Understanding the intricacies of telomerase regulation is crucial due to the potential health benefits of modifying its activity. Telomerase is composed of an RNA component and reverse transcriptase. However, additional factors required during biogenesis vary between species. Here we have identified fission yeast Lar7 as a member of the conserved LARP7 family, which includes the Tetrahymena telomerase-binding protein p65 and human LARP7. We show that Lar7 has conserved RNA-recognition motifs, which bind telomerase RNA to protect it from exosomal degradation. In addition, Lar7 is required to stabilise the association of telomerase RNA with the protective complex LSm2–8, and telomerase reverse transcriptase. Lar7 remains a component of the mature telomerase complex and is required for telomerase localisation to the telomere. Collectively, we demonstrate that Lar7 is a crucial player in fission yeast telomerase biogenesis, similarly to p65 in Tetrahymena, and highlight the LARP7 family as a conserved factor in telomere maintenance.

High-fidelity chromosome segregation relies on proper microtubule regulation. Kinesin-8 has been shown to destabilise microtubules to reduce metaphase spindle length and chromosome movements in multiple species. XMAP215/chTOG polymerases catalyse microtubule growth for spindle assembly, elongation and kinetochore-microtubule attachment. Understanding of their biochemical activity has advanced, but little work directly addresses the functionality and interplay of these conserved factors. We utilised the synthetic lethality of fission yeast kinesin-8 (Klp5-Klp6) and XMAP215/chTOG (Dis1) to study their individual and overlapping roles. We found that the non-motor kinesin-8 tailbox is essential for mitotic function; mutation compromises plus-end-directed processivity. Klp5-Klp6 induces catastrophes to control microtubule length and, surprisingly, Dis1 collaborates with kinesin-8 to slow spindle elongation. Together, they enforce a maximum spindle length for a viable metaphase–anaphase transition and limit elongation during anaphase A to prevent lagging chromatids. Our work provides mechanistic insight into how kinesin-8 negatively regulates microtubules and how this functionally overlaps with Dis1 and highlights the importance of spindle length control in mitosis.

Proper bipolar spindle assembly underlies accurate chromosome segregation. A cohort of 13 microtubule-associated proteins orchestrates spindle microtubule formation in a spatiotemporally 14 coordinated manner. Among them, the conserved XMAP215/TOG family of microtubule 15 polymerase plays a central role in spindle assembly. In fission yeast, two XMAP215/TOG members, 16Alp14 and Dis1, share essential roles in cell viability; however how these two proteins functionally 17 collaborate remains undetermined. Here we show the functional interplay and specification of 18 Alp14 and Dis1. Creation of new mutant alleles of alp14, which display temperature sensitivity in 19 the absence of Dis1, enabled us to conduct detailed analyses of a double mutant. We have found 20 that simultaneous inactivation of Alp14 and Dis1 results in early mitotic arrest with very short, 21 fragile spindles. Intriguingly, these cells often undergo spindle collapse, leading to a lethal "cut" 22 phenotype. By implementing an artificial targetting system, we have shown that Alp14 and Dis1 are 23 not functionally exchangeable and as such are not merely redundant paralogues. Intriguingly, while 24 Alp14 promotes microtubule nucleation, Dis1 does not. Our results uncover that the intrinsic 25 specification, not the spatial regulation, between Alp14 and Dis1 underlies the collaborative actions 26 of these two XMAP215/TOG members in mitotic progression, spindle integrity and genome 27 stability. 28 Keywords: Fission yeast; microtubule polymerase; XMAP215/TOG; mitotic spindle; spindle pole 29 body; kinetochore 30 31 progression, development and differentiation pathways [1]. During mitosis, spindle MTs assemble 34 to form a bipolar structure that emanates from the two spindle poles. The bipolar spindle functions35as the division apparatus for sister chromatids, generating pulling forces to move them towards 36 opposite poles to ensure equal partition of genetic material. Errors in this process can lead to cell 37 death and/or aneuploidy, a major risk factor for miscarriage, birth defects and tumourigenesis [2, 3]. 38MTs are intrinsically dynamic, a characteristic termed dynamic instability [4, 5]. In a living cell, 39 a cohort of proteins, collectively known as microtubule-associated proteins (MAPs), participate in 40 spindle assembly, stability and maintenance as structural and/or regulatory factors. A conserved 41 family of MAPs, here referred to as the XMAP215/TOG family, is deeply rooted in the eukaryotic 42 branch of the tree of life and arguably the most important regulator in MT organisation [6-9]. Int. J. Mol. Sci. 2019, 20, x FOR PEER REVIEW 2 of 16 [10], homologues have been identified in virtually all eukaryotic species [9]. The seminal 45 advancement of our understanding of the XMAP215/TOG family was the discovery that XMAP215 46 is a MT polymerase [11]; this protein is capable of incorporating α/β-heterodimers at the plus end of 47 pre-existing MTs. Since then, this activity has been proven for other members of the family from a 48 diverse ra...