Identification and analysis of Tef1p, a novel Rho1p-interacting protein
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Abstract
Rho GTPases act as molecular switches, occupying an active GTP or an inactive GDP-bound state. Rho1p, one member of the Rho family GTPases, has a role in the late stages of vacuole fusion through an unknown mechanism. We identified a ~50kDa Rho1p-interacting protein as Tef1p, a dual function GTPase with known roles in both protein translation and actin cytoskeletal organization. Tef1p is an aminoacyl-tRNA transferase in protein translation where GTP-bound Tef1p shuttles aminoacyl-charged tRNAs to ribosomes where they are transferred to the elongating peptide chain. Tef1p also binds actin filaments and organizes them into higher order cable structures such as stress fibers. We used nucleotide-state specific affinity pulldown methods to determine that Tef1p specifically interacts with Rho1p, but not as a downstream effector. The Tef1p::Rho1p interaction was calcium- dependent, suggesting that the interaction occurs late in membrane fusion. Our data suggest that Tef1p modulates Rho1p function, perhaps via complex formation.