The Journal of Cell Biology, Vol 89, 62-69, Copyright © 1981 by The Rockefeller University Press
Analytical study of microsomes and isolated subcellular membranes from rat liver. VII. Distribution of protein-bound sialic acid
A Amar-Costesec
Detailed investigations by quantitative centrifugal fractionation were
conducted to determine the subcellular distribution of protein-bound sialic
acid in rat liver. Homogenates obtained from perfused livers were
fractionated by differential centrifugation into nuclear fraction, large
granules, microsomes, and final supernate fraction, or were used to isolate
membrane preparations enriched in either plasma membranes or Golgi complex
elements. Large granule fractions, microsome fractions, and plasma membrane
preparations were subfractionated by density equilibration in linear
gradients of sucrose. In some experiments, microsomes or plasma membrane
preparations were treated with digitonin before isopycnic centrifugation to
better distinguish subcellular elements related to the plasma membrane or
the Golgi complex from the other cell components; in other experiments,
large granule fractions were obtained from Triton WR-1339-loaded livers,
which effectively resolve lysosomes from mitochondria and peroxisomes in
density gradient analysis. Protein-bound sialic acid and marker enzymes
were assayed in the various subcellular fractions. The distributions
obtained show that sialoglycoprotein is restricted to some particular
domains of the cell, which include the plasma membrane, phagolysosomes, and
possibly the Golgi complex. Although sialoglycoprotein is largely recovered
in the microsome fraction, it has not been detected in the endoplasmic
reticulum-derived elements of this subcellular fraction. In addition, it
has not been detected either in mitochondria or in peroxisomes. Because the
sialyltransferase activities are associated with the Golgi complex, the
cytoplasm appears compartmentalized into components which biogenetically
involve the Golgi apparatus and components which do not.