Biogenesis of peroxisomes: Isolation and characterization of two distinct peroxisomal populations from normal and regenerating rat liver

Abstract

According to Poole et al. (1970. J. Cell Biol. 45:408-415), newly synthesized peroxisomal proteins are incorporated uniformly into peroxisomes (PO) of different size classes, suggesting that rat hepatic PO form a homogeneous population. There is however increasing cytochemical and biochemical evidence that PO in rat liver are heterogenous, undergoing significant modulations in shape and size in process of PO morphogenesis (Yamamoto and Fahimi, 1987. J. Cell Biol. 105:713-722). In the present study, the kinetics of incorporation of newly synthesized proteins into distinct PO-subpopulations have been studied using short-term in vivo labeling (5-90 min). Two distinct "heavy" and "light" crude PO fractions were prepared by differential pelleting from normal and regenerating liver, and highly purified PO were subsequently isolated by density-dependent metrizamide gradient centrifugation according to Völkl and Fahimi (1985. Eur. J. Biochem. 149:257-265). The peroxisomal fractions banded at 1.20 and 1.24 g × cm-3. They differed in their mean diameters and form-factors and particularly in respect to the activity of β-oxidation enzymes which was higher in the "light" PO. Whereas the "light" PO exhibited a single immunoreactive band with the antibody to the 70-kD peroxisomal membrane protein the "heavy" PO contained an additional (68 kD) band. In pulse-labeling experiments "light" PO showed clearly a higher initial rate of incorporation than the "heavy" PO. The relative specific activity in the "heavy" PO fraction, however increased progressively reaching that of "light" PO by 90 min. These observations provide evidence for the existence of different PO populations in rat liver which differ in their morphological and biochemical properties as well as in their rates of incorporation of new proteins.