II. The function of molecular motor proteins in chromosome segregation
Chromosomal instability (CIN), i.e. the unequal segregation of the genome during mitosis, can result in aneuploidy, a hallmark of most solid tumors. Thus, the integrity and development of each eukaryotic organism is intimately tied to an error free process of mitotic chromosome segregation and cells have evolved complex mechanisms to accomplish this challenging task. Within this context, we are particular interested in the alignment of chromosomes at the spindle equator and the function of the mitotic kinesin Kif18A in this process.
Kif18A belongs to the kinesin-8 family and is characterized by its unique ability to not only move along microtubules but also to modulate their dynamic behavior once it reaches the plus-ends of kinetochore microtubules. Depletion of Kif18A in HeLa cells results in severe mitotic defects with misaligned chromosomes and elongated spindle phenotypes. To dissect the function of Kif18A at distinct mitotic stages, we performed a protein-based screen to identify small molecule inhibitors of Kif18A. These studies yielded in the identification of BTB-1, a small molecule that inhibits the ATPase activity of Kif18A in a reversible and ATP competitive manner.
Currently, we are further optimizing the chemical properties of BTB-1. In future studies, we plan to combine BTB-1 with time-resolved fluorescence microscopy to analyze the function of Kif18A in late mitosis.
The key function of Kif18A in chromosome congression suggests that the activity of Kif18A has to be tightly controlled during mitotic progression to ensure the precise positioning of chromosomes at the spindle equator. To gain insights into the underlying regulatory mechanisms, we are currently combining biochemical in vitro studies with mass spectrometry and live cell microscopy analyses. These combined approaches already revealed that Kif18A is subject to a complex regulatory network of posttranslational modifications.
Model of the antagonistic regulation of Kif18A by Cdk1 and PP1, for details see
Future studies will reveal how the different modifications adjust Kif18A function to the requirements of distinct mitotic stages and how these modifications impact the mechanochemical properties of Kif18A.