Characterization of DFG5，Involved in Cell Wall Biogenesis of Saccharomyces cerevisiae

Abstract:

Saccharomyces cerevisiae possesses cell wall-localized proteins（CWPs），which are essential for the growth. The majority of CWPs are initially synthesized as glycosylphosphatidylinositol（GPI）-anchored proteins，followed by transfer of the protein part to the β-glucan. Two GPI-anchored proteins localized at the plasma membrane，Dfg5 and Dcw1，have been proposed to participate in the transfer reaction，however，molecular functions of these homologous proteins are still unclear. In this study，we characterized the Dfg5 using a conditional mutant strain in which DFG5 was disrupted and DCW1 expression was controlled under MET3 promoter. We found that down-regulation of DCW1 expression in the mutant strain caused a great accumulation of our model CWP（Cwp1） in the plasma membrane. Site-directed mutagenesis of Dfg5 revealed that three residues conserved in glycoside hydrolase family 76 were necessary to complement the lethality of the mutant strain. In contrast，the growth defect was complemented by the expression of soluble form of Dfg5，which was secreted in culture medium due to a lack of its pro-peptide sequence. Taken together，these results suggested that at least Dfg5 acts as an enzyme to release CWPs from GPI anchor at plasma membrane.