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Etude de la régulation de l'épissage du transcrit primaire du virus de l'immunodeficience humaine : interaction et mécanisme d'action des protéines SR et hnRNP au niveau du site accepteur A3

Abstract : Alternative splicing of virus HIV-1 RNA plays a key role for virus multiplication. By the combined utilization of 4 donor and 8 acceptor splicing sites at least 30 distinct mRNAs are produced in infected cells. We studied the mechanism of regulation of one essential acceptor site (site A3). Its utilization is needed for production of tat mRNAs. Protein Tat is required for the viral RNA production. As Tat is toxic for the infected cells, its production is highly controlled at the splicing step. Two splicing silencers act at site A3 : ESS2 and ESS2p, they bind proteins hnRNP A1 and hnRNP H, respectively. We showed that by binding to ESS2p, protein hnRNP H limits site A3 activity by steric hindrance. By using molecular and structural methods, we explained how the interaction of hnRNP A1 with ESS2, located 69 nt downstream from site A3, blocks the utilization of this site. Our experimental data reveal that the cooperative binding and multimerization of hnRNP A1 on the 69 nt region located downstream from site A3 is initiated by simultaneous association of at least two hnRNP A1 molecules. One of them interacts directly with ESS2 by its RRM domains. The Gly domain of hnRNP A1 strongly stabilizes this complex. Mutations in ESS2 decrease hnRNP A1 binding on the entire RNA region and increase splicing efficiency. We showed that the activation of site A3 by proteins SRp40 and SC35 is due to their competition with hnRNP A1 for binding to ESS2. Finally, we found that HIV-1 RNAs of chronically infected cells, which lost their ability to produce new virions, contain 2 mutations in the A3 regulatory region that increase hnRNP A1 binding and block site A3 utilization.
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http://docnum.univ-lorraine.fr/prive/SCD_T_2006_0135_HALLAY.pdf
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https://hal.univ-lorraine.fr/tel-01746583
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Submitted on : Thursday, March 29, 2018 - 10:40:43 AM
Last modification on : Friday, February 26, 2021 - 3:02:02 PM

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  • HAL Id : tel-01746583, version 1

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Houda Hallay. Etude de la régulation de l'épissage du transcrit primaire du virus de l'immunodeficience humaine : interaction et mécanisme d'action des protéines SR et hnRNP au niveau du site accepteur A3. Biochimie, Biologie Moléculaire. Université Henri Poincaré - Nancy 1, 2006. Français. ⟨NNT : 2006NAN10135⟩. ⟨tel-01746583⟩

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