The Rusan Lab studies the role of centrosomes during animal development. The centrosome is a non-membrane bound organelle that serves as the main microtubule (MT) organizing center of most animal cells. Centrosomes function to initiate and maintain cell polarity, guide cell migration, direct intracellular cargos, and properly distribute other organelles. In mitosis, centrosomes are critical for accurate construction of the mitotic spindle to ensure faithful chromosome segregation to the two daughter cells. Thus, it is not a surprise that defects in centrosome function lead to a wide range of failures at the cellular level, which in turn, leads to tissue defects and many human diseases. The lab aims to determine how centrosomes are properly constructed from their individual parts and how centrosomes function in a wide range of cell types to avoid human diseases such as polycystic kidney disease, microcephaly, cancer and many others.
To study the centrosome, the Rusan Lab uses Drosophila genetics, cell biology, super-resolution microscopy, biochemistry, and modern molecular biology. Most recently, the lab undertook a targeted yeast-2-hybrid to define the direct protein-protein interaction (PPI) landscape of the centrosome. This centrosome ‘interactome’, in combination with the above mentioned techniques, is used to dig deep into the mechanisms of centrosome protein function. A general theme has emerged from these studies, that is, protein-protein network diversity is responsible for the diversity of centrosome functions across cell types and cell cycle stages. In other words, not all centrosomes are made equally. Understanding this centrosome PPI diversity is a priority because it could help explain why different mutations in centrosome proteins affect specific cell types and manifest different diseases.
The Rusan Lab’s current work focuses on two proteins, Pericentrin-Like-Protein (PLP) and Asterless (Asl), referred to as ‘linker’ proteins because they bridge the two main components of the centrosome – centrioles and the Pericentriolar Material (PCM). The centrosome interactome truly highlights how little is known about these two proteins. PLP and Asl interact with a large number of partners and the Rusan Lab is in the process of defining the role of each of these interactions with respect to short timescales (cell cycle) and long timescales (different developmental stages) in a variety of cell types. This work will shed light on how mutations in Pericentrin, the human ortholog of PLP, lead to microcephalic osteodysplastic primordal dwarfism type II (MOPD II).
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The centrosome undergoes massive structural and functional changes as it transitions through the cell cycle (Figure). These changes include doubling the number of centrosomes from one to two and modulating centrosome activity (number of microtubules nucleated and anchored). These two major events are controlled by the Centriole Duplication and Centrosome Maturation cycles, respectively.
One of our main goals is to understand how a centrosome is constructed from its individual component. With hundreds of proteins found at the centrosome, dozens of which are essential for proper centrosome function, we know relatively little about this massive organelle. We have put forward a large effort to identify direct protein-protein interaction at the centrosome. More importantly, we aim to test the importance of these interactions for centrosome, cell and organism function. To do this, we use a combination of Y2H analysis, modern molecular genetics and live cell imaging in animals to determine the role of these interactions in the Duplication and Maturation cycles.
Centrosomes in Development
The importance of centrosomes for proper cell and animal function has been appreciated for some time now. Diseases caused by defective centrosomes fall into two general categories, ciliopathies and cell division defects. For example, studies using genetic knockout cell lines show that loss of centrioles in mitotic cells can directly lead to aberrant mitotic spindle formation, defective chromosome segregation, and aneuploidy. Likewise, supernumerary centrosomes is a phenotype found in nearly all cancers, including brain, breast, colon, lung, pancreas and prostate. In fact, studies in Drosophila stem cells show that centrosome over-production can initiate stem cell tumors. Although a similar direct link has not been identified in mammalian cell transformation, increased centrosome number has been shown to cause aneuoploidy and microcephaly.
Our lab aims to understand how centrosomes guide proper tissue patterning by controlling cell polarity and mitotic division axes in order to avoid these detrimental defects. We investigate the mechanisms that regulate centrosome activity in Asymmetric Cell Divisions (ACD) using neural stem cells (figure) as our model, and in Symmetric Cell Divisions (SCD) using Drosophila embryos and spermatogonia as our models.
Centrosomes for Spindle Assembly
As the cornerstones of the bipolar mitotic spindle, the centrosomes ensure efficient and accurate chromosome attachment and segregation. I have had a longstanding interested in spindle assembly, with particular focus on the role of centrosomal microtubules (MTs), MT dynamics, pole focusing, and MT motors. We are interested in identifying common and unique mechanisms between symmetric and asymmetric cell divisions.
Recently, my lab has been interested in the role of Asp (Abnormal Spindle) and Cam (Calmodulin) in spindle assembly. Their role in spindle pole focusing truly highlights the cell-type diversity in tolerance to spindle assembly defects. We are currently exploring Asp as a model for microcephaly by investigating both its mitotic spindle dependent and independent roles.