Research topics in the Martin lab
Microtubule-dependent cell polarization
The cytoskeleton – microtubules and actin filaments – is essential for cell polarization. In rod-shaped fission yeast cells, microtubules are organized in a dynamic network aligned along the length of the cell and serve to transport polarity landmarks towards the extremities of the cell. Microtubules provide positional information for growth at cell extremities: cells with anomalies in their microtubule network grow at ectopic locations. We had previously demonstrated that one landmark transported by microtubules, the protein Tea4, binds an actin nucleator of the formin family, For3. This suggested a model in which positional information provided by microtubules directly links to actin assembly to promote polarized cell growth at cell poles. However, Tea4 function extends beyond formin recruitment. We are interested in further understanding how microtubules through Tea4 promote mark sites for polarized cell growth.
Actin cables and vesicle transport
The actin cytoskeleton plays fundamental roles in all cells for cell morphogenesis. In yeast cells, three main actin structures have been described: the cytokinetic actin ring, necessary for cell division, actin patches, which represent sites of endocytosis, and actin cables, long bundles of largely parallel linear actin filaments. Actin cables are assembled by For3, a member of the formin family of actin nucleators. These cables serve as tracks for myosin V-dependent transport of cargoes towards sites of polarized cell growth. Cargoes include membrane material and cell wall remodeling components essential for polarized cell growth. We have previously shown that actin cables undergo retrograde actin flow, where actin polymerization takes place at cell poles, where For3 is localized, and pushes the cable into the cell interior. For3 undergoes similar flow, detaching from cell poles and traveling with the cable into the cell interior. We are interested in defining how actin cables organize in the cell to efficiently transport cargoes towards cell poles. We have recently shown that type V myosins contribute to the organization of actin cables.
Surprisingly, myosin V transport and actin cables, while important for polarized cell growth, are not essential for it. We have recently shown that fission yeast cells also use a second polarization strategy. Secretion of cell wall-remodelling components at cell poles requires the exocyst complex, an eight-subunit complex that facilitates fusion of vesicles with the plasma membrane. Localization of this complex to cell poles is independent of the cytoskeleton. Disruption of actin cables or of the exocyst does not block polarized growth, but double disruption prevents polarized growth. For polar growth at cell poles, fission yeast thus rely both on the actin cables that transport vesicle cargoes and on the exocyst complex that promotes fusion of these vesicles with the plasma membrane.
While wildtype fission yeast cells transport vesicles towards cell tips along actin cables, we recently showed that this transport can be re-routed along microtubules to achieve very similar cell shape. To this aim, we engineered a chimeric motor protein between a kinesin motor domain and the cargo-binding region of the major type V myosin Myo52. This chimeric motor transports myosin cargos along microtubules and restores viability and elongated shape to cells lacking actin cables. Thus, cells are plastic enough to permit this re-wiring. This also demonstrates that vesicular transport to cell poles is sufficient for polarized cell growth, whatever route is used. The video-abstract below describes this study.
Coordination of cell growth and division
Cell cycle progression is monitored by checkpoints that ensure the fidelity of cell division and prevent unrestricted cell proliferation. Checkpoints also serve to couple cell size with division – a mechanism important to adapt to changing environmental conditions. While most studies on cell size homeostasis have focused on the links between size and biosynthetic activity, we have recently discovered a novel geometry-sensing mechanism by which fission yeast cells couple cell length with entry into mitosis. Conceptually, the system is remarkably simple: it is composed of a signal – the protein kinase Pom1 – forming concentration gradients from the ends of the cells, which inhibits a sensor – the protein kinase Cdr2, itself an activator of mitotic entry that inhibit the Wee1 kinase – placed at the cell equator. Since Pom1 concentration at the cell middle is higher in short cells than in long cells, this suggests a model where Pom1 inhibits Cdr2 until the cell has reached a sufficient length.
We have elucidated the molecular mechanism by which the gradients of Pom1 kinase form from cell poles. Pom1 associates with the plasma membrane through a basic region. Through autophosphorylation, which targets in particular in this basic region, membrane affinity is lowered. Local dephosphorylation of Pom1, which promotes membrane association, occurs at cell poles and depends on Tea4, which targets the type I phosphatase, Dis2, to this location. Thus, Pom1 gradients at the plasma membrane are simply organized by a local dephosphorylation event that drives Pom1 membrane association at cell poles. From there Pom1 likely diffuses at the plasma membrane and undergoes autophosphorylation reaction to drive membrane detachment.
We are further exploring the regulation of the Pom1-Cdr2 cell size homeostasis pathway both under normal growth conditions and in conditions of stress.
Cell polarization during mating
Yeast cells show prominent polarization during the mating process, when two cells of opposite mating type extend cellular projections towards each other. Fission yeast cells exist as homothallic strains, in which cells can switch mating type after division, yielding sister-cells of opposite mating types. We are interested in exploring the mechanisms by which cells re-orient their polarization machinery towards a mating partner.
We have shown that, in the early stages of the mating process before cells have selected a mating partner, or upon exposure to low-level pheromone levels, fission yeast cells display a dynamic site of polarization. The small GTPase Cdc42, a central eukaryotic regulator of cell polarization, forms clusters that appear and disappear dynamically at the plasma membrane. Direct effectors of Cdc42, such as formin and exocyst subunits, colocalize to these dynamic clusters, but not cell wall synthases. Consistently, cells do not grow during this phase. When cells experiment high-level pheromone signalling, they bypass this dynamic polarization phase, fail to orient growth towards neighbouring mating partners and mate preferentially to sister-cells. We are interested in defining the molecular mechanisms behind this dynamic polarization behaviour and partner choice. We also explore the cytoskeletal regulation of cell-cell fusion to produce a diploid progeny.