Nonviral Retroposon LINE Element 지도 교수 : 김 희 수 교수님 발 표 자 : 문 요 섭 참 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 proteins Lipid envelope

Virus groupPrototypesOther example Avian sarcoma and leukosis virusesRSVALV(RAV-1, 2) Reticuloendotheliosis virusesSNVREV CSV Mammalian leukemia and sarcoma virusesMo-MLVMO-MSV Ha-MSV, Fr-SFFV FeLV, SSAV Mammary tumor virusesMMTVHERV-K Primate type D virusesMPMVSRV-1, SMRV Human T-cell leukemia-related virusesHTLV-1HTLV-2, STLV BLV Immunodeficiency and lentivirusesHIV-1 Visna HIV-2 SIV CAEV EIAV Major Groups of Retroviruses

U3R U5 (A)n U3R 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 LTR Genomic Structure of Retrovirus 3`5`

Generation of LTR PBS Coding regionU3RRU5 PBS Coding regionU3RRU5 tRNA U5R tRNA PBS Coding regionU3R tRNA U5R PBS Coding regionU3R tRNA U5R PBS Coding regionU3R U5RU3 tRNA Genimic RNA DNA to form DNA copy tRNA extended removed Hybrid RNA First jump from 3` end DNA strand extended (A)n 5` 3` (A)n

U5RU3PBS tRNA U5RU3 PBS U5RU3 tRNA U5RU3 U5RU3 PBS U5RU3 U5RU3 PBS U5RU3 U5RU3 PBS Must hybrid RNA removed synthesized 3` end of second DNA strand and tRNA removed Remaining hybrid RNA Second jump 3` ends Both strands completed by synthesis from LTR 5` 3` Retroviral DNA

Release Budding Translation mRNA Transcription Transport to nucleus Reverse transcription Double strand of DNA Cytoplasm Absorption to Specific Receptor Assembly Provirus DNA Nuclear membrane Endocytosis Replication Mechanism of Retrovirus

TypeMechanism of MovementExamples 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)

ORF1ORF2P Poly(A) D D E Poly(A) TTTT Mobile Elements in Mammalian Genome MechanismClassExampleStructure Transposon LTR-containing Autonomous Retrotransposons Non-LTR retrotransposons Non-autonomous mariner Alu L1 Processed pseudogene Transposase EN RT terminator linker A-rich ‘cut’ or ‘copy and paste’ Retroviral-like Target primed reverse transcription(TPRT) TPRT? R-Alu L-Alu LTRORF1ORF2LTR Truncated HERVs

Mechanism of L1 Retrotransposon ORF1ORF2P Poly(A) EN RT AAAAn 5` AAAAn5` p40 ORF2 AAAAA 3`OH-TTTTT 5` ORF2 Poly(A) Cytoplasm Nucleus Transcription processing mRNA export Import or entry during mitosis transcription(TPRT) Target primed reverse at new chromosomal site Integration of truncated L1 modification RNP assembly; Transcription and Post-translational

Retroelement Retroposon SINE Retrotransposon Retrovirus LINE LTR P ORF1ORF2 gagpolenv RNA intermediate - LTR element+ LTR element - env+ env - RT+ RT P ORF1ORF2LTR 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 & Retrotransposon

Non-LTR Retro-element Retroelement Retroposon SINE Retrotransposon Retrovirus LINE RNA intermediate - LTR element+ LTR element - env+ env - RT+ RT LTRORF1ORF2LTR gagpolenvLTR P Poly(A) Human Alu ORF1ORF2P Poly(A) L1 Yeast Ty1/copia/truncated HERVs Human THE1

Evolution & Phylogeny of Non-LTR Retrotransposable Element Evolution & Phylogeny of Non-LTR Retrotransposable Element RT APE RT APERT APERT RNHAPERT APERT RNHAPERT RNHAPERT RNHAPERT RNHAPERT APERT APERT APE RNH CLADE CR1 CRE R2 R4 L1 RTE Tad R1 LOA I Jockey 250 aa

ORF1 (~1 kb) ~ ORF2 (~4 kb) ~ Target-site direct repeat Multiple stop codons (~1 kb) ~ Coding region A/T-rich region 5` 3` 5` Structure of LINE Element Structure of LINE Element

AAAAAAAAA TTTTTTTTTT TTTTTT AAAAAA AAAAAAAAAUUUUUU AAAAAAAAA TTTUUUUUU 3` 5` 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 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 Movement

3` 5` (A)n (T)n (A)n (T)n 5` 3` 3’` 3` L1 RNA ORF-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 Inverted segment 5` Truncated L1 5` Inverted/deleted L1 5` Inverted L1 Various Model for L1 Element

Transduction ORF2ORF1 An ENRT C An Genomic Diversity by L1 LINE Element An

Promoter Arrays & L1 Transposon Parental L1Md element 208 bp repeats 4 2/3 copies ORF-1 ORF-2(A)n SDR 3 2/3 2 2/3 1 2/3 2/3 mRNAs Functional progeny L1Md element + Non-functional progeny L1Md element

L1Hs L1Md 5`UTR 5’UTR ORF-1 ORF-2 3`UTR 3’UTR (A)n Comparative Analysis of L1Md & L1Hs

LINE SINE Transcription tRNA-related regiontRNA-unrelated region LINE-derived region Transcript 3` 5` 3` 5` 5’ 5`3` Translation Transcript 5`3` 5` 3` Reverse transcriptase Recognition of the 3’-end sequence Synthesis of cDNA New integration site Common 3` Sequence of LINE & SINE

Function of LINE Elements 1. Various Diseases 2. Evolutionary Change 3. Diversity & Phylogeny