Mechanistic and functional dissection of the multifunctional mitotic motor Kif15

• Applicant: Dr. Hauke Drechsler
• Subject Area: Biophysics ; Biochemistry ; Cell Biology
• Term: from 2019 to 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 417890911

Mitotic spindle assembly and maintenance is driven by forces generated from two functionally redundant tetrameric molecular motors; kinesin-5 (Eg5) and kinesin-12 (Kif15). Due to its central role in spindle assembly, current antiproliferative cancer treatment approaches aim to inhibit Eg5 activity thereby restricting tumour growth. However, cells chronically treated with Eg5-inhibitors quickly acquire inhibitor resistance by upregulating Kif15 expression. In general, the overexpression of Eg5 and Kif15 results in increased growth, survivability and invasiveness of tumour cells.While Eg5 mechanics during spindle assembly are well established, the mechanistic role of the multi-functional Kif15 in this process is still under debate. In particular, it remains elusive, which of the multiple but distinct cellular subpopulations of Kif15 on the mitotic spindle and chromosomes or which functions of Kif15 contribute to spindle assembly and the emergence of Eg5-inhbitior resistance. Interestingly, Kif15 also localises to actin structures during cell cleavage and cell migration. So far, Kif15’s link to the actin cytoskeleton has been little studied, but initial results imply that Kif15 is involved in cell migration control, which may have important implications for the metastatic behaviour of tumour cells overexpressing Kif15.To dissect the mechanics of spindle assembly, Eg5-inhibitor resistance and cell migration control by Kif15, my work programme aims to link cellular subpopulations and particular molecular functions of Kif15 to the respective mechanical processes. In a hypothesis-driven approach, I will study Kif15 variants that carry either point mutations or short deletions within a newly mapped regulatory hub in the stalk of Kif15 that abolish either i) the motor’s ability to tetramerise, ii) its interaction with particular cellular subcompartments or iii) its association with the actin cytoskeleton. As certain motor functions depend on its oligomerisation state, constitutively dimeric Kif15 (cdKif15) variants are expected to act as separation-of-function mutants. All Kif15 variants will be analysed for their substructure localisation in vivo, their oligomerisation state and their ability to bind known interaction partners. I further will biophysically and functionally characterise various Kif15 variants in vitro and test their capacity to rescue spindle assembly in the absence of Eg5-activity in vivo. I will also characterise Kif15’s link to the actin cytoskeleton on the molecular level using recombinant components and study the impact on interaction-deficient motors on cell cleavage and - with Dr. Anne Straube at Warwick University (UK) - on cell migration. Linking particular cellular subpopulations and functions of Kif15 to spindle assembly and cell migration control will identify potential targets of future cancer treatment approaches that seek to restrict tumour growth and metastasis while preventing Eg5-inhbitor resistance formation.

• DFG Programme: Research Grants