Human astrovirus C-terminal nsP1a protein is involved in RNA replication

Guix, Susana; Caballero, Santiago; Bosch, Albert; Pintó, Rosa M.

doi:10.1016/j.virol.2004.12.023

Cited by 32 publications

(38 citation statements)

References 31 publications

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“…The template used for the amplification of nsP1a/4 genotype XII and RdRp coding sequences was the plasmid pAVIC6 (kindly provided by S. Matsui, Gastroenterology Section, Veterans Administration Palo Alto Health Care System, Palo Alto, CA), which contains the full genome of HAstV-1. The templates used for the amplification of nsP1a/4 genotypes IV, V, and VI were the previously described plasmids pAVIC-IV, pAVIC-V, and pAVIC-VI (8), which contain the nsP1a/4 coding region of genotypes IV, V, and VI, respectively. The primers used to amplify all these sequences are shown in Table 1.…”

Section: Methodsmentioning

confidence: 99%

“…nsP1a/4 protein interacts with the viral polymerase. The involvement of nsP1a/4 in viral RNA replication as well as its interaction with the endoplasmic reticulum (7,8) suggests that this protein may be an integral part of the viral replication complex interacting with the RNA polymerase. The potential interaction between nsP1a/4 and RNA polymerase was studied through coimmunoprecipitation of these proteins from ex- ) was used as a negative control.…”

Section: Production Of Astrovirus Nsp1a/4 and Rdrp In Insect Cellsmentioning

confidence: 99%

“…Biological and functional analyses also showed a colocalization of the nsP1a/4 with the endoplasmic reticulum and viral RNA and revealed a role of this protein in RNA replication, as well as a relationship between the genetic diversity within the HVR of the nsP1a/4 coding region and the virus replication phenotype (7,8). Two patterns of replication that are associated with the nsP1a/4 genotype have been described: genotypes IV and V show higher levels of subgenomic RNA and lower levels of antigenomic RNA than genotypes VI and XII (8).…”

mentioning

confidence: 96%

“…In addition, computational analyses of the nsP1a/4 coding region performed in our laboratory revealed the presence of acidic regions and of glutamine-and proline-rich regions, a death domain, and several putative O-glycosylation and phosphorylation sites (6)(7)(8). Biological and functional analyses also showed a colocalization of the nsP1a/4 with the endoplasmic reticulum and viral RNA and revealed a role of this protein in RNA replication, as well as a relationship between the genetic diversity within the HVR of the nsP1a/4 coding region and the virus replication phenotype (7,8). Two patterns of replication that are associated with the nsP1a/4 genotype have been described: genotypes IV and V show higher levels of subgenomic RNA and lower levels of antigenomic RNA than genotypes VI and XII (8).…”

mentioning

confidence: 97%

“…Based on the HVR nucleotide genetic variability, 15 different nsP1a/4 protein HVR-derived genotypes (named using Roman numerals) have been established, and a restriction fragment length polymorphism (RFLP) typing method has been developed to consistently distinguish between genotypes (9). In addition, computational analyses of the nsP1a/4 coding region performed in our laboratory revealed the presence of acidic regions and of glutamine-and proline-rich regions, a death domain, and several putative O-glycosylation and phosphorylation sites (6)(7)(8). Biological and functional analyses also showed a colocalization of the nsP1a/4 with the endoplasmic reticulum and viral RNA and revealed a role of this protein in RNA replication, as well as a relationship between the genetic diversity within the HVR of the nsP1a/4 coding region and the virus replication phenotype (7,8).…”

mentioning

confidence: 99%

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The C-Terminal nsP1a Protein of Human Astrovirus Is a Phosphoprotein That Interacts with the Viral Polymerase

Fuentes

Guix

Bosch

et al. 2011

J Virol

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Human astrovirus nonstructural C-terminal nsP1a protein (nsP1a/4) colocalizes with the endoplasmic reticulum and viral RNA. It has been suggested that nsP1a/4 protein is involved in the RNA replication process in endoplasmic reticulum-derived intracellular membranes. A hypervariable region (HVR) is contained in the nsP1a/4 protein, and different replicative patterns can be distinguished depending on its variability. In the present work, both the astrovirus RNA-dependent RNA polymerase and four types (IV, V, VI, and XII) of nsP1a/4 proteins have been cloned and expressed in the baculovirus system to analyze their interactions. Different isoforms of each of the nsP1a/4 proteins exist: a nonphosphorylated isoform and different phosphorylated isoforms. While the polymerase accumulates as a monomer, the nsP1a/4 proteins accumulate as oligomers. The oligomerization domain of nsP1a/4-V is mapped between residues 176 and 209. For all studied genotypes, oligomers mainly contain the nonphosphorylated isoform. When RNA polymerase is coexpressed with nsP1a/4 proteins, they interact, likely forming heterodimers. The polymerase binding region has been mapped in the nsP1a/4-V protein between residues 88 and 176. Phosphorylated isoforms of nsP1a/4 type VI show a stronger interactive pattern with the polymerase than the nonphosphorylated isoform. This difference is not observed in genotypes IV and V, suggesting a role of the HVR in modulating the interaction of the nsP1a/4 protein with the polymerase through phosphorylation/dephosphorylation of some critical residues.

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Section: Methodsmentioning

confidence: 99%

Section: Production Of Astrovirus Nsp1a/4 and Rdrp In Insect Cellsmentioning

confidence: 99%

mentioning

confidence: 96%

mentioning

confidence: 97%

mentioning

confidence: 99%

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The C-Terminal nsP1a Protein of Human Astrovirus Is a Phosphoprotein That Interacts with the Viral Polymerase

Fuentes

Guix

Bosch

et al. 2011

J Virol

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Genetic analysis of the hypervariable region of the human astrovirus nsp1a coding region: Design of a new RFLP typing method

Guix

Caballero

Fuentes

et al. 2007

Journal of Medical Virology

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Human astroviruses (HAstV) are causative agents of viral gastroenteritis worldwide. A hypervariable region (HVR) is located close to the C-terminus of the nsP1a, and recent data support the involvement of the HVR-containing nonstructural protein in viral RNA replication processes, suggesting a correlation between variability in this region and pathogenic properties. The HVR of the C-terminal nsP1a coding region of 104 wild-type and reference isolates of HAstV was sequenced. A phylogenetic analysis was performed to identify different genotypes, and a restriction fragment length polymorphism (RFLP) method was designed. An extensive nucleotide and deduced amino acid sequence variability was observed, as well as many insertions and deletions that retained the reading frame. The resultant phylogenetic tree supported the subdivision of HAstV into the two previously described major genetic groups, genogroup A and B, and the identification of 12 genotypes (9 within genogroup A, and 3 within genogroup B), which could be identified by RFLP. A correlation analysis was performed between genotype information and viral load using information from 35 clinical samples. Significant differences were observed between the viral load in clinical samples and certain HAstV genotypes that belonged to the same serotype, confirming the influence of C-terminal nsP1a variability on the viral replication phenotype. The use of the new RFLP typing method based on the HVR of the C-terminal nsP1a coding region by diagnosticians would help to understand the relationship between different genotypes and the severity of the gastroenteritis.

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