Orderly assembly underpinning built-in asymmetry in the yeast centrosome duplication cycle requires cyclin-dependent kinase

Abstract

Asymmetric astral microtubule organization drives the polarized orientation of the S. cerevisiae mitotic spindle and primes the invariant inheritance of the old spindle pole body (SPB, the yeast centrosome) by the bud. This model has anticipated analogous centrosome asymmetries featured in self-renewing stem cell divisions. We previously implicated Spc72, the cytoplasmic receptor for the gamma-tubulin nucleation complex, as the most upstream determinant linking SPB age, functional asymmetry and fate. Here we used structured illumination microscopy and biochemical analysis to explore the asymmetric landscape of nucleation sites inherently built into the spindle pathway and under the control of cyclin-dependent kinase (CDK). We show that CDK enforces Spc72 asymmetric docking by phosphorylating Nud1/centriolin. Furthermore, CDK-imposed order in the construction of the new SPB promotes the correct balance of nucleation sites between the nuclear and cytoplasmic faces of the SPB. Together these contributions by CDK inherently link correct SPB morphogenesis, age and fate.

Ultrastructure expansion microscopy reveals the cellular architecture of budding and fission yeast

“…Like its metazoan counterpart, the SPB is duplicated once per cell cycle and has microtubule nucleation abilities ( Kilmartin, 2014 ). Our understanding of the S. cerevisiae SPB is grounded from electron microscopy (EM) ( Kilmartin, 2014 ; O'Toole et al, 1999 ) and has been beautifully complemented with live imaging and more recently SIM studies ( Burns et al, 2015 ; Geymonat et al, 2020 ; Unruh et al, 2018 ), as well as biochemical data ( Ruthnick et al, 2021 ). The SPB is embedded in the nuclear membrane as a feature of the yeast closed mitosis and is a cylindrical multi-layered organelle composed of outer, central and inner plaques ( Jaspersen and Winey, 2004 ) ( Fig.…”

Spindle pole body-associated Atg11, an autophagy-related protein, regulates microtubule dynamics essential for high-fidelity chromosome segregation

“…Lee et al, 2000; Miller, Cheng, & Rose, 2000; Segal & Bloom, 2001; Yeh et al, 1995). Two different premises suggest that the aMT asymmetry at the SPBs stems from the contribution of extrinsic and intrinsic factors (Geymonat et al, 2020). Kar9, present at both poles during the onset of SPB separation, represents the extrinsic control of the aMT asymmetry (Lengefeld & Barral, 2018).…”

“…On the other hand, the cyclin-dependent kinase (CDK) Cdc28/Cdk1, representing the intrinsic factor, facilitates the biased recruitment of SPB components such as Spc72 and the γ- tubulin complex (γ-TC) at the old SPB (dSPB) during the early cell cycle. Subsequently, this facilitates the assembly of the new SPB (mSPB) in the correct order with the recruitment of the outer plaque following the complete assembly of the inner plaque (Geymonat et al, 2020). Such asymmetry leads to the nucleation of aMTs from the dSPB necessary for the mitotic spindle positioning along the mother-bud axis (Markus, Kalutkiewicz, & Lee, 2012; Palmer, Sullivan, Huffaker, & Koshland, 1992; Shaw, Yeh, Maddox, Salmon, & Bloom, 1997; Sullivan & Huffaker, 1992).…”

“…In budding yeast, the spatial crosstalk between asymmetric nucleation of aMTs and cortical cues regulating the mitotic spindle alignment along the polarity axis requires the action of two functionally redundant pathways (Korinek, Copeland, Chaudhuri, & Chant, 2000; L. Lee et al, 2000;Miller, Cheng, & Rose, 2000;Segal & Bloom, 2001;Yeh et al, 1995). Two different premises suggest that the aMT asymmetry at the SPBs stems from the contribution of extrinsic and intrinsic factors (Geymonat et al, 2020). Kar9, present at both poles during the onset of SPB separation, represents the extrinsic control of the aMT asymmetry (Lengefeld & Barral, 2018).…”

“…On the other hand, the cyclin-dependent kinase (CDK) Cdc28/Cdk1, representing the intrinsic factor, facilitates the biased recruitment of SPB components such as Spc72 and the γtubulin complex (γ-TC) at the old SPB (dSPB) during the early cell cycle. Subsequently, this facilitates the assembly of the new SPB (mSPB) in the correct order with the recruitment of the outer plaque following the complete assembly of the inner plaque (Geymonat et al, 2020).…”