Anatomy of the fungal microtubule organizing center, the spindle pole body
Introduction
Microtubule organizing centers (MTOCs) are a diverse group of membrane-less organelles that serve as sites of microtubule nucleation in eukaryotes. In animal cells, centrosomes are the primary, nuclear-.associated MTOC that form the poles of the mitotic spindle. Each centrosome is composed of a pair of centrioles, which are surrounded by a cloud of pericentriolar material from which microtubules emanate. The centriole is a cylindrical structure made of triplet microtubules organized in ninefold symmetry by a central structure known as the cartwheel. Although centrioles, like those found in centrosomes, are found throughout all kingdoms of life, including fungi, most fungi evolved a centriole-less MTOC known as the spindle pole body (SPB) [1]. While SPBs and centrosomes are morphologically distinct, both organelles share components and regulators. Analysis of fungal SPBs have identified some of the most well conserved and important MTOC components, including γ-tubulin, first identified in the filamentous fungus Aspergillus nidulans [2] (Figure 1a).
In the electron microscope (EM), SPBs appear as small, layered disc-shaped structures associated with the nuclear envelope (Figure 1b). Perhaps best characterized is the Saccharomyces cerevisiae SPB (ScSPB), which is embedded in the nuclear membrane throughout the lifecycle and contains five layers: the outer, central and inner plaques and two intermediate layers (IL1 and IL2) [3,4]. The Schizosaccharomyces pombe SPB (SpSPB) is more amorphous than its budding yeast counterpart and is only inserted in the nuclear envelope during mitosis; throughout interphase, the SpSPB is located on the cytoplasmic face of the nuclear envelope, tethered to an electron density at the inner nuclear membrane [5,6]. Adjacent to both the ScSPB and SpSPB is an electron-dense region of the nuclear envelope known as the half-bridge. The ScSPB and SpSPB have provided a foundation for analysis of SPB structure in other fungi, including yeasts and fungi of clinical and agricultural significance, many of which share a similar laminar structure. Equally important, analysis of yeast SPB structure has provided fundamental insights into the function of all MTOCs, their role in microtubule nucleation and contribution to nuclear envelope breakdown.
Section snippets
Molecular composition of the SPB core
The ScSPB contains eighteen components present in multiple copies (reviewed in Ref. [7]) (Figure 1a). The locations of proteins within the SPB have been mapped using immunoEM and more recently structured-illumination microscopy (SIM) [8,9] (Figure 1b). Molecular modeling and in vitroreconstitution have further expanded our understanding of ScSPB architecture [10•]. The ScSPB is built around a hexagonal lattice of Spc42 [11,12] (Figure 2). Drennan and colleagues reconstituted the Spc42 lattice
Microtubule nucleation complexes
Microtubule nucleation in fungi, as in other eukaryotes, is mediated by the multisubunit γ-tubulin complex (γ-TuC). The γ-TuC is connected to the SPB core by two linker proteins, Spc72/Mto1 and Spc110/Pcp1, orthologs of human CDK5RAP2 and pericentrin, respectively [25, 26, 27] (Figure 1a). Conserved C-terminal motifs target Spc72/Mto1 (MASC, for Mto1 and Spc72 C-terminus) and Spc110/Pcp1 (PACT) to the cytoplasmic and nuclear faces of the SPB, respectively, while their N-termini contain a
Association of the SPB with the nuclear membrane
Most fungi undergo a closed or semi-closed mitosis where the nuclear envelope remains intact. Integration of the SPB into the nuclear envelope is considered necessary for nucleation of spindle microtubules. In S. cerevisiae, the ScSPB is inserted into the nuclear membrane during SPB duplication, and the pole remains membrane-associated for the duration of the yeast life cycle [46] (Figure 4a). In other fungi such as S. pombe, the pole is associated with the cytoplasmic face of the nuclear
Conclusion
The diverse architecture and unique ecology of yeasts provides a framework to understand the evolution of MTOCs, including their role in mitosis, nuclear envelope remodeling and asymmetric cell division. Key insights into the mechanisms of microtubule nucleation have originated in these tractable model systems, along with their cognate of microtubule regulatory factors. The closed mitosis of fungi necessitates a coordination between nuclear and cytoplasmic microtubule formation that we are just
Conflict of interest statement
Nothing declared.
References and recommended reading
Papers of particular interest, published within the period of review, have been highlighted as:
• of special interest
•• of outstanding interest
Acknowledgements
I am grateful to Tom Giddings, Andrew Bestul and Jingjing Chen for the images and to Shelly Jones, Marisa Segal and members of the Jaspersen lab for many insightful discussions and for comments on the manuscript. Research in my lab is supported by the Stowers Institute for Medical Research and the NIH-NIGMS under award number R01GM121443.
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