HRZZ projects

How centrosome age affects the formation, resolution, and fate of polar chromosomes

Dividing human cells face two key competing tasks: faithfully segregating sister chromatids while avoiding long mitotic delays. The mitotic spindle rapidly attaches and aligns most chromosomes on the metaphase plate, but polar chromosomes, which start behind the emerging spindle poles, offer a particular challenge. Recent work in the Tolić laboratory showed that polar chromosomes are especially prone to being mis-segregated in cancer cells. How polar chromosomes are formed and resolved is, however, poorly understood. Interestingly, the two spindle poles are not equal in their ability to deal with polar chromosomes, as work of the Meraldi laboratory has established that the pole containing the old centrosome accumulates more polar chromosomes. Thus, the central hypothesis of this project is that centrosome age affects the formation, resolution, and fate of polar chromosomes. The main goal of this collaborative effort is to decipher the molecular and biophysical mechanisms controlling polar chromosomes, by combining the expertise of the Meraldi laboratory in centrosome age-dependent asymmetries and the Tolić laboratory in the biomechanics of polar chromosomes. We will investigate and manipulate the spindle parameters affecting polar chromosomes in human cell lines with a large spectrum of polar chromosomes asymmetries, and in a physiological context of Drosophila neuroblasts. This project will synergistically benefit from the expertise developed in both laboratories, including cutting-edge live-cell imaging assays, super-resolution microscopy, laser ablation, as well as theoretical modelling in collaboration with Nenad Pavin. Overall, the project will reveal a mechanistic link between the initial location of the chromosomes with respect to the old and young centrosome with the accuracy of their segregation, providing understanding of the origin of a prominent cause of chromosome segregation errors in cancer cells.

2023-2027 CENTROPOL
Research area: Life Sciences
ResearcherProf Iva Tolić and Prof Patrik Meraldi
Host Institution: Ruđer Bošković Institute (Croatia) and University of Geneva (Switzerland)
Project: How centrosome age affects the formation, resolution, and fate of polar chromosomes (CentroPol)
HRZZ call: Research Projects – Swiss-Croatian Bilateral Projects
HRZZ funding: 193.786,41 EUR

Mechanisms of microtubule bundle formation required for spindle maturation

During mitosis, the cell forms a spindle that segregates chromosomes symmetrically into two daughter cells. The mitotic spindle consists of several classes of microtubules, including kinetochore fibers that interact with chromosomes via kinetochores, and overlap or interpolar bundles that consist of antiparallel microtubules extending from the opposite spindle poles. A precise spatial organization of microtubule bundles and kinetochores is necessary for spindle function, and it has been to a large extent described for a metaphase spindle. However, during prometaphase, microtubules are unevenly distributed over the area of the future metaphase spindle, whereas kinetochore fibers and the associated overlap bundles are not yet evident. A central question is how during prometaphase these unevenly distributed microtubules become rearranged into discrete well-organized bundles, which position kinetochores into the metaphase plate. Interestingly, our pilot experiments show that the number of overlap bundles increases during prometaphase, and that new bundles can form by lateral splitting of the existing ones. To dissect the molecular mechanisms of bundle formation, we will develop an approach based on new CRISPR-edited human cell lines, which will be studied by live-cell confocal and superresolution microscopy. To identify key molecular players in bundle formation, we will modify the expression of candidate kinetochore proteins, motor proteins, and non-motor microtubule crosslinkers. Moreover, we will develop an optogenetic approach to remove a crosslinker protein from the spindle, and an approach based on bioactive peptides to modulate motor velocity. Finally, we will identify the sites of kinetochore fiber and overlap bundle formation and detyrosination, as well as the time of their formation and interaction. This project will reveal how microtubule bundles form, giving rise to the highly organized architecture of the mature metaphase spindle.

Research area: Cell biology
ResearcherProf Iva Tolić
Host Institution: Ruđer Bošković Institute, Zagreb, Croatia
Project: Mechanisms of microtubule bundle formation required for spindle maturation (SPINDLEFORM)
HRZZ call: Cooperation Programme with Croatian Scientists in Diaspora “RESEARCH COOPERABILITY”
HRZZ funding: HRK 2.200.000,00

Oscillatory dynamics of the cytoskeleton

Oscillations in the cytoskeleton are of general importance in cell biology as a mean of spatial and temporal regulation of the cellular organization, cell division and motility. The project OSCITON focuses on two oscillatory processes. First, we will investigate the mechanism of microtubule-driven kinetochore oscillations during mitosis, and the role of kinesin-8 motors in this process. Second, we will study the oscillatory repolarization of Dictyostelium cells during random migration. This interdisciplinary project is designed to optimize the use of resources and bring together researchers with the expertise in a range of disciplines, including molecular and cellular biology, biochemistry, microscopy, and theoretical biophysics.

2015-2019 OSCITON
Research area: Cell biology
Researcher: Prof Iva Tolić
Host Institution: Ruđer Bošković Institute, Zagreb, Croatia
Project: Oscilatory dynamics of the cytoskeleton (OSCITON)
HRZZ funding: HRK 1.000.000,00

HRZZ_Logo_NEWDisclaimer: Any opinions, findings, and conclusions or recommendations expressed in this material are those of the author(s) and do not necessarily reflect the views of Croatian Science Foundation, Ministry of Science and Education and European Commission.