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Cleavage of a Neuroinvasive Human Respiratory Virus Spike Glycoprotein by Proprotein Convertases Modulates Neurovirulence and Virus Spread within the Central Nervous System.

Le Coupanec A, Desforges M, Meessen-Pinard M, Dubé M, Day R, Seidah NG, Talbot PJ - PLoS Pathog. (2015)

Bottom Line: Human coronaviruses (HCoV) are respiratory pathogens that may be associated with the development of neurological diseases, in view of their neuroinvasive and neurotropic properties.Taken together, these results are consistent with the adaptation of HCoV-OC43 to the CNS environment, resulting from the selection of quasi-species harboring mutations that lead to amino acid changes in viral genes, like the S gene in HCoV-OC43, which may contribute to a more efficient establishment of a less pathogenic but persistent CNS infection.This adaptative mechanism could potentially be associated with human encephalitis or other neurological degenerative pathologies.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Neuroimmunovirology, INRS-Institut Armand-Frappier, Université du Québec, Laval, Québec, Canada.

ABSTRACT
Human coronaviruses (HCoV) are respiratory pathogens that may be associated with the development of neurological diseases, in view of their neuroinvasive and neurotropic properties. The viral spike (S) glycoprotein is a major virulence factor for several coronavirus species, including the OC43 strain of HCoV (HCoV-OC43). In an attempt to study the role of this protein in virus spread within the central nervous system (CNS) and neurovirulence, as well as to identify amino acid residues important for such functions, we compared the sequence of the S gene found in the laboratory reference strain HCoV-OC43 ATCC VR-759 to S sequences of viruses detected in clinical isolates from the human respiratory tract. We identified one predominant mutation at amino acid 758 (from RRSR↓ G758 to RRSR↓R758), which introduces a putative furin-like cleavage (↓) site. Using a molecular cDNA infectious clone to generate a corresponding recombinant virus, we show for the first time that such point mutation in the HCoV-OC43 S glycoprotein creates a functional cleavage site between the S1 and S2 portions of the S protein. While the corresponding recombinant virus retained its neuroinvasive properties, this mutation led to decreased neurovirulence while potentially modifying the mode of virus spread, likely leading to a limited dissemination within the CNS. Taken together, these results are consistent with the adaptation of HCoV-OC43 to the CNS environment, resulting from the selection of quasi-species harboring mutations that lead to amino acid changes in viral genes, like the S gene in HCoV-OC43, which may contribute to a more efficient establishment of a less pathogenic but persistent CNS infection. This adaptative mechanism could potentially be associated with human encephalitis or other neurological degenerative pathologies.

No MeSH data available.


Related in: MedlinePlus

A delay in viral spread is observed in brains of rOC/SG758R -infected mice compared to rOC/ATCC after intracerebral infection in 21 day-old BALB/c female mice.Histological examination of virus spread within the brain. 21 day-old BALB/c mice received 102.5TCID50/10μL of rOC/ATCC or rOC/SG758R, or PBS by the IC route. Detection of viral antigens in the olfactory bulb (A) or in the hippocampus (B) of infected mice at 5 and 7 dpi at magnitude X40. Black arrows indicate viral particles staining for the S protein.
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ppat.1005261.g004: A delay in viral spread is observed in brains of rOC/SG758R -infected mice compared to rOC/ATCC after intracerebral infection in 21 day-old BALB/c female mice.Histological examination of virus spread within the brain. 21 day-old BALB/c mice received 102.5TCID50/10μL of rOC/ATCC or rOC/SG758R, or PBS by the IC route. Detection of viral antigens in the olfactory bulb (A) or in the hippocampus (B) of infected mice at 5 and 7 dpi at magnitude X40. Black arrows indicate viral particles staining for the S protein.

Mentions: Given our observation that reference virus was more neurovirulent compared to the mutant virus after inoculation by the IC route, we wished to evaluate whether this correlated with a difference in viral replication in the CNS. Brains and spinal cords were harvested and infectious virus titers were assayed every 2 days for a period of 21 dpi (Fig 3E and 3F). The difference in neurovirulence did not correlate with a significant difference in the amount of infectious viral particles in the brain (Fig 3E). However, there was a drastic difference in the production of infectious virus between both variants in the spinal cord (Fig 3F): virus titers of the reference strain (rOC/ATCC) were almost identical to what was detected in the brain, whereas the less virulent mutant (rOC/SG758R) reached the spinal cord only in one out of thirty infected mice. In this mouse, an important delay and a production of viral infectious particles close to the limit of detection suggested that mutant virus had difficulty reaching this portion of the CNS. Histological examination of infected mice revealed that the infected regions were similar following infection by both viruses in the brain, but that the kinetics were different (Fig 4). Indeed, as was the case after the IN route of infection, the IC route of infection also led to a delay in viral replication in the olfactory bulb and in the hippocampus, as no viral antigens were detected before 7 dpi for the mutant virus (compared to 5 dpi for the reference virus). As in 10 day-old BALB/c mice infected IN, when virus had spread to all regions of the brain, activation of astrocytes and microglial cells was evident in all infected regions (S2 Fig). As seen in 10 day-old mice after IN inoculation, even though no precise quantitation was performed, a slight increase in the number of astrocytes in the olfactory bulb (S2A Fig) and in the hippocampus (S2B Fig) could be observed in brains of mice infected by reference virus compared to the mutant virus. The same was observed for microglial cells at 7 dpi in the olfactory bulb (S2C Fig) and in the hippocampus (S2D Fig).


Cleavage of a Neuroinvasive Human Respiratory Virus Spike Glycoprotein by Proprotein Convertases Modulates Neurovirulence and Virus Spread within the Central Nervous System.

Le Coupanec A, Desforges M, Meessen-Pinard M, Dubé M, Day R, Seidah NG, Talbot PJ - PLoS Pathog. (2015)

A delay in viral spread is observed in brains of rOC/SG758R -infected mice compared to rOC/ATCC after intracerebral infection in 21 day-old BALB/c female mice.Histological examination of virus spread within the brain. 21 day-old BALB/c mice received 102.5TCID50/10μL of rOC/ATCC or rOC/SG758R, or PBS by the IC route. Detection of viral antigens in the olfactory bulb (A) or in the hippocampus (B) of infected mice at 5 and 7 dpi at magnitude X40. Black arrows indicate viral particles staining for the S protein.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4636366&req=5

ppat.1005261.g004: A delay in viral spread is observed in brains of rOC/SG758R -infected mice compared to rOC/ATCC after intracerebral infection in 21 day-old BALB/c female mice.Histological examination of virus spread within the brain. 21 day-old BALB/c mice received 102.5TCID50/10μL of rOC/ATCC or rOC/SG758R, or PBS by the IC route. Detection of viral antigens in the olfactory bulb (A) or in the hippocampus (B) of infected mice at 5 and 7 dpi at magnitude X40. Black arrows indicate viral particles staining for the S protein.
Mentions: Given our observation that reference virus was more neurovirulent compared to the mutant virus after inoculation by the IC route, we wished to evaluate whether this correlated with a difference in viral replication in the CNS. Brains and spinal cords were harvested and infectious virus titers were assayed every 2 days for a period of 21 dpi (Fig 3E and 3F). The difference in neurovirulence did not correlate with a significant difference in the amount of infectious viral particles in the brain (Fig 3E). However, there was a drastic difference in the production of infectious virus between both variants in the spinal cord (Fig 3F): virus titers of the reference strain (rOC/ATCC) were almost identical to what was detected in the brain, whereas the less virulent mutant (rOC/SG758R) reached the spinal cord only in one out of thirty infected mice. In this mouse, an important delay and a production of viral infectious particles close to the limit of detection suggested that mutant virus had difficulty reaching this portion of the CNS. Histological examination of infected mice revealed that the infected regions were similar following infection by both viruses in the brain, but that the kinetics were different (Fig 4). Indeed, as was the case after the IN route of infection, the IC route of infection also led to a delay in viral replication in the olfactory bulb and in the hippocampus, as no viral antigens were detected before 7 dpi for the mutant virus (compared to 5 dpi for the reference virus). As in 10 day-old BALB/c mice infected IN, when virus had spread to all regions of the brain, activation of astrocytes and microglial cells was evident in all infected regions (S2 Fig). As seen in 10 day-old mice after IN inoculation, even though no precise quantitation was performed, a slight increase in the number of astrocytes in the olfactory bulb (S2A Fig) and in the hippocampus (S2B Fig) could be observed in brains of mice infected by reference virus compared to the mutant virus. The same was observed for microglial cells at 7 dpi in the olfactory bulb (S2C Fig) and in the hippocampus (S2D Fig).

Bottom Line: Human coronaviruses (HCoV) are respiratory pathogens that may be associated with the development of neurological diseases, in view of their neuroinvasive and neurotropic properties.Taken together, these results are consistent with the adaptation of HCoV-OC43 to the CNS environment, resulting from the selection of quasi-species harboring mutations that lead to amino acid changes in viral genes, like the S gene in HCoV-OC43, which may contribute to a more efficient establishment of a less pathogenic but persistent CNS infection.This adaptative mechanism could potentially be associated with human encephalitis or other neurological degenerative pathologies.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Neuroimmunovirology, INRS-Institut Armand-Frappier, Université du Québec, Laval, Québec, Canada.

ABSTRACT
Human coronaviruses (HCoV) are respiratory pathogens that may be associated with the development of neurological diseases, in view of their neuroinvasive and neurotropic properties. The viral spike (S) glycoprotein is a major virulence factor for several coronavirus species, including the OC43 strain of HCoV (HCoV-OC43). In an attempt to study the role of this protein in virus spread within the central nervous system (CNS) and neurovirulence, as well as to identify amino acid residues important for such functions, we compared the sequence of the S gene found in the laboratory reference strain HCoV-OC43 ATCC VR-759 to S sequences of viruses detected in clinical isolates from the human respiratory tract. We identified one predominant mutation at amino acid 758 (from RRSR↓ G758 to RRSR↓R758), which introduces a putative furin-like cleavage (↓) site. Using a molecular cDNA infectious clone to generate a corresponding recombinant virus, we show for the first time that such point mutation in the HCoV-OC43 S glycoprotein creates a functional cleavage site between the S1 and S2 portions of the S protein. While the corresponding recombinant virus retained its neuroinvasive properties, this mutation led to decreased neurovirulence while potentially modifying the mode of virus spread, likely leading to a limited dissemination within the CNS. Taken together, these results are consistent with the adaptation of HCoV-OC43 to the CNS environment, resulting from the selection of quasi-species harboring mutations that lead to amino acid changes in viral genes, like the S gene in HCoV-OC43, which may contribute to a more efficient establishment of a less pathogenic but persistent CNS infection. This adaptative mechanism could potentially be associated with human encephalitis or other neurological degenerative pathologies.

No MeSH data available.


Related in: MedlinePlus