Retrotransposition of marked SVA elements by human L1s in cultured cells

Dustin C. Hancks, John L. Goodier, Prabhat K. Mandal, Ling E. Cheung, Haig H. Kazazian

Research output: Contribution to journalArticle

107 Citations (Scopus)

Abstract

Human retrotransposons generate structural variation and genomic diversity through ongoing retrotransposition and non-allelic homologous recombination. Cell culture retrotransposition assays have provided great insight into the genomic impact of retrotransposons, in particular, LINE-1(L1) and Alu elements; however, no such assay exists for the youngest active human retrotransposon, SINE-VNTR-Alu (SVA). Here we report the development of an SVA cell culture retrotransposition assay. We marked several SVAs with either neomycin or EGFP retrotransposition indicator cassettes. Engineered SVAs retrotranspose using L1 proteins supplemented in trans in multiple cell lines, including U2OS osteosarcoma cells where SVA retrotransposition is equal to that of an engineered L1. Engineered SVAs retrotranspose at 1-54 times the frequency of a marked pseudogene in HeLa HA cells. Furthermore, our data suggest a variable requirement for L1 ORF1p for SVA retrotransposition. Recovered engineered SVA insertions display all the hallmarks of LINE-1 retrotransposition and some contain 5′ and 3′ transductions, which are common for genomic SVAs. Of particular interest is the fact that four out of five insertions recovered from one SVA are full-length, with the 5′ end of these insertions beginning within 5 nt of the CMV promoter transcriptional start site. This assay demonstrates that SVA elements are indeed mobilized in trans by L1. Previously intractable questions regarding SVA biology can now be addressed.

Original languageEnglish (US)
Article numberddr245
Pages (from-to)3386-3400
Number of pages15
JournalHuman Molecular Genetics
Volume20
Issue number17
DOIs
StatePublished - Sep 1 2011

Fingerprint

Short Interspersed Nucleotide Elements
Alu Elements
Cultured Cells
Retroelements
Genomic Structural Variation
Cell Culture Techniques
Long Interspersed Nucleotide Elements
Pseudogenes
Neomycin
Homologous Recombination
Osteosarcoma
HeLa Cells
Cell Line

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics
  • Genetics(clinical)

Cite this

Hancks, D. C., Goodier, J. L., Mandal, P. K., Cheung, L. E., & Kazazian, H. H. (2011). Retrotransposition of marked SVA elements by human L1s in cultured cells. Human Molecular Genetics, 20(17), 3386-3400. [ddr245]. https://doi.org/10.1093/hmg/ddr245

Retrotransposition of marked SVA elements by human L1s in cultured cells. / Hancks, Dustin C.; Goodier, John L.; Mandal, Prabhat K.; Cheung, Ling E.; Kazazian, Haig H.

In: Human Molecular Genetics, Vol. 20, No. 17, ddr245, 01.09.2011, p. 3386-3400.

Research output: Contribution to journalArticle

Hancks, DC, Goodier, JL, Mandal, PK, Cheung, LE & Kazazian, HH 2011, 'Retrotransposition of marked SVA elements by human L1s in cultured cells', Human Molecular Genetics, vol. 20, no. 17, ddr245, pp. 3386-3400. https://doi.org/10.1093/hmg/ddr245
Hancks, Dustin C. ; Goodier, John L. ; Mandal, Prabhat K. ; Cheung, Ling E. ; Kazazian, Haig H. / Retrotransposition of marked SVA elements by human L1s in cultured cells. In: Human Molecular Genetics. 2011 ; Vol. 20, No. 17. pp. 3386-3400.
@article{6e4ae9d99c204824875b1c9daa997fed,
title = "Retrotransposition of marked SVA elements by human L1s in cultured cells",
abstract = "Human retrotransposons generate structural variation and genomic diversity through ongoing retrotransposition and non-allelic homologous recombination. Cell culture retrotransposition assays have provided great insight into the genomic impact of retrotransposons, in particular, LINE-1(L1) and Alu elements; however, no such assay exists for the youngest active human retrotransposon, SINE-VNTR-Alu (SVA). Here we report the development of an SVA cell culture retrotransposition assay. We marked several SVAs with either neomycin or EGFP retrotransposition indicator cassettes. Engineered SVAs retrotranspose using L1 proteins supplemented in trans in multiple cell lines, including U2OS osteosarcoma cells where SVA retrotransposition is equal to that of an engineered L1. Engineered SVAs retrotranspose at 1-54 times the frequency of a marked pseudogene in HeLa HA cells. Furthermore, our data suggest a variable requirement for L1 ORF1p for SVA retrotransposition. Recovered engineered SVA insertions display all the hallmarks of LINE-1 retrotransposition and some contain 5′ and 3′ transductions, which are common for genomic SVAs. Of particular interest is the fact that four out of five insertions recovered from one SVA are full-length, with the 5′ end of these insertions beginning within 5 nt of the CMV promoter transcriptional start site. This assay demonstrates that SVA elements are indeed mobilized in trans by L1. Previously intractable questions regarding SVA biology can now be addressed.",
author = "Hancks, {Dustin C.} and Goodier, {John L.} and Mandal, {Prabhat K.} and Cheung, {Ling E.} and Kazazian, {Haig H.}",
year = "2011",
month = "9",
day = "1",
doi = "10.1093/hmg/ddr245",
language = "English (US)",
volume = "20",
pages = "3386--3400",
journal = "Human Molecular Genetics",
issn = "0964-6906",
publisher = "Oxford University Press",
number = "17",

}

TY - JOUR

T1 - Retrotransposition of marked SVA elements by human L1s in cultured cells

AU - Hancks, Dustin C.

AU - Goodier, John L.

AU - Mandal, Prabhat K.

AU - Cheung, Ling E.

AU - Kazazian, Haig H.

PY - 2011/9/1

Y1 - 2011/9/1

N2 - Human retrotransposons generate structural variation and genomic diversity through ongoing retrotransposition and non-allelic homologous recombination. Cell culture retrotransposition assays have provided great insight into the genomic impact of retrotransposons, in particular, LINE-1(L1) and Alu elements; however, no such assay exists for the youngest active human retrotransposon, SINE-VNTR-Alu (SVA). Here we report the development of an SVA cell culture retrotransposition assay. We marked several SVAs with either neomycin or EGFP retrotransposition indicator cassettes. Engineered SVAs retrotranspose using L1 proteins supplemented in trans in multiple cell lines, including U2OS osteosarcoma cells where SVA retrotransposition is equal to that of an engineered L1. Engineered SVAs retrotranspose at 1-54 times the frequency of a marked pseudogene in HeLa HA cells. Furthermore, our data suggest a variable requirement for L1 ORF1p for SVA retrotransposition. Recovered engineered SVA insertions display all the hallmarks of LINE-1 retrotransposition and some contain 5′ and 3′ transductions, which are common for genomic SVAs. Of particular interest is the fact that four out of five insertions recovered from one SVA are full-length, with the 5′ end of these insertions beginning within 5 nt of the CMV promoter transcriptional start site. This assay demonstrates that SVA elements are indeed mobilized in trans by L1. Previously intractable questions regarding SVA biology can now be addressed.

AB - Human retrotransposons generate structural variation and genomic diversity through ongoing retrotransposition and non-allelic homologous recombination. Cell culture retrotransposition assays have provided great insight into the genomic impact of retrotransposons, in particular, LINE-1(L1) and Alu elements; however, no such assay exists for the youngest active human retrotransposon, SINE-VNTR-Alu (SVA). Here we report the development of an SVA cell culture retrotransposition assay. We marked several SVAs with either neomycin or EGFP retrotransposition indicator cassettes. Engineered SVAs retrotranspose using L1 proteins supplemented in trans in multiple cell lines, including U2OS osteosarcoma cells where SVA retrotransposition is equal to that of an engineered L1. Engineered SVAs retrotranspose at 1-54 times the frequency of a marked pseudogene in HeLa HA cells. Furthermore, our data suggest a variable requirement for L1 ORF1p for SVA retrotransposition. Recovered engineered SVA insertions display all the hallmarks of LINE-1 retrotransposition and some contain 5′ and 3′ transductions, which are common for genomic SVAs. Of particular interest is the fact that four out of five insertions recovered from one SVA are full-length, with the 5′ end of these insertions beginning within 5 nt of the CMV promoter transcriptional start site. This assay demonstrates that SVA elements are indeed mobilized in trans by L1. Previously intractable questions regarding SVA biology can now be addressed.

UR - http://www.scopus.com/inward/record.url?scp=80051702066&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=80051702066&partnerID=8YFLogxK

U2 - 10.1093/hmg/ddr245

DO - 10.1093/hmg/ddr245

M3 - Article

VL - 20

SP - 3386

EP - 3400

JO - Human Molecular Genetics

JF - Human Molecular Genetics

SN - 0964-6906

IS - 17

M1 - ddr245

ER -