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Gene activity in primary T cells infected with HIV89.6: intron retention and induction of genomic repeats.

Sherrill-Mix S, Ocwieja KE, Bushman FD - Retrovirology (2015)

Bottom Line: Analysis showed that activation was associated with a particular variant of ERV-9 long terminal repeats that contains an indel near the U3-R border.These data also allowed quantification of >70 splice forms of the HIV89.6 RNA and specified the main types of chimeric HIV89.6-host RNAs.Comparison to over 100,000 integration site sequences from the same infected cell populations allowed quantification of authentic versus artifactual chimeric reads, showing that 5' read-in, splicing out of HIV89.6 from the D4 donor and 3' read-through were the most common HIV89.6-host cell chimeric RNA forms.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Perelman School of Medicine at the University of Pennsylvania, 425 Johnson Pavilion, 3610 Hamilton Walk, Philadelphia, PA, 19104, USA. shescott@mail.med.upenn.edu.

ABSTRACT

Background: HIV infection has been reported to alter cellular gene activity, but published studies have commonly assayed transformed cell lines and lab-adapted HIV strains, yielding inconsistent results. Here we carried out a deep RNA-Seq analysis of primary human T cells infected with the low passage HIV isolate HIV89.6.

Results: Seventeen percent of cellular genes showed altered activity 48 h after infection. In a meta-analysis including four other studies, our data differed from studies of HIV infection in cell lines but showed more parallels with infections of primary cells. We found a global trend toward retention of introns after infection, suggestive of a novel cellular response to infection. HIV89.6 infection was also associated with activation of several human endogenous retroviruses (HERVs) and retrotransposons, of interest as possible novel antigens that could serve as vaccine targets. The most highly activated group of HERVs was a subset of the ERV-9. Analysis showed that activation was associated with a particular variant of ERV-9 long terminal repeats that contains an indel near the U3-R border. These data also allowed quantification of >70 splice forms of the HIV89.6 RNA and specified the main types of chimeric HIV89.6-host RNAs. Comparison to over 100,000 integration site sequences from the same infected cell populations allowed quantification of authentic versus artifactual chimeric reads, showing that 5' read-in, splicing out of HIV89.6 from the D4 donor and 3' read-through were the most common HIV89.6-host cell chimeric RNA forms.

Conclusions: Analysis of RNA abundance after infection of primary T cells with the low passage HIV89.6 isolate disclosed multiple novel features of HIV-host interactions, notably intron retention and induction of transcription of retrotransposons and endogenous retroviruses.

No MeSH data available.


Related in: MedlinePlus

Changes in the abundance of intronic regions with HIV infection. Expression of intronic and exonic regions was quantified as the proportion of reads mapping within the intron/exon out of the total reads mapping to the transcription units overlapping that intron/exon. a Comparison of the ratios of expression between infected and uninfected replicates in exclusively intronic or exonic regions of transcription units. b Reproducibility of intron retention between replicates. Each point quantifies the change in expression with HIV infection for a specific intronic region. The x-axis shows changes in gene activity accompanying infection for one set of replicates (Infected-1 and Infected-2 vs. Uninfected-1) and the y-axis shows the same data for different replicates (Infected-3 vs. Uninfected-2). c Reproducibility of intron retention between studies. The plot is arranged as in b but with all data from our study combined on the x-axis and corresponding data from Chang et al. [25] on the y-axis
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Fig3: Changes in the abundance of intronic regions with HIV infection. Expression of intronic and exonic regions was quantified as the proportion of reads mapping within the intron/exon out of the total reads mapping to the transcription units overlapping that intron/exon. a Comparison of the ratios of expression between infected and uninfected replicates in exclusively intronic or exonic regions of transcription units. b Reproducibility of intron retention between replicates. Each point quantifies the change in expression with HIV infection for a specific intronic region. The x-axis shows changes in gene activity accompanying infection for one set of replicates (Infected-1 and Infected-2 vs. Uninfected-1) and the y-axis shows the same data for different replicates (Infected-3 vs. Uninfected-2). c Reproducibility of intron retention between studies. The plot is arranged as in b but with all data from our study combined on the x-axis and corresponding data from Chang et al. [25] on the y-axis

Mentions: Cells respond to infection by shutting down macromolecular synthesis at multiple levels [70–74], so we investigated whether cells also showed perturbations in splicing efficiency after infection. As a probe, we created a database of cellular genomic regions annotated exclusively as exons or introns in all splice forms in the UCSC gene database [75] and quantified expression in these regions in infected and uninfected cells. We found a significant increase in intronic sequences relative to exonic sequence (Wilcoxon test ) (Fig. 3a). This increase in intronic sequence was reproducible between replicates in our study (Kendall’s , ) (Fig. 3b). We reanalyzed RNA-Seq data from Chang et al. [25] and also documented intron retention that correlated with the changes seen in our data (Kendall’s , ) (Fig. 3c).Fig. 3


Gene activity in primary T cells infected with HIV89.6: intron retention and induction of genomic repeats.

Sherrill-Mix S, Ocwieja KE, Bushman FD - Retrovirology (2015)

Changes in the abundance of intronic regions with HIV infection. Expression of intronic and exonic regions was quantified as the proportion of reads mapping within the intron/exon out of the total reads mapping to the transcription units overlapping that intron/exon. a Comparison of the ratios of expression between infected and uninfected replicates in exclusively intronic or exonic regions of transcription units. b Reproducibility of intron retention between replicates. Each point quantifies the change in expression with HIV infection for a specific intronic region. The x-axis shows changes in gene activity accompanying infection for one set of replicates (Infected-1 and Infected-2 vs. Uninfected-1) and the y-axis shows the same data for different replicates (Infected-3 vs. Uninfected-2). c Reproducibility of intron retention between studies. The plot is arranged as in b but with all data from our study combined on the x-axis and corresponding data from Chang et al. [25] on the y-axis
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4574318&req=5

Fig3: Changes in the abundance of intronic regions with HIV infection. Expression of intronic and exonic regions was quantified as the proportion of reads mapping within the intron/exon out of the total reads mapping to the transcription units overlapping that intron/exon. a Comparison of the ratios of expression between infected and uninfected replicates in exclusively intronic or exonic regions of transcription units. b Reproducibility of intron retention between replicates. Each point quantifies the change in expression with HIV infection for a specific intronic region. The x-axis shows changes in gene activity accompanying infection for one set of replicates (Infected-1 and Infected-2 vs. Uninfected-1) and the y-axis shows the same data for different replicates (Infected-3 vs. Uninfected-2). c Reproducibility of intron retention between studies. The plot is arranged as in b but with all data from our study combined on the x-axis and corresponding data from Chang et al. [25] on the y-axis
Mentions: Cells respond to infection by shutting down macromolecular synthesis at multiple levels [70–74], so we investigated whether cells also showed perturbations in splicing efficiency after infection. As a probe, we created a database of cellular genomic regions annotated exclusively as exons or introns in all splice forms in the UCSC gene database [75] and quantified expression in these regions in infected and uninfected cells. We found a significant increase in intronic sequences relative to exonic sequence (Wilcoxon test ) (Fig. 3a). This increase in intronic sequence was reproducible between replicates in our study (Kendall’s , ) (Fig. 3b). We reanalyzed RNA-Seq data from Chang et al. [25] and also documented intron retention that correlated with the changes seen in our data (Kendall’s , ) (Fig. 3c).Fig. 3

Bottom Line: Analysis showed that activation was associated with a particular variant of ERV-9 long terminal repeats that contains an indel near the U3-R border.These data also allowed quantification of >70 splice forms of the HIV89.6 RNA and specified the main types of chimeric HIV89.6-host RNAs.Comparison to over 100,000 integration site sequences from the same infected cell populations allowed quantification of authentic versus artifactual chimeric reads, showing that 5' read-in, splicing out of HIV89.6 from the D4 donor and 3' read-through were the most common HIV89.6-host cell chimeric RNA forms.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Perelman School of Medicine at the University of Pennsylvania, 425 Johnson Pavilion, 3610 Hamilton Walk, Philadelphia, PA, 19104, USA. shescott@mail.med.upenn.edu.

ABSTRACT

Background: HIV infection has been reported to alter cellular gene activity, but published studies have commonly assayed transformed cell lines and lab-adapted HIV strains, yielding inconsistent results. Here we carried out a deep RNA-Seq analysis of primary human T cells infected with the low passage HIV isolate HIV89.6.

Results: Seventeen percent of cellular genes showed altered activity 48 h after infection. In a meta-analysis including four other studies, our data differed from studies of HIV infection in cell lines but showed more parallels with infections of primary cells. We found a global trend toward retention of introns after infection, suggestive of a novel cellular response to infection. HIV89.6 infection was also associated with activation of several human endogenous retroviruses (HERVs) and retrotransposons, of interest as possible novel antigens that could serve as vaccine targets. The most highly activated group of HERVs was a subset of the ERV-9. Analysis showed that activation was associated with a particular variant of ERV-9 long terminal repeats that contains an indel near the U3-R border. These data also allowed quantification of >70 splice forms of the HIV89.6 RNA and specified the main types of chimeric HIV89.6-host RNAs. Comparison to over 100,000 integration site sequences from the same infected cell populations allowed quantification of authentic versus artifactual chimeric reads, showing that 5' read-in, splicing out of HIV89.6 from the D4 donor and 3' read-through were the most common HIV89.6-host cell chimeric RNA forms.

Conclusions: Analysis of RNA abundance after infection of primary T cells with the low passage HIV89.6 isolate disclosed multiple novel features of HIV-host interactions, notably intron retention and induction of transcription of retrotransposons and endogenous retroviruses.

No MeSH data available.


Related in: MedlinePlus