Molecular Biology of Rotaviruses
Rotavirus is a major cause of gastroenteritis in young mammals responsible for more than 200,000 deaths each year worldwide (mainly children under 5 in developing countries). Rotaviruses are also the source of significant economic losses in agriculture. Due to its multisegmented double-stranded RNA genome and the originality of its cycle (non-enveloped virus that does not release its genome into the cytoplasm. , budding in the ER with a transient lipid envelope, synthesis of non-polyadenylated messenger RNAs, etc.), rotaviruses offer an excellent model for many fundamental questions of biology; entry of viruses into the cell, modification of cellular genes expression, expression of viral genes (transcription and translation), selective molecular recognition (packaging of 11 double-stranded RNAs molecules into each viral capsid), molecular machine (transcription)
Despite the importance of rotavirus infections for human and veterinary healthes, many steps of the viral cycle are poorly understood. The goal of the rotavirus team is to better understand the biology of these viruses at the molecular level. The rotavirus team is conducting basic research aimed at understanding the functioning of rotavirus at the level of its interaction with the host, in particular at the level of the regulation of translation of viral genes and cellular genes.
Regulation of cellular and viral mRNA translation
The rotavirus NSP3 protein by interacting simultaneously with the 3 ′ end of rotavirus mRNAs and with the translation factor eIF4G, promotes viral translation. We were able to show that whereas rotavirus infection stimulates the expression of the viral reporter mRNAs and decreases the expression of the poly (A) RNAs, NSP3 alone increases the translation of the viral RNAs more than 100 times but also stimulates the expression of poly (A) RNAs. RT-qPCR analysis of the RNAs does not show stabilization of the viral RNAs by NSP3 and the use of NSP3 mutants shows that the increase in translation requires the two domains of NSP3 (RNA- and eIF4G- binding domains). Therefore, contrary to what had been suggested by others authors, the increase in translation of viral mRNAs by NSP3 is not explained by the simple stabilization of viral RNAs and the two functional domains of NSP3 are not independent. The inhibition of poly (A) RNA translation during infection is not due to the expression of the NSP3 protein alone and eviction of PABP from translation initiation complexes, but is a consequence of the saturation of the cellular machinery by the viral mRNAs.
Alternative splicing of the XBP1 gene
Expression of stress genes is altered by rotavirus infection. The study of the XBP1 stress gene during rotavirus infection allowed us to show that the rotavirus induces an alternative splicing of this gene. We were able to show by genetic analysis of reassortant viruses that NSP3 was responsible for this phenomenon. Using chimeras of NSP3 obtained by reverse genetics, we showed that the eIF4G-binding domain of NSP3 is involved.
It is quite remarquable that a strictly cytoplasmic virus, is able to perturb a nuclear function by interacting with a translation factor.
Entry of the rotavirus into the cells.
The mechanism of entry of rotavirus is still unclear. We have recently resumed work of the team showing that trypsin (protease necessary for the cleavage of the spicule protein VP4 necessary for the entry of the virus into the cells) remained associated with the viral particles. We are currently investigating the role of this association in virus entry by studying viruses carrying mutations on the VP4 protein.
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