Related Experiment Videos
Regulation of pH by the M2 protein of influenza A viruses
F Ciampor1, C A Thompson, S Grambas
1National Institute for Medical Research, London, U.K.
Abstract:
Inhibition of the function of the M2 protein by amantadine can cause a conformational change in the haemagglutinin (HA) of H7 influenza A viruses and the consequent expression of the low pH form of the glycoprotein on the surface of virus-infected cells. Immunofluorescence studies showed that this conversion occurs shortly after HA exists from the Golgi complex apparently during its transport through the trans Golgi network and using the pH probe, DAMP/anti-DNP, that it is the direct result of reduced vesicular pH. The lowest pHs encountered were estimated using mutant HAs differing in pH stability to be approximately 5.2 and 5.6 in virus-infected CEF or MDCK cells, respectively, in the absence of functional M2. Depending on the particular M2, this protein was responsible for increases in vesicular pH of up to 0.8 units. The influence of mutations in both HA and M2 on the maturation of native HA illustrates the important relationship between the structural and functional properties of these two proteins. Using the fluorescent probe SNARF-1 the M2 protein was also shown to be largely responsible for the 0.3-0.4 unit reduction in intracellular pH of virus-infected cells. The data thus provide further evidence for the pH regulatory function of M2 and its importance for the maturation of the HA glycoprotein.
Insights
Amantadine inhibits influenza A virus M2 protein, causing a conformational change in hemagglutinin (HA) and low pH expression. This highlights M2
Area of Science:
- Virology
- Molecular Biology
- Cell Biology
Background:
- Influenza A virus hemagglutinin (HA) undergoes conformational changes crucial for viral entry.
- The M2 protein of influenza A virus plays a role in regulating the pH of intracellular vesicles.
- Amantadine is a known inhibitor of the M2 protein.
Purpose of the Study:
- To investigate the effect of M2 protein inhibition by amantadine on HA conformation and expression.
- To determine the role of M2 protein in regulating vesicular and intracellular pH.
- To elucidate the relationship between M2 protein function and HA maturation.
Main Methods:
- Immunofluorescence studies to observe HA expression on infected cells.
- Use of pH probes (DAMP/anti-DNP and SNARF-1) to measure vesicular and intracellular pH.
- Employing mutant HAs with varying pH stability to estimate pH values.
- Analysis of the impact of mutations in both HA and M2 proteins.
Main Results:
- Amantadine-induced M2 inhibition leads to HA conformational changes and expression of the low pH form.
- This conversion is a direct result of reduced vesicular pH during HA transport through the Golgi.
- The M2 protein is responsible for increasing vesicular pH by up to 0.8 units and reducing intracellular pH by 0.3-0.4 units.
Conclusions:
- The M2 protein plays a critical role in regulating intracellular and vesicular pH.
- M2 protein function is essential for the proper maturation of the HA glycoprotein.
- The structural and functional interplay between HA and M2 is vital for influenza virus replication.