Nonviral Retroposon LINE Element

지도 교수 : 김 희 수 교수님발 표 자 : 문 요 섭참 가 자 : 김 영 균 , 윤 성 용

제 4 회 부산대학교 생물학과 학술제

Retrovirus (HIV)Retrovirus (HIV)

RNase

Viral RNA

Capsid : core proteins

Receptor

binding proteinsLipid envelope

Virus group Prototypes Other example

Avian sarcoma and leukosis viruses RSV ALV(RAV-1, 2)

Reticuloendotheliosis viruses SNV REVCSV

Mammalian leukemia and sarcoma viruses Mo-MLV MO-MSVHa-MSV, Fr-SFFVFeLV, SSAV

Mammary tumor viruses MMTV HERV-K

Primate type D viruses MPMV SRV-1, SMRV

Human T-cell leukemia-related viruses HTLV-1 HTLV-2, STLVBLV

Immunodeficiency and lentiviruses HIV-1Visna

HIV-2SIVCAEVEIAV

Major Groups of RetrovirusesMajor Groups of Retroviruses

U3 R U5

(A)n

U3 R U5 Host-cell DNA

Poly(A) polymerase

Start site Poly(A) site

RNA polymerase II

RNA-processing enzymes

Retroviral RNA genome

Integrated retroviral DNA

Primary transcript

Coding region LTRLTR

Genomic Structure of RetrovirusGenomic Structure of Retrovirus

3`5`

Generation of LTRGeneration of LTR

PBS

Coding region U3 RR U5 PBS

Coding region U3 RR U5

tRNAU5R

tRNA

PBS Coding region U3 R

tRNAU5R

PBS Coding region U3 R

tRNAU5R

PBS Coding region U3 R

U5RU3 tRNA

Genimic RNA

DNA

to form DNA copytRNA extended

removedHybrid RNA

First jump

from 3` end

DNA strand extended

(A)n5`

5`

5` 3`

3`

3`

3`

3`

3`

(A)n

(A)n

(A)n

(A)n

U5RU3PBS tRNA

U5RU3 PBS

U5RU3 tRNA

U5RU3

U5RU3 PBS

U5RU3

U5RU3 PBS

U5RU3

U5RU3 PBS

Must hybrid RNA removed

synthesized3` end of second DNA strand

and tRNA removedRemaining hybrid RNA

Second jump

3` ends

Both strandscompleted by

synthesis from

LTR LTR

5`

5`

5`

5`

5`

5`

5`

5`

3`

3`

3`

3`

3`

3`

3`

3`

3`

3`

Retroviral DNA

Release

Budding

Translation

mRNATranscription

Transport to nucleus

Reverse transcription

Double strand of DNA

Cytoplasm

Absorption to Specific Receptor

Assembly

Provirus DNA

Nuclear membrane

Endocytosis

Replication Mechanism of RetrovirusReplication Mechanism of Retrovirus

Type Mechanism of Movement Examples

DNA-Mediated Transposition

Bacterial insertion sequences (IS element)

Excision or copying of DNA and its insertion at target site

IS1, IS10

Bacterial transposons copying of DNA and its insertion at target site

Tn9

Eukayotic transposons Excision of DNA and its insertion at target site

P element (Drosophila)Ac and Ds elements (corn)

RNA-Mediated Transposition

Viral retrotransposons

Transcription into RNA from promoter in left LTR by RNA polymerase II followed by reverse transcription and insertion at target site

Ty element (yeast)Copia elements (Drosophila)

Nonviral retrotransposons

Transcription into RNA from internal promoter; folding of transcript to provide primer for reverse transcription followed by insertion at target site

F and G elements (Drosophila)LINE and SINE element (mammals)Alu sequences (humans)

ORF1 ORF2PPoly(A)

D D E

Poly(A)TTTT

Mobile Elements in Mammalian GenomeMobile Elements in Mammalian Genome

MechanismClass Example Structure

Transposon

LTR-containing

AutonomousRetrotransposons

Non-LTR

retrotransposonsNon-autonomous

mariner

Alu

L1

Processedpseudogene

Transposase

EN RT

terminatorlinkerA-rich

‘cut’ or ‘copy

and paste’

Retroviral-like

Target primed reverse

transcription(TPRT)

TPRT?

TPRT?

R-AluL-Alu

LTR ORF1 ORF2 LTRTruncatedHERVs

Mechanism of L1 RetrotransposonMechanism of L1 Retrotransposon

ORF1 ORF2PPoly(A)

EN RT

AAAAn

AAAAn

5`

5`

AAAAn5`p40

p40

ORF2 ORF2

AAAAA

3`OH-T

TTTT

5` ORF2

ORF2

Poly(A)

Cytoplasm Nucleus

Transcription processing

mRNA exportImport or entry

during mitosis

transcription(TPRT)Target primed reverse

at new chromosomal siteIntegration of truncated L1

modification

RNP assembly;Transcription and

Post-translational

Retroelement

Retroposon

SINE

Retrotransposon

Retrovirus

LINE

LTR

P

ORF1 ORF2

gag pol env

RNA intermediate

- LTR element + LTR element

- env + env

- RT + RT

P

ORF1 ORF2 LTR

LTR LTR

LTR LTR

Poly(A)

Poly(A)

Human Alu

L1

Yeast Ty1/copia/truncated HERVs

Human THE1

Full-length HERVs/exogenous retrovirus

Reverse transcriptase

Insertion sequence or transposon Retrotransposon

Donor DNA

Flanking DNA RNA polymerase

RNA intermediate

Donor DNA

Transposed mobile element

DNA intermediates

Target DNA

Transposon & RetrotransposonTransposon & Retrotransposon

Non-LTR Retro-elementNon-LTR Retro-element

Retroelement

Retroposon

SINE

Retrotransposon

Retrovirus

LINE

RNA intermediate

- LTR element + LTR element

- env + env

- RT + RTLTR ORF1 ORF2 LTR

LTR LTR

gag pol envLTR LTR

PPoly(A)

Human Alu

ORF1 ORF2PPoly(A)

L1

Yeast Ty1/copia/truncated HERVs

Human THE1

Evolution & Phylogeny of Non-LTR Retrotransposable ElementEvolution & Phylogeny of Non-LTR Retrotransposable Element

RT

APE

RT

RT

RT

RTAPE RT

APE RT

RNHAPE RT

APE RT

RNHAPE RT

RNHAPE RT

RNHAPE RT

RNHAPE RT

APE RT

APE RT

APE

RNH

CLADE

CR1

CRE

R2

R4

L1

RTE

Tad

R1

LOA

I

Jockey

250 aa

100

100

100

100

100

100

98

90

99

82

76

56

98

99

82

ORF1(~1 kb)~

ORF2 (~4 kb)~

Target-site direct repeat

Multiple

stop codons(~1 kb)~

Coding region A/T-rich region

5`

3`

3`

5`

Structure of LINE ElementStructure of LINE Element

AAAAAAAAATTTTTTTTTT

TTTTTTAAAAAA

AAAAAAAAA UUUUUU

AAAAAAAAA

TTTUUUUUU

3`

5`

5`

5`

5`

3`

3`

3`

cDNA

L1 element Flanking DNA

1) An RNA polymerase transcribes bottom strand

2) RNA fold back on itself with Us and As hybridizing

3) 3` end primes synthesis of DNA from RNA

template by L1 reverse transcriptase

4) cDNA serves as template for synthesis of double-stranded L1 DNA

5) Unknown mechanism insert L1 element into cellular DNA

Synthesis of L1 LINE ElementSynthesis of L1 LINE Element

ORF2ORF1

SDR

New SDR

???

+

L1 mRNA

DNA/RNA hybrid

Transcription

A(n) 3` 5`

Inserted

functional L1

ORF-2 product

Target site

Transposed

5` truncated L1

Retroposon Model of L1 MovementRetroposon Model of L1 Movement

3` 5` (A)n

(A)n

(T)n

(A)n

(A)n

(A)n (A)n

(A)n

(A)n(A)n

(A)n

(A)n

(T)n

(T)n

(T)n

(T)n

(T)n

(T)n

(T)n

(T)n

(T)n

(T)n

5`

5`

5`

5`

5` 5`

5`

5`

5`

5`

5`

5` 5`

5`

5`

5`5`

5`

5`5`

5`

3`

3`

3`

3`

3`

3`

3` 3`

3`

3`

3`

3`

3`

3`

3`

3`

3`

3’`

3`

3`

L1 RNAORF-2 RTase

3` cDNA

RNAase H

Random priming

S.S. nuclease (nick)

S.S. nuclease (complete)

Repair synthesis

Integration

S.S. nuclease(partial)

Repair synthesis

Integration

Integration

Inverted segment

Inverted segment

5` Truncated L1

5` Inverted/deleted L1

5` Inverted L1

Various Model for L1 ElementVarious Model for L1 Element

Transduction

ORF2ORF1

AnEN RT C

An

An

An

An

Genomic Diversity by L1 LINE ElementGenomic Diversity by L1 LINE Element

An

An

Promoter Arrays & L1 TransposonPromoter Arrays & L1 TransposonParental L1Md element

208 bp repeats4 2/3 copies ORF-1 ORF-2 (A)n

SDR

SDR

3 2/3

2 2/3

1 2/3

2/3

mRNAs

Functional progeny L1Md element

+

Non-functional progeny L1Md element

L1Hs

L1Md5`UTR

5’UTR

ORF-1

ORF-1 ORF-2

ORF-23`UTR

3’UTR

(A)n

(A)n

Comparative Analysis of L1Md & L1HsComparative Analysis of L1Md & L1Hs

LINE

SINE

Transcription Transcription

tRNA-related region tRNA-unrelated region

LINE-derived region

Transcript

3`

5`

5`

5`

5`

3`

3`

3`

3`

3`

3`

3`5`

5`

5`

5`

5’ 5`3` 3`

Translation

Transcript5` 3` 5` 3`

Reverse transcriptase

Recognition of the 3’-end sequence

Synthesis of cDNA Synthesis of cDNA

New integration site New integration site

Common 3` Sequence of LINE & SINECommon 3` Sequence of LINE & SINE

Function of LINE ElementsFunction of LINE Elements

1. Various Diseases

2. Evolutionary Change

3. Diversity & Phylogeny