A conserved and essential basic region mediates tRNA binding to the Elp1 subunit of the Saccharomyces cerevisiae Elongator complex.
Bottom Line: Since these modifications are required for the tRNAs to function efficiently, a translation defect caused by hypomodified tRNAs may therefore underlie the variety of phenotypes associated with Elongator dysfunction.The Elp1 carboxy-terminal domain contains a highly conserved arginine/lysine-rich region that resembles a nuclear localization sequence (NLS).Thus the conserved basic region in Elp1 may be essential for tRNA wobble uridine modification by acting as tRNA binding motif.
Affiliation: Centre for Gene Regulation & Expression, College of Life Sciences, MSI/WTB Complex, University of Dundee, Dundee, DD1 5EH, Scotland, UK.Show MeSH
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Mentions: Since it was recently shown that Elongator forms a dodecamer containing two copies each of its six subunits (Glatt et al., 2012), it was possible that Elp1–KR9A could be defective in assembly into this higher order complex. To test this, YCp–ELP1–6HA and YCp–elp1–KR9A–6HA expression plasmids were transformed into ELP1–GFP and elp1–KR9A–GFP strains to generate cells in which two differentially tagged copies of Elp1 were present. The ELP1–GFP strain remained sensitive to zymocin following introduction of YCp–ELP1–6HA, confirming that the increased ELP1 copy number did not cause significant loss of function (data not shown). Wild-type Elp1–GFP and Elp1–KR9A–GFP were immunoprecipitated using GFP–Trap and assayed for co-precipitation of the 6HA-tagged versions of Elp1 (Fig. 5A). GFP-tagged wild-type Elp1 co-precipitated Elp1–6HA as previously reported (Glatt et al., 2012) and could also associate to a similar extent with Elp1–KR9A–6HA. In the reciprocal experiment, GFP tagged Elp1–KR9A also co-precipitated both Elp1–6HA and Elp1–KR9A–6HA, again with no obvious defect in the interaction between any of the different combinations. Thus there is no apparent defect in Elp1 self-association within the Elongator complex due to the Elp1 basic region mutation.
Affiliation: Centre for Gene Regulation & Expression, College of Life Sciences, MSI/WTB Complex, University of Dundee, Dundee, DD1 5EH, Scotland, UK.