Biophysical and cellular insights into the membrane insertion mechanism of CLIC1

Chloride intracellular channel 1 (CLIC1) is a human protein expressed in the cytosol that has a remarkable feature – it can transition into an active chloride channel in nuclear, endoplasmic reticulum or plasma membranes. This metamorphic nature makes CLIC1 a potential drug target as overexpression of the membrane channel form in cells is implicated in neurodegenerative disease progression and tumour proliferation especially in cancers with poor prognosis such as glioblastomas. The mechanism of CLIC1 activation and membrane insertion, as well as the oligomerisation state and structure of the channel, still remain elusive, therefore my PhD focuses on deciphering more information about how and why CLIC1 forms a chloride channel. Combining biophysical and microscopy techniques we have discovered that, upon binding to divalent cations Ca2+ and Zn2+, CLIC1 relocalises and inserts into the plasma membrane to form an active chloride channel in both in vitro and in vivo experiments. Previous literature heavily implicates the role of cysteine oxidation in CLIC1 channel formation but the use of solution NMR studies confirmed that both the soluble and membrane bound forms of CLIC1 are in the same oxidation state, further supporting the hypothesis divalent cations are the trigger for this translocation from a cytosolic globular protein to integral membrane chloride channel. The identification of the molecular switch that promotes CLIC1 membrane insertion is a significant discovery as it provides a model that can enable mechanistic studies of CLIC1 translocation and structural investigation of the channel form, known to have clinical relevance. Additional research focused on how CLIC1 interacts with divalent cations in a cellular environment aiming to elucidate information about the conformational changes the protein undergoes in vivo.