Molecular basis of asymmetric cell division
c/o IFOM-IEO Campus
Via Adamello, 16 - 20139 Milan, Italy
We are interested in the molecular pathways governing asymmetric cell divisions, with emphasis on the role of the mitotic spindle orientation in determining the daughter cells' fate.
The proper execution of asymmetric divisions is crucial in generating tissue diversity during development, as well as for tissue homeostasis and regeneration in adult organisms. An increasing body of literature supports the notion that certain human cancers arise from abnormalities in adult stem cells asymmetric divisions able to alter the daughter cells' fate and lead to over-proliferation (the so called cancer stem cell hypothesis). Such failures in asymmetric divisions have to be expected if the pathways controlling the position of the cytokinesis plane are compromised. Indeed impairment of proteins involved in spindle alignment results in incorrect sibling fate and misproliferation during mammalian neurogenesis and skin development. Furthermore, genetic lesions affecting the spindle alignment in vertebrate gut epithelia correlate with colon cancer development.
Figure I. Relationship between the mitotic spindle orientation and the asymmetric outcome of a cell division. [+zoom]
To make a cell division asymmetric, the position of the mitotic spindle has to be tightly coordinated with the cortical polarity, so that daughter cells will be properly positioned within the tissue, will inherit unequal sets of fate determinants and follow differential fates (Figure I). This observation sets the stage for our studies, aimed at gaining insight into the structural and functional organization of the molecular machines responsible for spindle coupling to polarity cues during stem cells asymmetric divisions. To address this biological problem, we use a combination of high-resolution X-ray crystallography studies, biochemical analyses on reconstituted protein complexes and cell biology. Using the detailed molecular information delivered by our structural studies, we formulate precise models of how intrinsic properties of individual protein relate to the behavior of the mitotic spindle during asymmetric cell divisions, that we challenge in living cells. An emerging concept in the cancer field is that cancer stem cells may be responsible for relapse and resistance to anticancer therapies. In this view, a clear molecular description of processes underlying asymmetric cell divisions will be instrumental in identifying new stem-cell specific drug targets for therapeutical intervention.
Our activity is organized in two main research lines:
1. Structural and functional characterization of cortical force generators
Figure 1. Organizational principles of cortical force generators. [+zoom]
Cortical force generators are molecular motors orchestrating the correct placement of the mitotic spindle within the cell. They perform different tasks: 1) they localize at well defined spatially-restricted cortical sites in conjunction with polarity cues; 2) they capture astral microtubules emanating form the spindle pole; 3) they generate microtubule pulling forces whose resultant is responsible for the final position of the mitotic spindle (Figure 1). The core components of force generators and the non-canonical G-protein signaling pathway involved in their regulation are evolutionary conserved from nematode to mammals. Their central module consist of heterotrimeric NuMA/LGN/Gαi complexes, assembled on the GDP-loaded Gαi species. From a topological point of view, LGN has been depicted as the molecular link between Gαi subunits anchored at the plasma membrane and the microtubule associated protein NuMA. Intriguingly, in the apo form LGN works as a switch, held together by head-to-tail interactions. We are interested in understanding the molecular events triggering the LGN conformational transition required to assemble the NuMA/LGN/Gαi complexes, and in determining the architecture of the reconstituted trimer. A research interest intimately connected with the organizational principles underlying force generator assembly is their regulation by the Gαi GTP-cycle, controlled by the GEF RIC-8A. We study the recognition principles and the functional impact of the RIC-8A GEF activity in coordinating the cortical force generators activity with mitotic progression in space and time.
2. Molecular characterization of the interplay between polarity and cell division plane
Our second research line deals with the issue of how force generators are specifically recruited at sites of polarization. In several cellular systems, cortical polarization is established by asymmetrical distribution of the evolutionary conserved Par3/Par6/aPKC complex, which in turn defines the asymmetrical localization of fate determinants. How the mitotic spindle is coupled to the cellular polarity depends on the specific cellular system. During asymmetric divisions of Drosophila melanogaster neuroblasts, Par3/Par6/aPKC complexes localize at the apical site, and from here they recruit force generators via an adaptor named Inscuteable (Figure 2A).
Figure 2. Coupling of spindle orientation to cellular polarity in asymmetrically dividing cells and epithelia. [+zoom]Converging evidence supports the view that a similar pathway governs asymmetric divisions also during mammalian skin differentiation and neurogenesis. To shed light on the fundamental aspect of how cortical polarity drives the assembly of force generators at the correct sites, we study the interaction of LGN with Inscuteable and Par3, and its interplay with the NuMA/LGN/Gαi network. An emerging concept in the field of stem cells is that differential cues instruct the position of the cytokinesis plane (Figure 2B). To unveil the molecular network coupling the force generators to the cellular polarity in different environments, we are pursuing the identification of new tissue specific interactors of NuMA and LGN.