The ypt1-G80D and ypt1-3 mutants show defect in the secretory pathway

The putative translation initiation factor 5A (eIF5A) is essential for cell viability and the only cellular protein known to contain the unusual aminoacid residue hypusine. eIF5A has been implicated in translation initiation, cell proliferation, nucleocytoplasmic transport and mRNA decay, but the precise biological function of eIF5A is not clear. A screen for synthetic lethal mutations was carried out with one of the temperature-sensitive alleles of TIF51A (tif51A-3) to identify factors that functionally interacts with eIF5A and revealed a mutation (G80D) in the essential gene YPT1. This gene encodes a small GTPase, acting in the vesicular trafficking between the Endoplasmatic Reticulum and the Golgi. In order to assess the secretory pathway, we investigated the maturation of the vacuolar glycoprotein carboxipeptidase Y (CPY) and verified that the ypt1-G80D and ypt1-3 mutants accumulate precursor forms of CPY at the nonpermissive temperature (37ºC), indicating a defect in the secretory pathway. This is in agreement with the defect described in the ypt1-1 and ypt1-A136D mutants. Moreover, we were able to show that high-copy TIF51A supresses the temperature-sensitive phenotype of ypt1-3 but does not recover the CPY maturation defect of this mutant. We are also analyzing the secretory pathway in eIF5A mutants. Although the genetic interactions between YPT1 and TIF51A suggest that the proteins encoded by these genes are intimately associated, further analysis will be necessary to clarify the functional interaction of Ypt1 and eIF5A.