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RIG-I is required for the inhibition of measles virus by retinoids.

Soye KJ, Trottier C, Richardson CD, Ward BJ, Miller WH - PLoS ONE (2011)

Bottom Line: Vitamin A can significantly decrease measles-associated morbidity and mortality.Retinoid signaling was also found to act in combination with IFN to induce high levels of RIG-I expression.IRF-1 is known to be regulated by retinoids and MeV, but we found recruitment of IRF-1 to the RIG-I promoter by retinoids alone.

View Article: PubMed Central - PubMed

Affiliation: McGill University Health Center Research Institute, Department of Infectious Diseases, McGill University, Montreal, Quebec, Canada.

ABSTRACT
Vitamin A can significantly decrease measles-associated morbidity and mortality. Vitamin A can inhibit the replication of measles virus (MeV) in vitro through an RARα- and type I interferon (IFN)-dependent mechanism. Retinoid-induced gene I (RIG-I) expression is induced by retinoids, activated by MeV RNA and is important for IFN signaling. We hypothesized that RIG-I is central to retinoid-mediated inhibition of MeV in vitro. We demonstrate that RIG-I expression is increased in cells treated with retinoids and infected with MeV. The central role of RIG-I in the retinoid-anti-MeV effect was demonstrated in the Huh-7/7.5 model; the latter cells having non-functional RIG-I. RAR-dependent retinoid signaling was required for the induction of RIG-I by retinoids and MeV. Retinoid signaling was also found to act in combination with IFN to induce high levels of RIG-I expression. RIG-I promoter activation required both retinoids and MeV, as indicated by markers of active chromatin. IRF-1 is known to be regulated by retinoids and MeV, but we found recruitment of IRF-1 to the RIG-I promoter by retinoids alone. Using luciferase expression constructs, we further demonstrated that the IRF-1 response element of RIG-I was required for RIG-I activation by retinoids or IFN. These results reveal that retinoid treatment and MeV infection induces significant RIG-I. RIG-I is required for the retinoid-MeV antiviral response. The induction is dependent on IFN, retinoids and IRF-1.

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RIG-I expression is regulated by MeV and ATRA.(A) U937 cells were infected with MeV at an MOI of 0.1 and treated with increasing doses of ATRA or DMSO for 24 hours. Some samples were also treated with 1000 U/mL IFNβ for 24 hours, with or without ATRA. RNA was extracted and analyzed for RIG-I expression by qPCR. Data presented are representative of three experiments performed in triplicate (N = 3). (B) U937 cells were infected with MeV at an MOI of 0.1 and treated with ATRA or DMSO (1 µM) for 48 hours, or with 2000 U/mL IFNβ as a positive control. Samples were analyzed by western blot for RIG-I and β-actin expression and quantified. ***p<0.001.
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pone-0022323-g001: RIG-I expression is regulated by MeV and ATRA.(A) U937 cells were infected with MeV at an MOI of 0.1 and treated with increasing doses of ATRA or DMSO for 24 hours. Some samples were also treated with 1000 U/mL IFNβ for 24 hours, with or without ATRA. RNA was extracted and analyzed for RIG-I expression by qPCR. Data presented are representative of three experiments performed in triplicate (N = 3). (B) U937 cells were infected with MeV at an MOI of 0.1 and treated with ATRA or DMSO (1 µM) for 48 hours, or with 2000 U/mL IFNβ as a positive control. Samples were analyzed by western blot for RIG-I and β-actin expression and quantified. ***p<0.001.

Mentions: We have previously shown that MeV can be inhibited in a number of cells lines including U937 cells and PBMCs [8]. To determine the involvement of RIG-I in the retinoid-mediated inhibition of MeV, the regulation of RIG-I expression during MeV infection with and without ATRA treatment was investigated in U937 cells. These cells are neoplastic and histiocytic progenitors of monocytes that have been extensively used in immunological studies [38]. They can be infected with MeV and are partially responsive to pharmacological doses of retinoids [8]. RIG-I mRNA and protein are expressed at very low levels in untreated U937 cells. MeV infection alone resulted in a small increase in RIG-I mRNA, while ATRA treatment alone had no discernible effect on RIG-I expression in this cell line. Importantly, U937 cells infected with MeV and treated with increasing doses of ATRA showed a dose response in RIG-I expression at the mRNA level (Figure 1A) and increased expression at the protein level (Figure 1B) over the induced over-expression of RIG-I by the artificial treatment with exogenous IFNβ. The IFNβ (positive control) could induce RIG-I expression as expected (Figure 1A). The combination of ATRA and IFNβ treatment resulted in higher levels of RIG-I expression than IFNβ alone (Figure 1A). Additionally, in our system we observe the up-regulation of a number of ISGs including IRF-7 [9] and MDA-5 (data not shown). RIG-I and MDA-5 have both been implicated in the induction of IFN in response to MeV [39]. The importance of RIG-I in this antiviral response due to its regulation by retinoids. To date, there has been no evidence that MDA-5 is a retinoid responsive gene.


RIG-I is required for the inhibition of measles virus by retinoids.

Soye KJ, Trottier C, Richardson CD, Ward BJ, Miller WH - PLoS ONE (2011)

RIG-I expression is regulated by MeV and ATRA.(A) U937 cells were infected with MeV at an MOI of 0.1 and treated with increasing doses of ATRA or DMSO for 24 hours. Some samples were also treated with 1000 U/mL IFNβ for 24 hours, with or without ATRA. RNA was extracted and analyzed for RIG-I expression by qPCR. Data presented are representative of three experiments performed in triplicate (N = 3). (B) U937 cells were infected with MeV at an MOI of 0.1 and treated with ATRA or DMSO (1 µM) for 48 hours, or with 2000 U/mL IFNβ as a positive control. Samples were analyzed by western blot for RIG-I and β-actin expression and quantified. ***p<0.001.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3139622&req=5

pone-0022323-g001: RIG-I expression is regulated by MeV and ATRA.(A) U937 cells were infected with MeV at an MOI of 0.1 and treated with increasing doses of ATRA or DMSO for 24 hours. Some samples were also treated with 1000 U/mL IFNβ for 24 hours, with or without ATRA. RNA was extracted and analyzed for RIG-I expression by qPCR. Data presented are representative of three experiments performed in triplicate (N = 3). (B) U937 cells were infected with MeV at an MOI of 0.1 and treated with ATRA or DMSO (1 µM) for 48 hours, or with 2000 U/mL IFNβ as a positive control. Samples were analyzed by western blot for RIG-I and β-actin expression and quantified. ***p<0.001.
Mentions: We have previously shown that MeV can be inhibited in a number of cells lines including U937 cells and PBMCs [8]. To determine the involvement of RIG-I in the retinoid-mediated inhibition of MeV, the regulation of RIG-I expression during MeV infection with and without ATRA treatment was investigated in U937 cells. These cells are neoplastic and histiocytic progenitors of monocytes that have been extensively used in immunological studies [38]. They can be infected with MeV and are partially responsive to pharmacological doses of retinoids [8]. RIG-I mRNA and protein are expressed at very low levels in untreated U937 cells. MeV infection alone resulted in a small increase in RIG-I mRNA, while ATRA treatment alone had no discernible effect on RIG-I expression in this cell line. Importantly, U937 cells infected with MeV and treated with increasing doses of ATRA showed a dose response in RIG-I expression at the mRNA level (Figure 1A) and increased expression at the protein level (Figure 1B) over the induced over-expression of RIG-I by the artificial treatment with exogenous IFNβ. The IFNβ (positive control) could induce RIG-I expression as expected (Figure 1A). The combination of ATRA and IFNβ treatment resulted in higher levels of RIG-I expression than IFNβ alone (Figure 1A). Additionally, in our system we observe the up-regulation of a number of ISGs including IRF-7 [9] and MDA-5 (data not shown). RIG-I and MDA-5 have both been implicated in the induction of IFN in response to MeV [39]. The importance of RIG-I in this antiviral response due to its regulation by retinoids. To date, there has been no evidence that MDA-5 is a retinoid responsive gene.

Bottom Line: Vitamin A can significantly decrease measles-associated morbidity and mortality.Retinoid signaling was also found to act in combination with IFN to induce high levels of RIG-I expression.IRF-1 is known to be regulated by retinoids and MeV, but we found recruitment of IRF-1 to the RIG-I promoter by retinoids alone.

View Article: PubMed Central - PubMed

Affiliation: McGill University Health Center Research Institute, Department of Infectious Diseases, McGill University, Montreal, Quebec, Canada.

ABSTRACT
Vitamin A can significantly decrease measles-associated morbidity and mortality. Vitamin A can inhibit the replication of measles virus (MeV) in vitro through an RARα- and type I interferon (IFN)-dependent mechanism. Retinoid-induced gene I (RIG-I) expression is induced by retinoids, activated by MeV RNA and is important for IFN signaling. We hypothesized that RIG-I is central to retinoid-mediated inhibition of MeV in vitro. We demonstrate that RIG-I expression is increased in cells treated with retinoids and infected with MeV. The central role of RIG-I in the retinoid-anti-MeV effect was demonstrated in the Huh-7/7.5 model; the latter cells having non-functional RIG-I. RAR-dependent retinoid signaling was required for the induction of RIG-I by retinoids and MeV. Retinoid signaling was also found to act in combination with IFN to induce high levels of RIG-I expression. RIG-I promoter activation required both retinoids and MeV, as indicated by markers of active chromatin. IRF-1 is known to be regulated by retinoids and MeV, but we found recruitment of IRF-1 to the RIG-I promoter by retinoids alone. Using luciferase expression constructs, we further demonstrated that the IRF-1 response element of RIG-I was required for RIG-I activation by retinoids or IFN. These results reveal that retinoid treatment and MeV infection induces significant RIG-I. RIG-I is required for the retinoid-MeV antiviral response. The induction is dependent on IFN, retinoids and IRF-1.

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Related in: MedlinePlus