Post by Nadica (She/Her) on Nov 24, 2024 1:13:17 GMT
SARS-CoV-2 infection in hiPSC-derived neurons is cathepsin-dependent and causes accumulation of HIF1alpha and phosphorylated tau - Preprint posted Nov 23, 2024
Abstract
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been shown to infect the human brain and a subset of human neurons in vitro. We have previously demonstrated that the virus enters the human induced pluripotent stem cell (hiPSC)-derived neurons via an endosomal-lysosomal pathway, which is dependent on low levels of angiotensin-converting enzyme 2 (ACE2) and independent of transmembrane serine protease 2 (TMPRSS2). Here, we use hiPSC-derived neurons overexpressing ACE2 in co-culture with human astrocytes to show that the infection with both SARS-CoV-2 Wuhan and Omicron XBB.1.5 variants is dependent on cathepsins and can be efficiently blocked by an inhibitor of cathepsin B (CA-074-ME). The result was reproducible in non-transgenic hiPSC-derived cortical organoids. The cathepsin L inhibitor SB412515 was less effective against the Wuhan strain but equally effective against the Omicron variant. Using PCR and reinfection assays, we show that SARS-CoV-2 can replicate in neurons in 2D co-cultures. Interestingly, the infectivity of the newly produced virions declined at 24 hours post-infection despite a further increase in released viral RNA at later time points, suggesting the possible activation of an antiviral response in neurons and/or astrocytes, which is supported by a correspondent increase in the levels of secreted cytokines. Furthermore, the number of infected neurons decreased within five days, suggesting that SARS-CoV-2 infection eventually leads to the death of the target neuronal cell in vitro. The infection also caused the accumulation of the hypoxia-inducible stress factor HIF1-alpha in infected neurons under normoxia. Finally, we confirm and expand the previous finding that in SARS-CoV-2 infected neurons, the microtubule-associated protein tau is hyperphosphorylated at multiple loci, including S202/T205, and mislocalized to the soma of the infected neurons. Hyperphosphorylation and mislocalization of tau are hallmarks of Alzheimer's disease (AD) and other tauopathies. Our data provides further evidence supporting the neurodegenerative potential of SARS-CoV-2 infection.
Abstract
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been shown to infect the human brain and a subset of human neurons in vitro. We have previously demonstrated that the virus enters the human induced pluripotent stem cell (hiPSC)-derived neurons via an endosomal-lysosomal pathway, which is dependent on low levels of angiotensin-converting enzyme 2 (ACE2) and independent of transmembrane serine protease 2 (TMPRSS2). Here, we use hiPSC-derived neurons overexpressing ACE2 in co-culture with human astrocytes to show that the infection with both SARS-CoV-2 Wuhan and Omicron XBB.1.5 variants is dependent on cathepsins and can be efficiently blocked by an inhibitor of cathepsin B (CA-074-ME). The result was reproducible in non-transgenic hiPSC-derived cortical organoids. The cathepsin L inhibitor SB412515 was less effective against the Wuhan strain but equally effective against the Omicron variant. Using PCR and reinfection assays, we show that SARS-CoV-2 can replicate in neurons in 2D co-cultures. Interestingly, the infectivity of the newly produced virions declined at 24 hours post-infection despite a further increase in released viral RNA at later time points, suggesting the possible activation of an antiviral response in neurons and/or astrocytes, which is supported by a correspondent increase in the levels of secreted cytokines. Furthermore, the number of infected neurons decreased within five days, suggesting that SARS-CoV-2 infection eventually leads to the death of the target neuronal cell in vitro. The infection also caused the accumulation of the hypoxia-inducible stress factor HIF1-alpha in infected neurons under normoxia. Finally, we confirm and expand the previous finding that in SARS-CoV-2 infected neurons, the microtubule-associated protein tau is hyperphosphorylated at multiple loci, including S202/T205, and mislocalized to the soma of the infected neurons. Hyperphosphorylation and mislocalization of tau are hallmarks of Alzheimer's disease (AD) and other tauopathies. Our data provides further evidence supporting the neurodegenerative potential of SARS-CoV-2 infection.