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Basal Autophagy Is Required for Herpes simplex Virus-2 Infection.

Yakoub AM, Shukla D - Sci Rep (2015)

Bottom Line: Thus, we investigated the importance of basal autophagy for HSV-2 infection, using pharmacological autophagy suppression or cells genetically deficient in an autophagy-essential gene (ATG5).Interference with basal autophagy flux in cells significantly reduced viral replication and diminished the infection.These results indicate that basal autophagy plays an indispensable role required for a productive infection.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Microbiology and Immunology, University of Illinois, Chicago, IL USA, 60612 [2] Department of Ophthalmology and Visual Sciences, University of Illinois Medical Center, Chicago, IL USA, 60612.

ABSTRACT
Autophagy is a conserved catabolic process of the cell, which plays an important role in regulating plethora of infections. The role of autophagy in Herpes simplex virus-2 (HSV-2) infection is unknown. Here, we found that HSV-2 does not allow induction of an autophagic response to infection, but maintains basal autophagy levels mostly unchanged during productive infection. Thus, we investigated the importance of basal autophagy for HSV-2 infection, using pharmacological autophagy suppression or cells genetically deficient in an autophagy-essential gene (ATG5). Interference with basal autophagy flux in cells significantly reduced viral replication and diminished the infection. These results indicate that basal autophagy plays an indispensable role required for a productive infection. Importantly, this study draws a sharp distinction between induced and basal autophagy, where the former acts as a viral clearance mechanism abrogating infection, while the latter supports infection.

No MeSH data available.


Related in: MedlinePlus

Validation of autophagy deficiency in ATG5−/− cells.(a) LC3-GFP-expressing WT or ATG5−/− MEFs were cultured in regular serum-containing medium (Fed), or starved for 3 hr. The cells were then fixed and imaged using confocal microscopy at 63x magnification. (b) Quantification of the number of LC3 punctae per cells, from confocal microscopy experiments performed as in (a). An average of 50 cells was assessed and used for counting. (c) Quantification of cells with ≥10 LC3 punctae, from confocal microscopy experiments performed as in (a). An average of 50 cells was assessed.
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f3: Validation of autophagy deficiency in ATG5−/− cells.(a) LC3-GFP-expressing WT or ATG5−/− MEFs were cultured in regular serum-containing medium (Fed), or starved for 3 hr. The cells were then fixed and imaged using confocal microscopy at 63x magnification. (b) Quantification of the number of LC3 punctae per cells, from confocal microscopy experiments performed as in (a). An average of 50 cells was assessed and used for counting. (c) Quantification of cells with ≥10 LC3 punctae, from confocal microscopy experiments performed as in (a). An average of 50 cells was assessed.

Mentions: Before using the ATG5−/− cells in our virus assays, we wanted to validate their autophagy deficiency. We monitored basal autophagosomal levels in normal feeding conditions and induced autophagosomal levels under nutrient loss (starvation). Using GFP-LC3-expressing MEFs, we were able to detect low basal levels of autophagy in wild-type (WT) cells, but much less levels in ATG5−/− cells (Fig. 3a–c). In addition, ATG5−/− cells were also defective in mounting an autophagic response to starvation (Fig. 3a–c). These results validated the autophagy-deficient nature of these cells.


Basal Autophagy Is Required for Herpes simplex Virus-2 Infection.

Yakoub AM, Shukla D - Sci Rep (2015)

Validation of autophagy deficiency in ATG5−/− cells.(a) LC3-GFP-expressing WT or ATG5−/− MEFs were cultured in regular serum-containing medium (Fed), or starved for 3 hr. The cells were then fixed and imaged using confocal microscopy at 63x magnification. (b) Quantification of the number of LC3 punctae per cells, from confocal microscopy experiments performed as in (a). An average of 50 cells was assessed and used for counting. (c) Quantification of cells with ≥10 LC3 punctae, from confocal microscopy experiments performed as in (a). An average of 50 cells was assessed.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4528227&req=5

f3: Validation of autophagy deficiency in ATG5−/− cells.(a) LC3-GFP-expressing WT or ATG5−/− MEFs were cultured in regular serum-containing medium (Fed), or starved for 3 hr. The cells were then fixed and imaged using confocal microscopy at 63x magnification. (b) Quantification of the number of LC3 punctae per cells, from confocal microscopy experiments performed as in (a). An average of 50 cells was assessed and used for counting. (c) Quantification of cells with ≥10 LC3 punctae, from confocal microscopy experiments performed as in (a). An average of 50 cells was assessed.
Mentions: Before using the ATG5−/− cells in our virus assays, we wanted to validate their autophagy deficiency. We monitored basal autophagosomal levels in normal feeding conditions and induced autophagosomal levels under nutrient loss (starvation). Using GFP-LC3-expressing MEFs, we were able to detect low basal levels of autophagy in wild-type (WT) cells, but much less levels in ATG5−/− cells (Fig. 3a–c). In addition, ATG5−/− cells were also defective in mounting an autophagic response to starvation (Fig. 3a–c). These results validated the autophagy-deficient nature of these cells.

Bottom Line: Thus, we investigated the importance of basal autophagy for HSV-2 infection, using pharmacological autophagy suppression or cells genetically deficient in an autophagy-essential gene (ATG5).Interference with basal autophagy flux in cells significantly reduced viral replication and diminished the infection.These results indicate that basal autophagy plays an indispensable role required for a productive infection.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Microbiology and Immunology, University of Illinois, Chicago, IL USA, 60612 [2] Department of Ophthalmology and Visual Sciences, University of Illinois Medical Center, Chicago, IL USA, 60612.

ABSTRACT
Autophagy is a conserved catabolic process of the cell, which plays an important role in regulating plethora of infections. The role of autophagy in Herpes simplex virus-2 (HSV-2) infection is unknown. Here, we found that HSV-2 does not allow induction of an autophagic response to infection, but maintains basal autophagy levels mostly unchanged during productive infection. Thus, we investigated the importance of basal autophagy for HSV-2 infection, using pharmacological autophagy suppression or cells genetically deficient in an autophagy-essential gene (ATG5). Interference with basal autophagy flux in cells significantly reduced viral replication and diminished the infection. These results indicate that basal autophagy plays an indispensable role required for a productive infection. Importantly, this study draws a sharp distinction between induced and basal autophagy, where the former acts as a viral clearance mechanism abrogating infection, while the latter supports infection.

No MeSH data available.


Related in: MedlinePlus