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Molecular characterization of a retrotransposon in the Rhynchosciara americana genome and its association with telomere

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Abstract

Non-LTR retrotransposons, also known as long interspersed nuclear elements (LINEs), are transposable elements that encode a reverse transcriptase and insert into genomic locations via RNA intermediates. The sequence analysis of a cDNA library constructed from mRNA of the salivary glands of R. americana showed the presence of putative class I elements. The cDNA clone with homology to a reverse transcriptase was the starting point for the present study. Genomic phage was isolated and sequenced and the molecular structure of the element was characterized as being a non-LTR retrotransposable element. Southern blot analysis indicated that this transposable element is represented by repeat sequences in the genome of R. americana. Chromosome tips were consistently positive when this element was used as probe in in-situ hybridization. Real-time RT-PCR showed that this retrotransposon is transcribed at different periods of larval development. Most interesting, the silencing of this retrotransposon in R. americana by RNA interference resulted in reduced transcript levels and in accelerated larval development.

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Authors and Affiliations

  1. Departamento de Biologia Celular e do Desenvolvimento, Avenida Professor Lineu Prestes, 1524 Y ICBI Y sala 307, Universidade de São Paulo, São Paulo, CEP 05508-900, Brazil

    Paula Rezende-Teixeira, Fábio Siviero, Amanda S. Brandão & Gláucia M. Machado-Santelli

  2. Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, SP, Brazil

    Paula Rezende-Teixeira, Fábio Siviero & Roberto Vicente Santelli

  3. Programa de Pós-Graduação Interunidades em Biotecnologia, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo, SP, Brazil

    Fábio Siviero, Amanda S. Brandão & Gláucia M. Machado-Santelli

Authors
  1. Paula Rezende-Teixeira
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  2. Fábio Siviero
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  3. Amanda S. Brandão
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  4. Roberto Vicente Santelli
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  5. Gláucia M. Machado-Santelli
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Corresponding author

Correspondence to Paula Rezende-Teixeira.

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Figure S1.

Northern blot hybridized with putative reverse transcriptase cDNA probe from Rhynchosciara americana. Lanes: (1) fourth instar, second period; (2) fourth instar, third period; (3) fourth instar, fourth period; and (4) fourth instar, fifth period. The arrows indicate the transcript sizes in about of 9.0°kb and 6.5°kb. (TIFF 78.8KB)

Figure S2.

(A) Relative expression of the GAPDH gene in salivary glands of Rhynchosciara americana during normal larval development. Real-time RT-PCR analysis of mRNA extracted from different periods of the fourth instar of larval development. (B) Real-time RT-PCR analysis of mRNA extracted from dsRaRT-treated larvae. (TIFF 561KB)

Figure S3.

Appearance of larvae after RNAi treatment. (a) and (b) Sibling larvae at 18 and 20 days, respectively, after injection of dsRNA of the RaTART retrotransposon. (c) and (d) Control sibling larvae at 18 and 20 days of experiment, respectively, both with characteristics of second-period fourth instar larvae and normal morphological aspects. (TIFF 6.57MB)

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Rezende-Teixeira, P., Siviero, F., Brandão, A.S. et al. Molecular characterization of a retrotransposon in the Rhynchosciara americana genome and its association with telomere. Chromosome Res 16, 729–742 (2008). https://doi.org/10.1007/s10577-008-1223-9

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  • DOI: https://doi.org/10.1007/s10577-008-1223-9

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