BioTechniques News
Maddy Chapman
Researchers seem to have demonstrated that the much-debated interleukin, IL-39, can’t be secreted by human cells, but it could serve as a valuable engineered tool for immunology and drug design.
A recent research effort from a team at the Technical University of Munich (Germany), working in HEK293 cells, has shed light on the debate surrounding the possibility of a naturally occurring human interleukin 39 (IL-39) and used its findings to engineer a modified version of the cytokine that could prove useful in therapeutic development.
Interleukins play a critical role in the stimulation and modulation of immune cells. A subset of these interleukins, known as the IL-12 family, is composed of one of three α subunits and one of two β subunits, resulting in a potential six members of the family. However, only four members have been definitively characterized in humans, and much debate surrounds the possibility that a fifth, IL-39, which is composed of the α subunit IL-23α and the β subunit EBI3 and is found in mice, exists in humans.
Seeking to address this debate, the team, led by corresponding author Matthias Feige, transfected human IL-23α and EBI3 into HEK293T cells before using coimmunoprecipitation experiments, involving pulldown of IL-23α with FLAG tag and an immunoblot for an HA tag added onto EBI3 to establish whether the two components were associated within cells. This was followed by secretion assays comparing proteins in cell lysate vs cell media to establish whether the subunits assembled together and were successfully expressed to form fully folded IL-39 in the HEK293T cells.
They found no such association from the coimmunoprecipitation or any evidence of secretion of IL-23α induced by EBI3, indicating that IL-39 did not naturally assemble in these cells.
Not satisfied with this fact alone, the team used MODELLER to model the structure of a potential IL-23α-EBI3 heterodimer and found several critical weaknesses, such as cavities in key positions, which destabilized the molecule and would not allow it to form.
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Toying with several mutant versions of EBI3 designed to stabilize the heterodimer, they found that there was some indication that IL-23α and EBI3 could potentially assemble in the cell, but only very weakly. With further engineering of both components, the researchers were able to design a version of IL-39 that could be secreted by the cells and even demonstrated the ability to bind the receptors IL-23R and gp130 on human immune cells, stimulating inflammatory pathways.
It is important to note that there is still the possibility of the heterodimer forming outside of cells once secreted but, speaking to the impact of their study, the authors explain that they have created the tools needed to fully address the question of whether IL-39 can be endogenously produced under certain conditions, once and for all.
They also explain that the importance of their findings extends beyond this question:
“This highlights that rational engineering of synthetic cytokines provides a tool to regulate immune cells that express the receptors needed to respond to cytokines that do not occur naturally or are produced only under specific circumstances. Engineered IL- 39 may thus provide a way to specifically induce immune cell activation.”
Perhaps the toolbox with which researchers can both investigate and directly treat immunological conditions and common diseases has just grown by one…
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