Inhibition of Ras activity coordinates cell fusion with cell–cell contact during yeast mating

Abstract

The yeast cell wall is digested to allow cell fusion during sexual reproduction. How cells coordinate this process with cell–cell contact to prevent lysis is unclear. Merlini et al. show that the Ras GAP protein Gap1, which is recruited to sites of Ras-GTP, restricts Ras activity and protects cells from lysis due to premature fusion attempts.

Differential GAP requirement for Cdc42-GTP polarization during proliferation and sexual reproduction

“…This phenotype, similar to expression of GTP-locked Cdc42, is consistent with the idea that spatial restriction of Cdc42 activity is a dynamic process that relies not only on local activation at cell tips, but also on GTP hydrolysis to counteract Cdc42 diffusion. An essentially identical outcome is observed for Ras1 GTPase normally selectively active at cell poles: upon deletion of Ras1 only GAP, the GTPase is now active around the entire cell cortex (Merlini et al, 2018). By contrast, cells lacking all three GAPs were still almost fully mating-competent and retained the ability to polarize active Cdc42, as labeled by the CRIB reporter or the Cdc42-GTP-binding Scd2 scaffold.…”

“…Indeed, the intrinsic GTPase activity of Cdc42 we observed in vitro ( Fig. 2C; Cdc42-GTP t 1/2 < 5 min) is relatively high compared to that of Cdc42 in other species (Zhang et al, 1999) or Ras1 in S. pombe (Merlini et al, 2018), and may thus suffice to be effective against an estimated lateral diffusion of Cdc42-GTP of 2µm 2 /min .…”

“…However, local activation is not sufficient to generate locally restricted Cdc42 activity: cells also need to inactivate Cdc42 to prevent the spread of the active form. For instance, we recently observed that the cell pole-restricted activity of the related small GTPase Ras1 requires inactivation by its GAP, the deletion of which leads to loss of spatial information and Ras1-GTP distribution over the whole plasma membrane (Merlini et al, 2018). In fission yeast, two Cdc42 GAPs, Rga4 and Rga6, both of which localize to cell sides, contribute to the spatial restriction of active Cdc42 to the cell tips, with rga4∆ and rga6∆ mutant cells exhibiting enlarged cell width (Revilla-Guarinos et al, 2016;Tatebe et al, 2008).…”

Optogenetics reveals Cdc42 local activation by scaffold-mediated positive feedback and Ras GTPase

“…In a similar manner, Scd1 overexpression causes cell rounding of gap1∆ mutants but not wildtype cells (unpublished data), indicating that cells lacking spatial Ras1dependent cues become sensitive to perturbations in feedback control. Ras1 modulation of the positive feedback might be particularly relevant during sexual reproduction for partner selection and mating, when cells abolish tip growth and pheromone signaling positions active Ras1 (Merlini et al, 2016;Merlini et al, 2018). We propose that by superimposing positive feedback and Ras1-dependent regulation, cells buffer fluctuations of individual regulators to robustly define and position zones of Cdc42 activity.…”

“…Notably, scd2 deletion causes cell rounding (Chang et al, 1994;Kelly and Nurse, 2011b) but does not severely affect Cdc42 activity, in contrast to scd1∆ , implying the existence of alternative mechanisms for Cdc42 activation. Several pieces of data implicate the small GTPase Ras1 as a Cdc42 regulator upstream of Scd1 (Chang et al, 1994;Merlini et al, 2018;Weston et al, 2013): Ras1 binds Scd1 directly (Chang et al, 1994), is localized to the plasma membrane and activated at the cell ends like Cdc42 , and its deletion causes partial cell rounding (Fukui et al, 1986).…”