tetano
Editor, Senior Moderator
Appl Microbiol Biotechnol
. 2025 Mar 24;109(1):74.
doi: 10.1007/s00253-025-13423-3. Quantification of intracellular influenza A virus protein dynamics in different host cells after seed virus adaptation
Jan Küchler[SUP] 1 [/SUP], Patricia Opitz[SUP] 2 [/SUP], Ingo Jordan[SUP] 3 [/SUP], Yvonne Genzel[SUP] 4 [/SUP], Dirk Benndorf[SUP] 5 [/SUP], Udo Reichl[SUP] 4 2 [/SUP]
Affiliations
Influenza A virus is a major human pathogen, and its replication is widely studied. One important aspect for effective virus propagation is the host cell, since cellular properties can limit or favor virus entry, viral genome and viral protein synthesis and virus release. To establish detailed mathematical models for these processes, quantitative experimental data on the intracellular dynamics of viral compounds together with the number of infectious and non-infectious virus particles released are required. In this study, we report results obtained from an optimized mass spectrometry assay for the quantification of viral proteins that was applied to compare the production of influenza A virus HA, NP, NA, M1, and NS1 proteins for different seed viruses and host cells of batch cultures. With canine MDCK cell-adapted seed virus, a maximum of about 1.0E+08 copies/cell were found for all five viral proteins after infection of avian AGE1.CR and human HEK293 cells. These intracellular levels are about fivefold lower than in MDCK cells. However, after five passages of seed virus adaptation, intracellular protein copy numbers comparable to those in MDCK cells were achieved. Highest levels were found for the NS1 protein with about 1.0E+09 copies/cell. Furthermore, the onset of virus particle release started earlier for both cell lines (about 3-6 h). In contrast, the maximum virus titers did not change for AGE1.CR cells but increased for HEK293 cells. Nevertheless, the highest HA titers were always obtained for MDCK cells. Overall, the experimental data indicate that influenza A virus replication is different due to specifics of innate host cell immune response, viral protein production, precursor consumption, and degradation rates. KEY POINTS: • Application of absolute quantification for five major proteins of influenza A virus. • NS1 protein most abundant protein with 1.0E+09 copies/cell at the end of infection. • Virus adaptation leads to earlier release and higher virus titers in HEK293 cell.
Keywords: Absolute quantification; Influenza A virus; Mass spectrometry; Virus adaptation
. 2025 Mar 24;109(1):74.
doi: 10.1007/s00253-025-13423-3. Quantification of intracellular influenza A virus protein dynamics in different host cells after seed virus adaptation
Jan Küchler[SUP] 1 [/SUP], Patricia Opitz[SUP] 2 [/SUP], Ingo Jordan[SUP] 3 [/SUP], Yvonne Genzel[SUP] 4 [/SUP], Dirk Benndorf[SUP] 5 [/SUP], Udo Reichl[SUP] 4 2 [/SUP]
Affiliations
- PMID: 40126655
- DOI: 10.1007/s00253-025-13423-3
Influenza A virus is a major human pathogen, and its replication is widely studied. One important aspect for effective virus propagation is the host cell, since cellular properties can limit or favor virus entry, viral genome and viral protein synthesis and virus release. To establish detailed mathematical models for these processes, quantitative experimental data on the intracellular dynamics of viral compounds together with the number of infectious and non-infectious virus particles released are required. In this study, we report results obtained from an optimized mass spectrometry assay for the quantification of viral proteins that was applied to compare the production of influenza A virus HA, NP, NA, M1, and NS1 proteins for different seed viruses and host cells of batch cultures. With canine MDCK cell-adapted seed virus, a maximum of about 1.0E+08 copies/cell were found for all five viral proteins after infection of avian AGE1.CR and human HEK293 cells. These intracellular levels are about fivefold lower than in MDCK cells. However, after five passages of seed virus adaptation, intracellular protein copy numbers comparable to those in MDCK cells were achieved. Highest levels were found for the NS1 protein with about 1.0E+09 copies/cell. Furthermore, the onset of virus particle release started earlier for both cell lines (about 3-6 h). In contrast, the maximum virus titers did not change for AGE1.CR cells but increased for HEK293 cells. Nevertheless, the highest HA titers were always obtained for MDCK cells. Overall, the experimental data indicate that influenza A virus replication is different due to specifics of innate host cell immune response, viral protein production, precursor consumption, and degradation rates. KEY POINTS: • Application of absolute quantification for five major proteins of influenza A virus. • NS1 protein most abundant protein with 1.0E+09 copies/cell at the end of infection. • Virus adaptation leads to earlier release and higher virus titers in HEK293 cell.
Keywords: Absolute quantification; Influenza A virus; Mass spectrometry; Virus adaptation