THE HANSENULA-POLYMORPHA PER9 GENE ENCODES A PEROXISOMAL MEMBRANE-PROTEIN ESSENTIAL FOR PEROXISOME ASSEMBLY AND INTEGRITY

We have cloned and characterized the Hansenula polymorpha PERS gene by functional complementation of the per9-1 mutant of H. polymorpha, whi ch is defective in peroxisome biogenesis. The predicted product, Per9p , is a polypeptide of 52 Id)a with sequence similarity to Pas3p, a pro tein involved in peroxisome biogenesis in Saccharomyces cerevisiae, In a per9 disruption strain (Delta per9), peroxisomal matrix and membran e proteins are present at wild-type levels, The matrix proteins accumu lated in the cytoplasm, However, the location of the membrane proteins remained obscure; fully induced Delta per9 cells lacked residual pero xisomal vesicles (''ghosts''), Analysis of the activity of the PER9 pr omoter revealed that PER9 expression was low in cells grown on glucose , but was enhanced during growth of cells on peroxisome-inducing subst rates, The highest expression levels were observed in cells grown on m ethanol, Localization studies revealed that Per9p is an integral membr ane protein of the peroxisome. Targeting studies suggested that Per9p may be sorted to the peroxisome via the endoplasmic reticulum, Overexp ression of PER9 induced a significant increase in the number of peroxi somes per cell, a result that suggests that Per9p may be involved in p eroxisome proliferation and/or membrane biosynthesis, When PERS expres sion was placed under the control of a strongly regulatable promoter a nd switched off, peroxisomes were observed to disintegrate over time i n a manner that suggested that Per9p may be required for maintenance o f the peroxisomal membrane.