Researchers capture live footage of virus infecting cell

by Stephanie Dutchen, Harvard Medical School

Schematic representation of the principal entry routes SARS-CoV-2 uses for infection. 

Entry starts with membrane attachment and ends with S protein–catalyzed membrane

 fusion releasing the viral contents into the cytosol. Fusion activity depends on two

 proteolytic cleavage steps, namely, one typically carried out by furin in the producing cell 

and the second by TMPRSS2 on the cell surface on in endosomes of the target cell. 

Alternatively, endosomal cathepsins can carry out both cleavages. Exposure of the virus 

to an acidic milieu is essential for membrane fusion, genome penetration, and productive 

infection. Fusion and penetration occur only in acidic early and late endosomal/lysosomal 

compartments but not at the cell surface, even when the furin and TMPRSS2 cleavages 

have both occurred. Fusion and penetration can occur at the cell surface of cells expressing

 TMPRSS2 if the extracellular pH is ∼6.8.

 Credit: Proceedings of the National Academy of Sciences (2022). DOI: 10.1073/pnas.2209514119

In a first, scientists have captured on video all the steps a virus follows as it enters and infects a living cell in real time and in three dimensions.

Scientists achieved the feat by using advanced imaging called lattice light sheet microscopy as well as chemical and genetic manipulation.

The first part of the video shown here follows a virus engineered to sprout SARS-CoV-2 spike proteins (labeled pink) as it is captured at a cell surface and engulfed by a cellular compartment called an endosome. The virus then fuses with the endosome membrane and injects its genetic material (labeled blue) inside the cell—the steps necessary to kick off a cycle of viral infection and replication.

The second part of the video shows many such viruses inside the cell. The video covers 4 minutes of activity, with snapshots taken every 4 seconds.

The findings, published Sept. 1 in PNAS, provide new insights into the fundamental mechanics of viral infection and could point the way to new methods for intervening before the onset of COVID-19.

The researchers' work reveals that viruses can't fuse with the membrane and release their genomes unless they're bathed in a slightly acidic environment. Experiments indicated that the pH must fall between 6.2 and 6.8, just shy of neutral and on par with bodily fluids such as saliva and urine. Endosomes have such acidity, and the team's measurements confirmed that this is also the pH range inside a typical human nose, where SARS-CoV-2 infection often begins.

Credit: Harvard Medical School

"Amusingly enough, measuring the pH of the nostril cavity has rarely been done before," noted co-senior author Tomas Kirchhausen, professor of cell biology in the Blavatnik Institute at Harvard Medical School and HMS professor of pediatrics at Boston Children's Hospital.

The acidic environment allows enzymes in the endosome or on the cell surface—including TMPRSS2, a key enabler of SARS-CoV-2 infection—to cut the spike protein and facilitate membrane fusion, the team found.

The work was led by the labs of Kirchhausen; former HMS professor Sean Whelan, now at Washington University in St. Louis; and Giuseppe Balistreri at the University of Helsinki. Alex Kreutzberger, HMS instructor in pediatrics in the Kirchhausen lab, is first author of the paper.Powerful new antibody neutralizes all known SARS-CoV-2 variants

More information: Alex J. B. Kreutzberger et al, SARS-CoV-2 requires acidic pH to infect cells, Proceedings of the National Academy of Sciences (2022). DOI: 10.1073/pnas.2209514119

Journal information: Proceedings of the National Academy of Sciences 

Provided by Harvard Medical School