p16INK4a is a tumour suppressor protein that regulates cell cycle progression by binding to and inhibiting the cyclin-dependent kinases 4 and 6 (CDK4/6). It is described as a tumour suppressor as it induces cell cycle arrest, and its function is therefore frequently lost in cancer.
p16 is a 16 kDa protein comprised of four ankyrin repeats joined by three loop regions. Recently, our group has discovered that the single cysteine residue of p16 is able to undergo oxidation under relatively mild oxidizing conditions which leads to disulfide-dependent homo‑dimerization. This dimerization leads to subsequent transition from the native alpha‑helical structure of p16, into beta-sheet based amyloid fibrils. In this rearranged amyloid fibril state. p16 is unable to perform its normal function as a CDK4/6 inhibitor.1
We recently had the opportunity to send samples to the International Space Station (ISS). Radiation is well known to generate reactive oxygen species, and as we now know that oxidizing conditions trigger the ‘shapeshifting’ of p16 from monomeric to amyloid state, we aimed to explore the effects of cosmic radiation and microgravity conditions aboard the ISS on this unique transition. Analyses of the structural state and functionality of p16 following cosmic radiation exposure were carried out using SDS-PAGE, thioflavin T aggregation assays, kinase functional assays, transmission electron microscopy, mass spectrometry and filter trap assays. Results from this detailed analysis show intriguing differences between ISS samples compared to control samples. To further explore the effects of radiation, experiments were conducted at higher radiation exposures using caesium-137 radiation.
Overall, this work contributes to our understanding of the mechanism and potential pathogenic role of p16 amyloid formation.