Biogenesis of the yeast vacuole (lysosome)

Abstract

The vacuolar proteinase yscB (PrB) has been implicated in the final maturation of procarboxypeptidase yscY (pro‐CpY) to the mature wild‐type form CpY^b of 61 kDa. In PrB‐deficient mutants, only the proteinase yscA processed form CpY^a of 62 kDa is found [Mechler, B., Müller, H. & Wolf, D. H. (1987) EMBO J. 6, 2157–2163]. We report now that, akin to CpY, two forms of mature proteinase yscA (PrA) can be distinguished. In PrB‐deficient mutant cells, PrA^a, migrating at about 43 kDa in SDS/PAGE, is found, whereas PrA^b, found in wild‐type cells, had the known molecular mass of 42 kDa. In the PrB‐deficient strain, pro‐PrA and pro‐CpY matured only to the higher‐molecular‐mass forms, PrA^a and CpY^a, and the maturation of both precursors was slower than in the isogenic wild‐type strain. Pulse‐labeling experiments indicated that the mature forms, PrA^b or CpY^b, are generated directly in the PrB‐containing wild‐type strain in vivo. In vitro experiments showed that PrB is able to trigger maturation of its 42‐kDa pro‐PrB precursor to mature PrB in the absence of PrA. Mature PrB and its proteolytic activity, however, shows a higher stability in the presence of mature PrA. The data indicate a molecular and kinetic participation of proteinase yscB in vacuolar hydrolase precursor maturation.