Associations among PH and SH3 domain-containing proteins and Rho-type GTPases in Yeast.

Abstract

Abstract. The src homology region 3 (SH3) domainbearing protein Bemlp and the Rho-type GTPase Cdc42p are important for bud emergence in Saccharomyces cerevisiae. Here, we present evidence that through its second SH3 domain, Bemlp binds to the structurally and functionally similar proteins Boilp and Boi2p, each of which contain an SH3 and a pleckstrin homology (PH) domain. Deletion of BOll and BOI2 together leads to impaired morphogenesis and poor viability. A PH domain-bearing segment of Boilp that lacks the Bemlp-binding site is necessary and sufficient for function. This segment of Boilp displays a twohybrid interaction with Cdc42p, suggesting that Boilp either binds directly to or is part of a larger complex that contains Cdc42p. Consistent with these possibilities, overexpression of Boilp inhibits bud emergence, but this inhibition is counteracted by cooverexpression of Cdc42p. Increased expression of the Rho-type GTPase Rho3p, which is implicated in bud growth, suppresses the growth defects of boil boi2 mutants, suggesting that Boilp and Boi2p may also play roles in the activation or function of Rho3p. These findings provide an example of a tight coupling in function between PH domain-bearing proteins and both Rho-type GTPases and SH3 domain-containing proteins, and they raise the possibility that Boilp and Boi2p play a role in linking the actions of Cdc42p and Rho3p.T HE src homology region 3 (SH3) 1 and pleckstrin homology (PH) domains are present in many proteins that are involved in signal transduction and the organization of the cortical cytoskeleton (Pawson, 1995). The binding sites for SH3 domains generally appear to be short, proline-rich sequences . Less is known about the binding sites for PH domains. The findings that the PH domains in some proteins overlap with sequences that can bind to the 13y subunit of some trimeric G proteins (Touhara et al., 1994) and that some PH domains can bind PIP2 (Harlan et al., 1994), however, raise the possibility that a general role of PH domains might be to target proteins to membranes.Rho-type GTPases also are involved in signal transduction and the organization of the cortical cytoskeleton

A Novel Cellular Protein (MTBP) Binds to MDM2 and Induces a G1 Arrest That Is Suppressed by MDM2

“…binding protein). Data base analysis detected two yeast genes whose protein products possessed significant homology to MTBP: BOI1 and BOI2 (36,37). The two proteins encoded by these genes (Boi1p and Boi2p, respectively) exhibit an overall amino acid identity of 38%, but this is concentrated into four regions (I-IV) that possess identities of 71, 65, 78, and 69%, respectively.…”

“…MTBP Inhibits Cell Growth-Several MDM2-binding proteins are regulators of cell growth; and indeed, both of the MTBP partial homologs (Boi1p and Boi2p) have been shown to have growth inhibitory activity (36,37). Therefore, we investigated the effect of MTBP expression on cell growth in culture.…”

Yeast and fungal morphogenesis from an evolutionary perspective

“…Through multiple protein interaction domains, these proteins may facilitate transient protein complex formation but are unlikely to act as static reference site for the localization of the bound proteins. One example is Bem1, which has binding sites for a number of proteins involved in cell polarization, including Cdc42 GTP , Cdc24, Cla4 and Boi1,2 proteins [67,118]. Whereas Bem1 may engage in hetero-oligomeric interactions, it has been shown to rapidly cycle on and off the plasma membrane and is therefore unlikely to provide any significant anchorage for its binding partners [69].…”

Evidence for physical and functional interactions among two Saccharomyces cerevisiae SH3 domain proteins, an adenylyl cyclase-associated protein and the actin cytoskeleton.

“…Vertebrate actin-associated proteins containing SH3 domains include proteins with structural roles such as fodrin and nebulin (Merilainen et al, 1993;Wang et al, 1996), unconventional myosins (Bement et al, 1994;Stoffler et al, 1995;Goodson et al, 1996), kinases and associated molecules (Flynn et al, 1993;Turner and Miller, 1994;Hildebrand et al, 1996), and other actin-binding proteins such as cortactin (Wu and Parsons, 1993). In S. cerevisiae, SH3-containing proteins include the actin-associated proteins Abplp (Drubin et al, 1990), Slalp (Holtzman et al, 1993), and Rvs167p (Bauer et al, 1993), as well as Bemlp and the recently identified Boilp and Boi2p, which have roles in the maintenance of normal cellular and cytoskeletal morphology (Chenevert et al, 1992;Bender et al, 1996;Matsui et al, 1996). A number of yeast proteins implicated in cytoskeletal function, including Abplp, Slalp, Boilp, Boi2p, and Srv2p (Field et al, 1990), contain proline-rich regions which include amino acid sequences similar to known binding sites for SH3 domains.…”