Subcellular Structure and cellular Dynamics

Our lab is interested in understanding how centrosomes (organelle that functions as the main microtubule (MT) organizing center) and cilia (organelle essential for motility or sensory perception) regulate various cellular processes and how these influence development, proliferation and the establishment of diseases.

Fig. 1The two centrosomes of the cell (shown in yellow) nucleate and organize the MT network (red) to assemble a bipolar mitotic spindle. The chromosomes are shown in blue.

At the ultra structural level, the centrosome comprises a pair of centrioles (barrel shape microtubule (MT) structures) surrounded by a matrix of proteins, the pericentriolar material or PCM. The PCM is the site of MT nucleation during somatic mitosis and it is now well established that centrosomes are not essential for mitosis but increase the efficiency of mitotic spindle assembly and cytokinesis. Centrosomes have, however, essential roles in polarity establishment and asymmetric cell division. For example, during fertilization of the C. elegans embryo, the centrosome, brought in by the sperm, specifies the posterior of the embryo. In addition, spindle positioning during asymmetric cell division also relies on the presence of functional centrosomes.

Fig. 2Electron microscopy (EM) picture of a cross section of Drosophila sperm tail flagellum.
 

In differentiated cells, centrioles can localize near the plasma membrane and behave as basal bodies. Basal bodies nucleate two types of MT structures: motile cilia or flagella and primary cilia. In recent years, we have realised how important these structures are. For example, primary cilia are crucial for mechano, chemo or photo perception in all animals whereas motile cilia or flagella are essential for movement of fluids in our body or sperm motility.

Centrosome and cilia dysfunction are associated with a variety of human diseases such as microcephaly, lissencefaly, cancer or infertility. It is therefore, essential, to characterize the basic mechanisms that regulate centriole replication and cilia assembly in the context of a developing organism to understand how mutations in centrosome, basal body and cilia components can lead to the establishment of disease.

We use a variety of different approaches that include molecular and cell biology methods, genetics and tissue culture to answer to these questions.

Fig. 3Paramecium tetraurelia stained with basal body markers.

Our work on centriole replication is primarily focused on the characterization of centriole replication mutants in the fruit fly Drosophila melanogaster. We are also using other model organisms such as the multiciliated Paramecium to investigate the role of centriole replication components in basal body duplication and cilia assembly.

Another area of research in the lab investigates how centrosomes and MTs influence spindle positioning during stem cell divisions. Stem cells divide asymmetrically to generate another stem cell (self-renewal) and a differentiating cell. Asymmetric cell division defects can lead to an imbalance between self-renewal and differentiation, which can contribute to over-proliferation and tumour formation. We use Drosophila neural stem cells (neuroblasts) to investigate the fundamental mechanisms that regulate these divisions.

Movie 1: Wild-type neuroblast co-expressing RFP-Tubulin to visualize the mitotic spindle and GFP-Pon, an adaptor protein that binds to Numb, a cell fate determinant. This cell divides asymmetrically and produces a large neuroblast and a small cell, the ganglion mother cell. GFP-Pon is only inherited by the GMC.

Movie 2: DSas-4 mutant neuroblast. This cell does not contain centrosomes and the spindle fails to be aligned along the polarity axis. The cell divides symmetrically and the two daughter cells inherit GFP-Pon.

We have recently characterized Drosophila fly lines that contain supernumerary centrosomes. We have noticed that during cell division, cells with extra centrosomes manage to assemble a functional bipolar spindle. Moreover, we have shown that centrosome amplification can also lead to over-proliferation and tumour formation. We are currently investigating which mechanism can either facilitate or inhibit tumour formation when extra centrosomes are present.

Movie 3: Wild-type cell co-expressing RFP-Tubulin to visualize the mitotic spindle and GFP-DSas-4 to visualize the centrosomes. The two centrosomes assemble a bipolar mitotic spindle and after division each cell inherits a single centrosome.

Movie 4: A cell with three centrosomes. As the cell enter into mitosis, the extra centrosomes cluster and form the poles of a bipolar spindle. Centrosome clustering is an efficient process since we rarely observed defects in cell division.