Abstract
Autophagy is a basic biological metabolic process involving in intracellular membrane transport pathways that recycle cellular components and eliminate intracellular microorganisms within the lysosome. Autophagy also plays an important part in virus infection and propagation. However, some pathogens, including viruses, have evolved unique trick to escape or exploit autophagy. This study explores the mechanism of autophagy induction by porcine hemagglutinating encephalomyelitis virus (PHEV) in Neuro-2a cells, and examines the role of autophagy in PHEV replication. PHEV triggered autophagy in Neuro-2a cells is dependent on the presence of bulk double- or single-membrane vacuoles, the accumulation of GFP-LC3 fluorescent dots, and the LC3 lipidation. In addition, PHEV induced an incomplete autophagic effect because the degradation level of p62 did not change in PHEV-infected cells. Further validation was captured using LysoTracker and lysosome-associated membrane protein by indirect immunofluorescence labeling in PHEV-infected cells. We also investigated the change in viral replication by pharmacological experiments with the autophagy inducer rapamycin or the autophagy inhibitor 3-MA, and the lysosomal inhibitor chloroquine (CQ). Suppression of autophagy by 3-MA increased viral replication, compared with the mock treatment, while promoting of autophagy by rapamycin reduced PHEV replication. CQ treatment enhanced the LC3 lipidation in PHEV-infected Neuro-2a cells but lowered PHEV replication. These results show that PHEV infection induces atypical autophagy and causes the appearance of autophagosomes but blocks the fusion with lysosomes, which is necessary for the replication of PHEV in nerve cells.
Highlights
Porcine hemagglutinating encephalomyelitis virus (PHEV) is a single-stranded, positive-sense RNA coronavirus and is classified as a member of the Coronaviridae family (Andries and Pensaert, 1981)
We found that the induction of autophagy with rapamycin (200 μM) for 2 h effectively upregulated the alteration of LC3-I to LC3-II but significantly inhibited porcine hemagglutinating encephalomyelitis virus (PHEV) replication at 24 and 48 hpi in Neuro-2a cells (Figure 6A)
PHEV is a neurotropic virus that transmits along the peripheral nerves to the central nervous system (CNS) (Li et al, 2013)
Summary
Porcine hemagglutinating encephalomyelitis virus (PHEV) is a single-stranded, positive-sense RNA coronavirus and is classified as a member of the Coronaviridae family (Andries and Pensaert, 1981). PHEV Induces Atypical Autophagy in neurological damage by virus infection (Andries and Pensaert, 1980). Nervous lesions caused by pathogens often lead to abnormal nerve cell morphology, unnatural protein accumulation, vesicular transport or neuronal gene transcription disorders, and synaptic transmission obstacles (Xilouri and Stefanis, 2010). As a virus invading the nervous system, rabies virus (RABV) can cause severe encephalitis and induce analogous cytopathic features (Peng et al, 2016). A more detailed description of the mechanism should be revealed, especially the biological changes of the host cells induced by infection
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