A stable microtubule bundle formed through an orchestrated multistep process controls quiescence exit.

Cells fine-tune microtubule assembly in both space and time to give rise to distinct edifices with specific cellular functions. In proliferating cells, microtubules are highly dynamics, and proliferation cessation often leads to their stabilization. One of the most stable microtubule structures identified to date is the nuclear bundle assembled in quiescent yeast. In this article, we characterize the original multistep process driving the assembly of this structure. This Aurora B-dependent mechanism follows a precise temporality that relies on the sequential actions of kinesin-14, kinesin-5, and involves both microtubule-kinetochore and kinetochore-kinetochore interactions. Upon quiescence exit, the microtubule bundle is disassembled via a cooperative process involving kinesin-8 and its full disassembly is required prior to cells re-entry into proliferation. Overall, our study provides the first description, at the molecular scale, of the entire life cycle of a stable microtubule structure in vivo and sheds light on its physiological function.

A new partial loss of function allele of rad-54.L.

RAD-54.L is required for the repair of meiotic double-strand DNA breaks (DSBs), playing an essential role in promoting removal of recombinase RAD-51 and normal completion of meiotic recombination. Failure to complete meiotic DSB repair leads to 100% lethality of embryos produced by rad-54.L null mutant mothers. Here we report a new partial loss of function allele, rad-54.L(me139) , that may prove useful for investigating meiotic mechanisms by providing a sensitized genetic background that reduces but does not eliminate the essential functions of RAD-54.L.

Tubulin isotypes - functional insights from model organisms.

The microtubule cytoskeleton is assembled from the α- and ß-tubulin subunits of the canonical tubulin heterodimer, which polymerizes into microtubules, and a small number of other family members, such as γ-tubulin, with specialized functions. Overall, microtubule function involves the collective action of multiple α- and ß-tubulin isotypes. However, despite 40 years of awareness that most eukaryotes harbor multiple tubulin isotypes, their role in the microtubule cytoskeleton has remained relatively unclear. Various model organisms offer specific advantages for gaining insight into the role of tubulin isotypes. Whereas simple unicellular organisms such as yeast provide experimental tractability that can facilitate deeper access to mechanistic details, more complex organisms, such as the fruit fly, nematode and mouse, can be used to discern potential specialized functions of tissue- and structure-specific isotypes. Here, we review the role of α- and ß-tubulin isotypes in microtubule function and in associated tubulinopathies with an emphasis on the advances gained using model organisms. Overall, we argue that studying tubulin isotypes in a range of organisms can reveal the fundamental mechanisms by which they mediate microtubule function. It will also provide valuable perspectives on how these mechanisms underlie the functional and biological diversity of the cytoskeleton.

Tubulin isotypes optimize distinct spindle positioning mechanisms during yeast mitosis.

Microtubules are dynamic cytoskeleton filaments that are essential for a wide range of cellular processes. They are polymerized from tubulin, a heterodimer of α- and ß-subunits. Most eukaryotic organisms express multiple isotypes of α- and ß-tubulin, yet their functional relevance in any organism remains largely obscure. The two α-tubulin isotypes in budding yeast, Tub1 and Tub3, are proposed to be functionally interchangeable, yet their individual functions have not been rigorously interrogated. Here, we develop otherwise isogenic yeast strains expressing single tubulin isotypes at levels comparable to total tubulin in WT cells. Using genome-wide screening, we uncover unique interactions between the isotypes and the two major mitotic spindle positioning mechanisms. We further exploit these cells to demonstrate that Tub1 and Tub3 optimize spindle positioning by differentially recruiting key components of the Dyn1- and Kar9-dependent mechanisms, respectively. Our results provide novel mechanistic insights into how tubulin isotypes allow highly conserved microtubules to function in diverse cellular processes.

Discrete regions of the kinesin-8 Kip3 tail differentially mediate astral microtubule stability and spindle disassembly.

To function in diverse cellular processes, the dynamic properties of microtubules must be tightly regulated. Cellular microtubules are influenced by a multitude of regulatory proteins, but how their activities are spatiotemporally coordinated within the cell, or on specific microtubules, remains mostly obscure. The conserved kinesin-8 motor proteins are important microtubule regulators, and family members from diverse species combine directed motility with the ability to modify microtubule dynamics. Yet how kinesin-8 activities are appropriately deployed in the cellular context is largely unknown. Here we reveal the importance of the nonmotor tail in differentially controlling the physiological functions of the budding yeast kinesin-8, Kip3. We demonstrate that the tailless Kip3 motor domain adequately governs microtubule dynamics at the bud tip to allow spindle positioning in early mitosis. Notably, discrete regions of the tail mediate specific functions of Kip3 on astral and spindle microtubules. The region proximal to the motor domain operates to spatially regulate astral microtubule stability, while the distal tail serves a previously unrecognized role to control the timing of mitotic spindle disassembly. These findings provide insights into how nonmotor tail domains differentially control kinesin functions in cells and the mechanisms that spatiotemporally control the stability of cellular microtubules.