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Correlation of LNCR rasiRNAs expression with heterochromatin formation during development of the holocentric insect Spodoptera frugiperda.

ABSTRACT: Repeat-associated small interfering RNAs (rasiRNAs) are derived from various genomic repetitive elements and ensure genomic stability by silencing endogenous transposable elements. Here we describe a novel subset of 46 rasiRNAs named LNCR rasiRNAs due to their homology with one long non-coding RNA (LNCR) of Spodoptera frugiperda. LNCR operates as the intermediate of an unclassified transposable element (TE-LNCR). TE-LNCR is a very invasive transposable element, present in high copy numbers in the S. frugiperda genome. LNCR rasiRNAs are single-stranded RNAs without a prominent nucleotide motif, which are organized in two distinct, strand-specific clusters. The expression of LNCR and LNCR rasiRNAs is developmentally regulated. Formation of heterochromatin in the genomic region where three copies of the TE-LNCR are embedded was followed by chromatin immunoprecipitation (ChIP) and we observed this chromatin undergo dynamic changes during development. In summary, increased LNCR expression in certain developmental stages is followed by the appearance of a variety of LNCR rasiRNAs which appears to correlate with subsequent accumulation of a heterochromatic histone mark and silencing of the genomic region with TE-LNCR. These results support the notion that a repeat-associated small interfering RNA pathway is linked to heterochromatin formation and/or maintenance during development to establish repression of the TE-LNCR transposable element. This study provides insights into the rasiRNA silencing pathway and its role in the formation of fluctuating heterochromatin during the development of one holocentric organism.

SUBMITTER: Stanojcic S

PROVIDER: S-EPMC3184123 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

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Correlation of LNCR rasiRNAs expression with heterochromatin formation during development of the holocentric insect Spodoptera frugiperda.

Stanojcic Slavica S Gimenez Sylvie S Permal Emmanuelle E Cousserans François F Quesneville Hadi H Fournier Philippe P d'Alençon Emmanuelle E

PloS one 20110930 9

Repeat-associated small interfering RNAs (rasiRNAs) are derived from various genomic repetitive elements and ensure genomic stability by silencing endogenous transposable elements. Here we describe a novel subset of 46 rasiRNAs named LNCR rasiRNAs due to their homology with one long non-coding RNA (LNCR) of Spodoptera frugiperda. LNCR operates as the intermediate of an unclassified transposable element (TE-LNCR). TE-LNCR is a very invasive transposable element, present in high copy numbers in th ...[more]

PMID: 21980354

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Project description:Chikungunya virus-like particles (VLPs) have potential to be used as a prophylactic vaccine based on testing in multiple animal models and are currently being evaluated for human use in a Phase I clinical trial. The current method for producing these enveloped alphavirus VLPs by transient gene expression in mammalian cells presents challenges for scalable and robust industrial manufacturing, so the insect cell baculovirus expression vector system was evaluated as an alternative expression technology. Subsequent to recombinant baculovirus infection of Sf21 cells in standard culture media (pH 6.2-6.4), properly processed Chikungunya structural proteins were detected and assembled capsids were observed. However, an increase in culture pH to 6.6-6.8 was necessary to produce detectable concentrations of assembled VLPs. Since this elevated production pH exceeds the optimum for growth medium stability and Sf21 culture, medium modifications were made and a novel insect cell variant (SfBasic) was derived by exposure of Sf21 to elevated culture pH for a prolonged period of time. The high-pH adapted SfBasic insect cell line described herein is capable of maintaining normal cell growth into the typical mammalian cell culture pH range of 7.0-7.2 and produces 11-fold higher Chikungunya VLP yields relative to the parental Sf21 cell line. After scale-up into stirred tank bioreactors, SfBasic derived VLPs were chromatographically purified and shown to be similar in size and structure to a VLP standard derived from transient gene expression in HEK293 cells. Total serum anti-Chikungunya IgG and neutralizing titers from guinea pigs vaccinated with SfBasic derived VLPs or HEK293 derived VLPs were not significantly different with respect to production method, suggesting that this adapted insect cell line and production process could be useful for manufacturing Chikungunya VLPs for use as a vaccine. The adaptation of Sf21 to produce high levels of recombinant protein and VLPs in an elevated pH range may also have applications for other pH-sensitive protein or VLP targets.

Dataset Information

Correlation of LNCR rasiRNAs expression with heterochromatin formation during development of the holocentric insect Spodoptera frugiperda.

Publications

Correlation of LNCR rasiRNAs expression with heterochromatin formation during development of the holocentric insect Spodoptera frugiperda.

Similar Datasets

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