A novel assay allows simultaneous detection of
individual proteins and their interactions in living cells.
Dr. Ruth Brack-Werner, Director of the GSF
Institute of Molecular Virology (IMV) explains the decisive advantage
of the new approach: "In previous assays, signals were generated only
by interacting proteins, whereas the individual partners remained
undetected. However, the absence of signals could not be used to rule
out protein interactions since the absence of one or both interaction
partners would have the same effect. To overcome this problem
Brack-Werner and her team developed the so-called extended bimolecular
fluorescence complementation (exBiFC) which allows simultaneous
monitoring of individual proteins and their interactions.
Brack-Werner and her colleagues' groundbreaking
research work focusses on mechanisms that control replication of the
human immunodeficiency virus (HIV), which causes AIDS. "HIV
replication is based on the interaction of cellular proteins with
viral proteins. Interactions involving viral regulatory factors have a
direct impact on the amount of virus produced by the HIV host cell",
Brack-Werner explains. "Preventing HIV proteins from interacting with
their crucial partners is a promising approach to developing novel
therapies." Therefore the GSF-scientists developed and validated
exBiFC with the HIV Rev protein, which is an accelerator of HIV
production. Various assays investigating Rev interactions in
artificial settings indicate that the activity of Rev depends on the
interaction of Rev molecules with each other and with cellular
proteins. The latter include Exportin 1, which transports proteins
from the nucleus to the cytoplasm and RISP, a modulator of HIV gene
expression discovered by the Brack-Werner team in previous studies.
Brack-Werner and her team demonstrated that exBIFC allows
visualization of interactions of Rev with itself and with Exportin1
and RISP in living cells. In addition they were able to compare the
strengths of the interactions of Rev with its partners by analysing
the intensities of the signals in cell images.
ExBiFC has a wide range of potential appllications
and represents an important tool for the elucidation of protein
interaction networks and discovery of novel antiviral factors. Thus
exBIFC has an enormous potential in the battle against leading global
health problems such as infectious diseases and cancers.
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