ABSTRACT
The cell entry mechanism of SARS-CoV-2, the causative agent of the COVID-19 pandemic, is not fully understood. Most animal viruses hijack cellular endocytic pathways as an entry route into the cell. Here, we show that in cells that do not express serine proteases such as TMPRSS2, genetic depletion of all dynamin isoforms blocked the uptake and strongly reduced infection with SARS-CoV-2 and its variant Delta. However, increasing the viral loads partially and dose-dependently restored infection via a thus far uncharacterized entry mechanism. Ultrastructural analysis by electron microscopy showed that this dynamin-independent endocytic processes appeared as 150-200 nm non-coated invaginations and was efficiently used by numerous mammalian viruses, including alphaviruses, influenza, vesicular stomatitis, bunya, adeno, vaccinia, and rhinovirus. Both the dynamin-dependent and dynamin-independent infection of SARS-CoV-2 required a functional actin cytoskeleton. In contrast, the alphavirus Semliki Forest virus, which is smaller in diameter, required actin only for the dynamin-independent entry. The presence of TMPRSS2 protease rescued SARS-CoV-2 infection in the absence of dynamins. Collectively, these results indicate that some viruses such as canine parvovirus and SARS-CoV-2 mainly rely on dynamin for endocytosis-dependent infection, while other viruses can efficiently bypass this requirement harnessing an alternative infection entry route dependent on actin.