Junín Virus Promotes Autophagy To Facilitate the Virus Life Cycle

Abstract

Junín virus (JUNV), a member of the family Arenaviridae, is the etiological agent of Argentine hemorrhagic fever (AHF), a potentially deadly endemicepidemic disease affecting the population of the most fertile farming land of Argentina. Autophagy is a degradative process with a crucial antiviral role; however, several viruses subvert the pathway to their benefit. We determined the role of autophagy in JUNV-infected cells by analyzing LC3, a cytoplasmic protein (LC3-I) that becomes vesicle membrane associated (LC3-II) upon induction of autophagy. Cells overexpressing enhanced green fluorescent protein (EGFP)-LC3 and infected with JUNV showed an increased number of LC3 punctate structures, similar to those obtained after starvation or bafilomycin A1 treatment, which leads to autophagosome induction or accumulation, respectively. We also monitored the conversion of LC3-I to LC3-II, observing LC3-II levels in JUNV-infected cells similar to those observed in starved cells. Additionally, we kinetically studied the number of LC3 dots after JUNV infection and found that the virus activated the pathway as early as 2 h postinfection (p.i.), whereas the UV-inactivated virus did not induce the pathway. Cells subjected to starvation or pretreated with rapamycin, a pharmacological autophagy inductor, enhanced virus yield. Also, we assayed the replication capacity of JUNV in Atg5 knockout or Beclin 1 knockdown cells (both critical components of the autophagic pathway) and found a significant decrease in JUNV replication. Taken together, our results constitute the first study indicating that JUNV infection induces an autophagic response, which is functionally required by the virus for efficient propagation. IMPORTANCE Mammalian arenaviruses are zoonotic viruses that cause asymptomatic and persistent infections in their rodent hosts but may produce severe and lethal hemorrhagic fevers in humans. Currently, there are neither effective therapeutic options nor effective vaccines for viral hemorrhagic fevers caused by humanpathogenic arenaviruses, except the vaccine Candid no. 1 against Argentine hemorrhagic fever (AHF), licensed for human use in areas of endemicity in Argentina. Since arenaviruses remain a severe threat to global public health, more in-depth knowledge of their replication mechanisms would improve our ability to fight these viruses. Autophagy is a lysosomal degradative pathway involved in maintaining cellular homeostasis, representing powerful anti-infective machinery. We show, for the first time for a member of the family Arenaviridae, a proviral role of autophagy in JUNV infection, providing new knowledge in the field of host-virus interaction. Therefore, modulation of virus-induced autophagy could be used as a strategy to block arenavirus infections.

Silencing long non‐coding RNA DLX6‐AS1 or restoring microRNA‐193b‐3p enhances thyroid carcinoma cell autophagy and apoptosis via depressing HOXA1

“…LC3 was a cytoplasmic protein (LC3‐I), which was converted into vesicular membrane‐associated protein (LC3‐II) after inducing autophagy. 22 LC3‐II was an important marker molecule necessary for the formation of autophagosomes and selective recruitment of substrates, which increased with the increase of autophagosome membranes. 23 Detection of LC3‐II and LC3‐I by Western blot assay manifested that LC3‐II/LC3‐I ratio increased by DLX6‐AS1 inhibition.…”

“…CCK-8, colony formation and Transwell assays, along with flow cytometry, was in application in identifying DLX6-AS1 regulation-induced effects on TC cell progression. The findings concluded that depleted DLX6-AS1 restrained K1 cell proliferation, invasion, migration and triggered apoptosis (Figure2B-F).LC3 was a cytoplasmic protein (LC3-I), which was converted into vesicular membrane-associated protein (LC3-II) after inducing autophagy 22. LC3-II was an important marker molecule necessary for the formation of autophagosomes and selective recruitment of substrates, which increased with the increase of autophagosome membranes 23.…”

Differential Immune Responses to Hemorrhagic Fever-Causing Arenaviruses

“…The role of autophagy during arenavirus infection is largely unknown, but recent research has provided evidence of a proviral role for autophagy. Two groups observed that both virulent and attenuated strains of JUNV induce autophagy early in infection [86,87]. Induction of autophagy prior to infection results in increased NP expression [87].…”

“…The OW MOPV also transiently induces autophagy during the first two days of infection, though LASV does not [88]. Nevertheless, depleting the essential autophagic vesicle protein ATG5 impairs MOPV, LASV, and JUNV replication [86][87][88], indicating that both OW and NW arenaviruses may utilize components of the autophagy pathway to aid in replication. While ATG5 is the key component in autophagy, there is increasing evidence that ATG5 also plays roles in non-autophagy processes, including negative regulation of RIG-I or MDA5 [89][90][91].…”

“…The mechanism behind autophagy induction in arenavirus infection is unclear. One group determined that, unlike replication-competent JUNV, UV-inactivated JUNV did not induce autophagy up to 9 hours post-exposure [86]. However, another group observed autophagy induction 24 hours after exposure to UV-inactivated JUNV [87], though it is unclear whether this was virus-driven.…”