Ras and the Rho Effector Cla4 Collaborate to Target and Anchor Lte1 at the Bud Cortex

Abstract

Previously published online as a Cell Cycle E-publication: http://www.landesbioscience.com/journals/cc/abstract.php?id=1785 KEY WORDS SUPPLEMENTARY MATERIALSupplementary Figures 1S and 2S can be found at http://www.landesbioscience.com/seshanCC4-7-sup.pdf. Report Ras and the Rho Effector Cla4 Collaborate to Target and Anchor Lte1 at the Bud Cortex ABSTRACTLte1, a protein important for exit from mitosis, localizes to the bud cortex as soon as the bud forms and remains there until cells exit from mitosis. Ras, the Rho GTPase Cdc42 and its effector, the protein kinase Cla4, are required for Lte1's association with the bud cortex. Here we investigate how Ras and the Cdc42 effector Cla4 regulate the localization of Lte1. We find that Ras2 and Lte1 associate in stages of the cell cycle when Lte1 is phosphorylated and associated with the bud cortex and that this association requires CLA4. Additionally, RAS1 and RAS2 are required for CLA4-dependent Lte1 phosphorylation. Our findings suggest that Cla4-dependent phosphorylation promotes the initial association of Lte1 with Ras at the bud cortex and that Ras is required to stabilize phosphorylated forms of Lte1 at the bud cortex. Our results also raise the interesting possibility that the localization of Lte1 affects the protein's ability to promote mitotic exit.

p21-Activated Kinases Cla4 and Ste20 Regulate Vacuole Inheritance in Saccharomyces cerevisiae

“…6C and D), we hypothesized that Cla4 may act indirectly to affect Vac17 protein levels. Cla4 plays a role in the proper localization and activation of the guanine nucleotide exchange factor Lte1 in the bud (37,38,66,67,81,88). Asynchronous cultures of wild-type and GAL-CLA4 lte1 cells were grown to mid-log phase, 2% galactose was added at the zero time point, and samples were taken every hour for examination of Vac17-ProA levels.…”

A Role For Lte1p (a Low Temperature Essential Protein Involved in Mitosis) in Proprotein Processing in the Yeast Secretory Pathway

“…It is debated whether the CDC25 homology (GEF) domain contained in the C-terminal region of Lte1p is normally required for its mitotic function (10) although this is a currently favored view (11). TEM1 was originally discovered in a screen for high copy suppressor genes that rescue cold-sensitive growth arrest of lte1 cells (8).…”

“…The timing of Tem1p activation by Lte1p occurs when the spindle pole body migrates into the bud. Lte1p localization at the bud appears to be regulated by its interactions with Ras2p (10), its phosphorylation state (11), and interactions with cell polarity proteins (14). However, Lte1p function during S-phase is largely unknown.…”

“…Lte1p has a large, modular-appearing primary structure including small GEF homology domains embedded in the C-and N-terminal regions (9). Overexpression of lte1-mini (truncated to lack these domains) can suppress the cold-sensitive growth arrest of lte1D cells, raising the possibility that Lte1p might not be a GEF for Tem1p (10); however, other analysis supports the hypothesis that GEF function of Lte1p activates Tem1p (11). Importantly, while Tem1p expression becomes significant only during mitosis (telophase), Lte1p is stably expressed throughout the cell cycle (12), suggesting other possible roles for Lte1p in interphase cells.…”

Interpreting spatial information and regulating mitosis in response to spindle orientation: Figure 1.

“…Localization of Lte1 to the daughter cell cortex is dependent on the small GTPase proteins Cdc42 and Ras in addition to other polarity proteins, Cla4 and Kel1, but precisely how they work is not understood (Seshan et al 2002;Yoshida et al 2003;Seshan and Amon 2005). Maintaining Lte1 in the daughter cell requires the septins, structural proteins that are key components of the bud neck.…”