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Type I Interferon Regulates the Expression of Long Non-Coding RNAs.

Carnero E, Barriocanal M, Segura V, Guruceaga E, Prior C, Börner K, Grimm D, Fortes P - Front Immunol (2014)

Bottom Line: Surprisingly, both ISR2 and 8 were significantly upregulated in cultured cells and livers from patients infected with HCV.This is relevant as genome-wide guilt-by-association studies predict that ISR2, 8, and 12 may function in viral processes, in the IFN pathway and the antiviral response.Therefore, we propose that these lncRNAs could be induced by IFN to function as positive or negative regulators of the antiviral response.

View Article: PubMed Central - PubMed

Affiliation: Department of Gene therapy and Hepatology, Center for Applied Medical Research (CIMA), University of Navarra , Pamplona , Spain.

ABSTRACT
Interferons (IFNs) are key players in the antiviral response. IFN sensing by the cell activates transcription of IFN-stimulated genes (ISGs) able to induce an antiviral state by affecting viral replication and release. IFN also induces the expression of ISGs that function as negative regulators to limit the strength and duration of IFN response. The ISGs identified so far belong to coding genes. However, only a small proportion of the transcriptome corresponds to coding transcripts and it has been estimated that there could be as many coding as long non-coding RNAs (lncRNAs). To address whether IFN can also regulate the expression of lncRNAs, we analyzed the transcriptome of HuH7 cells treated or not with IFNα2 by expression arrays. Analysis of the arrays showed increased levels of several well-characterized coding genes that respond to IFN both at early or late times. Furthermore, we identified several IFN-stimulated or -downregulated lncRNAs (ISRs and IDRs). Further validation showed that ISR2, 8, and 12 expression mimics that of their neighboring genes GBP1, IRF1, and IL6, respectively, all related to the IFN response. These genes are induced in response to different doses of IFNα2 in different cell lines at early (ISR2 or 8) or later (ISR12) time points. IFNβ also induced the expression of these lncRNAs. ISR2 and 8 were also induced by an influenza virus unable to block the IFN response but not by other wild-type lytic viruses tested. Surprisingly, both ISR2 and 8 were significantly upregulated in cultured cells and livers from patients infected with HCV. Increased levels of ISR2 were also detected in patients chronically infected with HIV. This is relevant as genome-wide guilt-by-association studies predict that ISR2, 8, and 12 may function in viral processes, in the IFN pathway and the antiviral response. Therefore, we propose that these lncRNAs could be induced by IFN to function as positive or negative regulators of the antiviral response.

No MeSH data available.


Related in: MedlinePlus

Validation of IDRs and ISRs at different times after IFN treatment. Expression levels of 24 ISRs (A) and 16 IDRs (B) were evaluated by qRT-PCR in HuH7 cells treated with 0 or 10,000 units/ml of IFNα for 6, 12, 24, 48, or 72 h. GAPDH was also evaluated by qRT-PCR and used as a reference to calculate the relative levels of each transcript. The ratio of levels with IFN versus no IFN is shown for each candidate at each time point. The results have been clustered and are shown in the form of a heat map. The color scale is shown at the top. Red denotes upregulation and green downregulation. A maximum of linear units from −5 to +5 (A) or −3 to +3 (B) has been set as indicated in the color scale.
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Figure 3: Validation of IDRs and ISRs at different times after IFN treatment. Expression levels of 24 ISRs (A) and 16 IDRs (B) were evaluated by qRT-PCR in HuH7 cells treated with 0 or 10,000 units/ml of IFNα for 6, 12, 24, 48, or 72 h. GAPDH was also evaluated by qRT-PCR and used as a reference to calculate the relative levels of each transcript. The ratio of levels with IFN versus no IFN is shown for each candidate at each time point. The results have been clustered and are shown in the form of a heat map. The color scale is shown at the top. Red denotes upregulation and green downregulation. A maximum of linear units from −5 to +5 (A) or −3 to +3 (B) has been set as indicated in the color scale.

Mentions: We next wanted to validate the IFN effect on these candidates in independent samples using a different technique. Furthermore, we wanted to determine whether the levels of ISRs and IDRs were altered early after IFN treatment. Therefore, the expression levels of 24 ISRs and 16 IDRs were evaluated by qRT-PCR in HuH7 cells treated with 0 or 10,000 units/ml of IFNα for 6, 12, 24, 48, or 72 h. The fold-change observed for each candidate at each time point is shown in Figure 3 and Table S5 in Supplementary Material. At 72 h post-treatment, 11 ISRs and 8 IDRs showed a fold-change higher or lower, respectively, than 1.5. Only ISR13 and 20 were not significantly upregulated, or IDR3, 5, 9, 12, and 13 were not significantly downregulated, at any time tested. However, the fold-change was relatively low in most of the cases, indicating a weak response to IFN. Moreover, 40% of the candidates showed low overall expression levels (Table S4 in Supplementary Material and data not shown). Interestingly, ISR2 and ISR8 were induced more than 50-fold at 6 h post-IFN treatment, and ISR1 was induced more than 20-fold at later times. Therefore, we decided to study these candidates further. We also opted to focus on ISR10 and 12, as they were upregulated at later time points. IDR1 and IDR2, which were downregulated at most times studied, were also analyzed further.


Type I Interferon Regulates the Expression of Long Non-Coding RNAs.

Carnero E, Barriocanal M, Segura V, Guruceaga E, Prior C, Börner K, Grimm D, Fortes P - Front Immunol (2014)

Validation of IDRs and ISRs at different times after IFN treatment. Expression levels of 24 ISRs (A) and 16 IDRs (B) were evaluated by qRT-PCR in HuH7 cells treated with 0 or 10,000 units/ml of IFNα for 6, 12, 24, 48, or 72 h. GAPDH was also evaluated by qRT-PCR and used as a reference to calculate the relative levels of each transcript. The ratio of levels with IFN versus no IFN is shown for each candidate at each time point. The results have been clustered and are shown in the form of a heat map. The color scale is shown at the top. Red denotes upregulation and green downregulation. A maximum of linear units from −5 to +5 (A) or −3 to +3 (B) has been set as indicated in the color scale.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4222131&req=5

Figure 3: Validation of IDRs and ISRs at different times after IFN treatment. Expression levels of 24 ISRs (A) and 16 IDRs (B) were evaluated by qRT-PCR in HuH7 cells treated with 0 or 10,000 units/ml of IFNα for 6, 12, 24, 48, or 72 h. GAPDH was also evaluated by qRT-PCR and used as a reference to calculate the relative levels of each transcript. The ratio of levels with IFN versus no IFN is shown for each candidate at each time point. The results have been clustered and are shown in the form of a heat map. The color scale is shown at the top. Red denotes upregulation and green downregulation. A maximum of linear units from −5 to +5 (A) or −3 to +3 (B) has been set as indicated in the color scale.
Mentions: We next wanted to validate the IFN effect on these candidates in independent samples using a different technique. Furthermore, we wanted to determine whether the levels of ISRs and IDRs were altered early after IFN treatment. Therefore, the expression levels of 24 ISRs and 16 IDRs were evaluated by qRT-PCR in HuH7 cells treated with 0 or 10,000 units/ml of IFNα for 6, 12, 24, 48, or 72 h. The fold-change observed for each candidate at each time point is shown in Figure 3 and Table S5 in Supplementary Material. At 72 h post-treatment, 11 ISRs and 8 IDRs showed a fold-change higher or lower, respectively, than 1.5. Only ISR13 and 20 were not significantly upregulated, or IDR3, 5, 9, 12, and 13 were not significantly downregulated, at any time tested. However, the fold-change was relatively low in most of the cases, indicating a weak response to IFN. Moreover, 40% of the candidates showed low overall expression levels (Table S4 in Supplementary Material and data not shown). Interestingly, ISR2 and ISR8 were induced more than 50-fold at 6 h post-IFN treatment, and ISR1 was induced more than 20-fold at later times. Therefore, we decided to study these candidates further. We also opted to focus on ISR10 and 12, as they were upregulated at later time points. IDR1 and IDR2, which were downregulated at most times studied, were also analyzed further.

Bottom Line: Surprisingly, both ISR2 and 8 were significantly upregulated in cultured cells and livers from patients infected with HCV.This is relevant as genome-wide guilt-by-association studies predict that ISR2, 8, and 12 may function in viral processes, in the IFN pathway and the antiviral response.Therefore, we propose that these lncRNAs could be induced by IFN to function as positive or negative regulators of the antiviral response.

View Article: PubMed Central - PubMed

Affiliation: Department of Gene therapy and Hepatology, Center for Applied Medical Research (CIMA), University of Navarra , Pamplona , Spain.

ABSTRACT
Interferons (IFNs) are key players in the antiviral response. IFN sensing by the cell activates transcription of IFN-stimulated genes (ISGs) able to induce an antiviral state by affecting viral replication and release. IFN also induces the expression of ISGs that function as negative regulators to limit the strength and duration of IFN response. The ISGs identified so far belong to coding genes. However, only a small proportion of the transcriptome corresponds to coding transcripts and it has been estimated that there could be as many coding as long non-coding RNAs (lncRNAs). To address whether IFN can also regulate the expression of lncRNAs, we analyzed the transcriptome of HuH7 cells treated or not with IFNα2 by expression arrays. Analysis of the arrays showed increased levels of several well-characterized coding genes that respond to IFN both at early or late times. Furthermore, we identified several IFN-stimulated or -downregulated lncRNAs (ISRs and IDRs). Further validation showed that ISR2, 8, and 12 expression mimics that of their neighboring genes GBP1, IRF1, and IL6, respectively, all related to the IFN response. These genes are induced in response to different doses of IFNα2 in different cell lines at early (ISR2 or 8) or later (ISR12) time points. IFNβ also induced the expression of these lncRNAs. ISR2 and 8 were also induced by an influenza virus unable to block the IFN response but not by other wild-type lytic viruses tested. Surprisingly, both ISR2 and 8 were significantly upregulated in cultured cells and livers from patients infected with HCV. Increased levels of ISR2 were also detected in patients chronically infected with HIV. This is relevant as genome-wide guilt-by-association studies predict that ISR2, 8, and 12 may function in viral processes, in the IFN pathway and the antiviral response. Therefore, we propose that these lncRNAs could be induced by IFN to function as positive or negative regulators of the antiviral response.

No MeSH data available.


Related in: MedlinePlus