SGT2 and MDY2 interact with molecular chaperone YDJ1 in Saccharomyces cerevisiae

Abstract

In Saccharomyces cerevisiae, Sgt2 was thought to be the homologue of vertebrate SGT (small glutamine tetratricopeptide repeat-containing protein). SGT has been known to interact with both Hsp70 and Hsp90. However, it was not clear whether Sgt2 might have a similar capacity. Here, we showed that Ssa1/Ssa2 (yeast heat shock cognate [Hsc]70), Hsc82 (yeast Hsp90), and Hsp104 coprecipitated with Sgt2 from yeast lysates. Another molecular chaperone, Ydj1, known to interact with Ssa1 and Hsc82, also coprecipitated with Sgt2. Synthetic lethality between SGT2 and YDJ1 was observed after the cells were under stress, although Sgt2 might not interact physically with Ydj1. We also found that Mdy2 interacted with the N-terminal region of Sgt2 and that Mdy2 appeared to interact physically with Ydj1. Mdy2 therefore may mediate the association of Ydj1 and Sgt2. In addition, the mating efficiency of mdy2⌬, sgt2⌬, and mdy2⌬sgt2⌬ strains was reduced to a similar extent. Compared with mdy2⌬ and ydj1⌬ cells, ydj1⌬mdy2⌬ cells, however, showed a further suppression in mating efficiency. Moreover, MDY2 interacted genetically with YDJ1. These results suggest that protein complexes containing Sgt2 and Mdy2 bring molecular chaperones together to carry out certain chaperoning functions.

A Structural Model of the Sgt2 Protein and Its Interactions with Chaperones and the Get4/Get5 Complex

“…The Sgt2 Dimer Binds a Single Copy of Get5 at a Canonical Ubl Interface-Yeast two-hybrid assays (7,14) and analytical SEC (5) indicated that the N terminus of Sgt2 and the Ubl domain of Get5 are necessary for binding between these two proteins; however, molecular details of this interaction remain to be defined. We used a nickel affinity pulldown assay to further characterize the complex.…”

“…Additional yeast two-hybrid and pulldown assays demonstrated that an N-terminal construct of Sgt2 was necessary and sufficient for binding to Get5, and consequently Get4, in an interaction also dependent upon the Ubl domain of Get5 (7,14). A direct role for Sgt2 in the TA targeting pathway was shown by two independent genetic interaction analyses indicating a strong functional connection with other GET pathway members (23,24).…”

“…The Sgt2 dimer has a larger hydrodynamic radius than expected for a globular protein suggesting an extended conformation (11,12). SGT interacts with a variety of proteins, notably heat-shock proteins and their cognates (referred to here in general as HSC) such as Hsc70 and Hsp90, which bind directly to the TPR domain (11)(12)(13)(14). SGT binding to HSCs appears to modulate the chaperone ATPase activity and folding rates dependent on other HSC co-chaperones.…”

Biogenesis of tail-anchored proteins: the beginning for the end?

“…The GET4 and GET5 genes appear to be highly conserved, raising the possibility that their counterparts are present in the mammalian TM-recognition complex (see also Stefanovic and Hegde, 2007). Although Get4 has not been functionally characterized, Get5 has been found to associate with Sgt2, a factor involved in the coordination of several more ubiquitous chaperones such as Hsc70 and Hsp90 (Angeletti et al, 2002;Liou et al, 2007). Intriguingly, Get4 and Get5 have been suggested to associate with ribosomes on the basis of a proteomic analysis of yeast ribosomeassociated complexes (Fleischer et al, 2006).…”

Interaction Surface and Topology of Get3-Get4-Get5 Protein Complex, Involved in Targeting Tail-anchored Proteins to Endoplasmic Reticulum

“…Briefly, the pathway begins when Sgt2 recognizes the TMD of a nascent TA protein by its Met-rich C-terminal domain and then binds to a ubiquitin-like domain of Get5 via its N-terminal dimerization domain (11,12). The N-terminal region of Get5 (referred to as Get5N) then binds to the C-terminal region of Get4 to form a stable complex, predominantly through hydrophobic interactions (13).…”