Function and interactions of a protein bridge between the inner membrane complex and subpellicular microtubules in Toxoplasma gondii.

Toxoplasma gondii is an intracellular parasite that utilizes peripheral membrane and cytoskeletal structures for essential functions such as host cell invasion and replication. These include the inner membrane complex (IMC) and the underlying longitudinal subpellicular microtubules (SPMT) that provide support for the IMC and give the parasite its distinctive crescent shape. Although the IMC and SPMTs have been studied separately, the mechanisms linking these adjacent structures remain largely unknown. This study identifies a protein named IMT1 that localizes to the maternal IMC and SPMTs and appears to tether the IMC to the microtubules. We disrupt the IMT1 gene to assess function and then use deletion analyses and mutagenesis to reveal regions of the protein that are necessary for binding to the IMC cytoskeleton or SPMTs. Using proximity labeling, we identify candidate IMT1 interactors in the IMC or SPMTs. Exploration of these candidates reveals that the loss of IMT1 results in a dramatic reduction of the microtubule-associated protein TLAP2 and that IMT1 binds directly to the cytoskeletal IMC proteins IMC1, IMC18, and IMC24. Together, these interactions reveal a novel bridge that connects two key cytoskeletal structures and provides new insight into the organization of the structural backbone of T. gondii.