the AP [61,62]. While vir-

tually all animal retrotransposons of the Gypsy superfamily

use �1 translational frameshifting at the end of gag to

translate pol, most plant Gypsy elements appear to employ

a single ORF spanning gag and pol, with a few containing a

+1 frameshift or a stop codon between the two ORFs [60].

In the classical model for retrotransposon and retrovirus

replication (Figure 3), the translated and processed Gag

binds and encapsidates the retrotransposon RNAs into a

virus-like particle (VLP) together with RT-RNAseH and

IN [63,64��]. Two RNA molecules are packaged into each

VLP, directed by the dimerization (DIS) and packaging

(PSI) signals [65]. In plants, these signals have been

identified only for BARE1 and BARE2 of barley [58].

Evidence for formation of virus-like particles has, how-

ever, been demonstrated only for the native BARE1 in

barley [62] and the tobacco Tto1 under an inducible

promoter in Arabidopsis [66].

The encapsidation of the RNA represents a branchpoint

between translation and reverse transcription in the retro-

transposon life cycle (Figure 3). During reverse transcrip-

tion, RNaseH degrades the RNA template. Hence, the

RNA must be translated first if the same molecule is to

serve as a template both for cDNA and protein. Reverse

transcription is primed generally from a tRNA matching

the primer binding site (PBS) flanking the 30 end of the 50

LTR (Figures 1 and 4). A (�)-strand strong-stop cDNA is

synthesized, which cannot be further extended without a

jump to the 30 end of the template. Once the (�)-strand

reaches the polypurine tract (PPT) on the 50 side of the 30

LTR within the RNA, synthesis of the (+)-strand cDNA

begins, likewise requiring a template jump in order to

complete (Figure 4). The two strand jumps homogenize

the ends of the final double-stranded cDNA, filling in the

missing 50 and 30 segments of the 50 and 30 LTRs respect-

ively as well as making the two LTRs identical. Detailed

verification of the steps of reverse transcription has not

been made for plant retrotransposons as they have for the

model Ty1 of yeast [67]. However, given both the struc-

tural conservation of plant retrotransposons for features

such as the PBS [68] and the presence of expected cDNA

intermediates in a transgenic test system [69], the mech-

anisms are likely very similar.

In order to be integrated into the chromosome, the retro-

transposon cDNA must find its way into the nucleus

(Figure 3). Nuclear entry is moderated by the nuclear

localization signal (NLS), which in retroviruses can be

found in various retroviral proteins [70,71]. For fungal

retrotransposons, the NLS has been found in both Gag

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Figure 3

AU

G

PBS + PSI-DS

gag ORFAAA

pol ORF

AAA

AAA

AAA

INRHRT

GAG

GAGGAG

GAG

INRHRTAPGAG

AAA

AAA

1

2

3

46

7

8

5

LTR

LTR

gag

ap

in

rt-rh

Current Opinion in Virology

Lifecycle of LTR retrotransposons. An element of superfamily Copia is shown integrated into the genome, within the nucleus (magenta curve). The

steps are: (1) transcription of a copy integrated into the genome from the promoter in the long terminal repeat (LTR); (2) nuclear export; (3) translation

or, alternatively, packaging of two transcripts into a virus-like particle (VLP); (4) translation either of separate gag and pol ORFs or of one common ORF

to produce the capsid protein Gag and a polyprotein containing aspartic proteinase (AP), RT, RNAseH (RH), and integrase (IN), the order of the protein

as in elements of superfamily Gypsy; (5) assembly of a VLP from Gag containing RNA transcripts, IN, RT-RH; (6) reverse transcription by RT; (7)

localization of the VLP to the nucleus; (8) passage of the cDNA – IN complex into the nucleus and integration of the cDNA into the genome. The details

are essentially as presented earlier [91,115].

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6 Virus replication in animals and plants

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Figure 4

R AAAA

U5 PBS U3 R PPT

U5 R

5′R

(a)

(b)

(c)

(d)

(e)

(f)

(g)

(h)

AAAA U5 PBS PPT internal domain

3′U3 R

AAAA U3 R PPT U5 R PBS

U5 R

AAAA U3 R PPT PBS

U5 R

U3 R PBS

U5 R U3 PPT

PPT U3 R U5

U3 R PBS

U5 R U3 PPT

PBS

U5

U3

U3 R U5

R

internal domain

internal domain

internal domain

internal domain

internal domain

U5 R U3 PPT

U3 R U5

U5 R U3

U3 R U5

PBS

PBS PPT

internal domain

internal domain

PBS

PBS

PPT

U3

R U

5

PBS internal dom

ain

Current Opinion in Virology

Reverse transcription of LTR retrotransposons. (a) Attachment of a tRNA primer at the primer-binding site (PBS) of the retrotransposon transcript

(black line), adjacent to the 30 end of the 50 LTR regions R and U5, to initiate reverse transcription. (b) Extension of the minus-strand cDNA (shown as a

purple line) to the end of the transcript to form minus-strand strong-stop DNA (�sssDNA); (c) Degradation of the RNA from the RNA/DNA hybrid by

RNaseH, exposing the repeat (R) domain that is present at both ends of the transcript. (d) Strand jumping of the exposed �sssDNA to the 30 end of the

transcript mediated by hybridization of the R domain. (e) Extension of the minus-strand and degradation of the hybridized regions of the RNA by

RNase H, ultimately exposing the polypurine tract (PPT) of the (�)-strand cDNA, so that plus-strand cDNA (red line) synthesis begins from

complementary PPT RNA fragments (short black lines) as primers. The plus strand is extended to the 50 end of the minus-strand cDNA, thereby

forming a complementary copy of the PBS and producing plus-strand strong-stop DNA (+sssDNA). (f) The RNA primers are removed by RNAseH,

exposing the PBS on the +sssDNA. (g) Strand jumping of the +sssDNA, mediated by hybridization of the PBS domains, and continuation of cDNA

synthesis, each strand serving as a template for the other. (h) Completion of cDNA synthesis to generate a double-stranded linear molecular with intact

LTRs at either end. The details and representation are essentially as presented earlier [91,116].

[72] and IN [73]. There is very little information on NLS

signals in plant retrotransposons. While we have evidence

that BARE Gag contains an NLS (Gomez-Orte et al.,unpublished), the only other clue is for Grande of maize

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Current Opinion in Virology 2013, 3:1–11

and other Zea spp., where gene23, transcribed under its

own promoter from the opposite strand as gag and pol,encodes a functional NLS. Its precise role in localizing

Grande cDNA to the nucleus remains unclear [74].

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In contrast to the LINEs and SINEs, integration of

retrotransposons into the genome is accomplished by a

specialized enzyme, integrase. The IN makes staggered

cuts at the target site, trims extra nucleotides from the 30

termini of the LTRs, and also joins the 30 termini to the

free 50 ends at the staggered cuts. The mechanism of

integration appears to be highly conserved among retro-

transposons and retroviruses, with retroviral IN forming

the interpretive model for the others [75]. Moreover, IN

and bacteriophage Mu transposase share the catalytic

DDE motif and other structural features, implying a

common origin [76–78]. Mechanistic studies on retro-

transposon INs have advanced farthest with those of yeast

Ty1 and Ty3, in keeping with the general advanced state

of these systems [79]. Unravelling the target specificity of

Ty3 IN for integration at Pol III transcription initiation

sites has been of particular interest [80�] as has been,

conversely, the question of substrate specificity for Ty1IN [81].

There has been little functional investigation of plant

INs, although structural predictions for the BARE IN

have been made [82] and processing of cDNA by Tnt1 IN

examined [69]. Plant IN studies have focused, instead, on

evolutionary perspectives for the diversity present in

large retrotransposon families [83]. The INs of the wide-

spread chromovirus group of Gypsy elements [84] are

interesting for possession at their C-terminus of chromo-

domains, structurally similar to members of the hetero-

chromatin protein 1 (HP1) family, which can confer

particular integration patterns to the various clades of

chromoviruses [85]. Functional analysis of the chromo-

domain has been made in a yeast system [86]. For plants,

sequence analysis of chromovirus IN has been the

primary approach [85,87–89], although transcription and

propagation of a whole element, LORE1a, has been

examined in the model legume Lotus japonica [90�].

Replication of non-autonomous LTRretrotransposonsAnalyses of plant genomes tend to reveal large numbers

of LTR retrotransposons that contain stop codons inter-

rupting their ORFs, lack one or more coding domains, or

both. These non-autonomous retrotransposons may none-

theless be transcriptionally competent. For non-autonom-

ous LTR retrotransposons, the steps of packaging,

reverse transcription, nuclear localization, and integration

may be blocked by lack of self-encoded proteins. Never-

theless, all steps potentially can be complemented in

trans if a translationally or enzymatically defective retro-

transposon possesses the correct recognition signals for

proteins encoded by autonomous elements [63,91].

Among non-autonomous elements in plants are con-

served, abundant, and insertionally polymorphic families

such as BARE2, which receives Gag, which it cannot

produce, from BARE1 [58]. In contrast, the Large

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Retrotransposon Derivative (LARD) elements have

replaced gag and pol with a long internal domain of

conserved RNA structure having no coding capacity

[92]. The Terminal Repeat retrotransposons In Miniature

(TRIMs) have non-coding internal domains reduced to

just 100–300 bp, but are nevertheless abundant, wide-

spread, active, and insertionally polymorphic in plants

[93–95]. The Cassandra family of TRIMs is both inser-

tionally polymorphic, distributed throughout the plant

kingdom, and transcribed by a Pol III promoter from 5S

sequences found in its LTRs [96,97]. Non-autonomous,

structurally conserved retrotransposons such as BARE2,

TRIMs, and Cassandra appear to represent replicational

strategies based on parasitizing proteins from other retro-

transposons. However, if the host retrotransposons pro-

viding proteins in trans lose their activity, their non-

autonomous partners will become inactive. This appears

to have happened to TRIMs in rice [98].

Control of retrotransposon replicationThe complex lifecycles of LINEs and LTR retrotran-

sposons present multiple points for regulation and epi-

genetic silencing of activity in order to control the many

consequences of replication that were mentioned at the

outset. Indeed, plant retrotransposons can be variously

silenced both transcriptionally by DNA methylation

[99��,100] and chromatin modification [101] and post-

transcriptionally [102]. As with viruses, some retrotran-

sposons are able to evade silencing [103��,104�]. More-

over, retrotransposon activity, which can be sharply

tissue-specific [90�,105��] can conversely lead to post-

transcriptional silencing of neighbouring genes

[10,55,106].

ConclusionsAnalysis of retrotransposon replication shows a unity that

transcends both the borders separating the Kingdoms of

eukaryotic evolution and the diverse forms of these

abundant and ancient entities. The replicational mech-

anisms of retroviruses, LTR retrotransposons, LINEs,

and their non-autonomous partners all rely on RT. RT

appears to originate in an ancient family, rvt, able to

incorporate both ribonucleotides and deoxyribonucleo-

tides [33��]. The great parting of the ways between

LINEs and the retroviruses and LTR retrotransposons

was the innovation of IN for integration of a pre-formed

cDNA. The LTRs of retrotransposons and retroviruses

share a universal 50 TG. . ... CA 30 structure within term-

inal inverted repeats, which provide the recognition and

binding sites for this enzyme. Indeed, a recent analysis

lends weight to the view that LTRs arose once, making

the retrotransposons and retroviruses monophyletic [22�].

Although retroviruses and superfamily Gypsy retrotran-

sposons share a common order of their gag and pol genes,

the env gene is generally thought to distinguish the

retroviruses. It encodes the envelope protein enabling

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8 Virus replication in animals and plants

COVIRO-298; NO. OF PAGES 11

retroviruses to leave the host cell and infect other cells.

However, env-bearing clades of Gypsy [107] and Copiaelements [108,109] are widespread in plants. The intra-

cellular or extracellular role of their env domains never-

theless remains to be established. Taken together, the

evidence suggests that the viral forms — the retro-

viruses — represent a gain of function within the Gypsyretrotransposons. Indeed, an analysis of gag and polsuggests that the Retroviridae is polyphyletic, having

arisen at least three times from within the Gypsy clade

[110]. In this context, the endogenous retroviruses

(ERVs), which have lost their infectivity, have merely

returned to their old life style [111].

The role of the envelope protein in retroviral infectivity

and its widespread occurrence in plants raises the question

of its role in ‘horizontal transfers’ of plant retrotransposons

between reproductively isolated species. Horizontal trans-

fer is often invoked when a group of retrotransposons is

found in an organism phylogenetically distant from the

clade in which it is expected. For example, TRIM

elements were heretofore found only in plants, but have

been recently discovered in red harvester ants [112].

Claims of horizontal transfer need to be considered in light

of the availability of sequence data bridging the phyloge-

netic divide. Nonetheless, well-supported cases of hori-

zontal transfer of retrotransposons in plants have been

reported [113]. Evidence for horizontal transfer of env-

bearing elements in plants is, however, still lacking, sharply

different from the situation in Drosophila [114].

For the plants, what remains unresolved, aside from many

mechanistic details, is how retrotransposon replication

takes place within the context of the plant life cycle so

that new copies are passed through the germline to the

next generation. The plant life cycle differs greatly from

that of animals or fungi. In plants, the germline is laid

down late in development, following many somatic cell

divisions and the eventual transition of an apical meris-

tem to a floral meristem. The reasons that LTR retro-

transposons in particular have been so successful in plants

remain to be established.

AcknowledgementsThomas Wicker, Francois Sabot, and Jan Buchmann are thanked for usefuldiscussions. Funding from Academy of Finland (Decision 134079) isgratefully acknowledged.

References and recommended readingPapers of particular interest, published within the period of review,have been highlighted as:

� of special interest

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The authors establish that the well-studied Tnt1 retrotransposon oftobacco remains active even when Tnt1 promoters driving transgenesbecome silenced by methylation.

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Jaaskelainen M, Chang W, Moisy C, Schulman AH:Retrotransposon BARE displays strong tissue-specificdifferences in expression. New Phytol 2013. in press.

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n Virol (2013), http://dx.doi.org/10.1016/j.coviro.2013.08.009

Current Opinion in Virology 2013, 3:1–11