Ak. Rajasekaran et al., A SIMPLE BIOCHEMICAL APPROACH TO QUANTITATE ROUGH ENDOPLASMIC-RETICULUM, American journal of physiology. Cell physiology, 37(2), 1995, pp. 308-316
In this report we demonstrate that the changes in size of the rough en
doplasmic reticulum (RER) can be determined by quantifying the membran
e-bound ribosomal population separated by cell fractionation and sucro
se density gradient analysis. Total cell membranes, rather than micros
omes, were used as the source of membrane-bound ribosomes to eliminate
potential losses during the preparation of microsomes. Bound ribosome
s were assayed after quantitative release and recovery from total cell
membranes using puromycin in the presence of high-salt buffer. Using
this analysis, we demonstrate a 4.2-fold increase in RER in estrogen-t
reated male Xenopus laevis liver. Furthermore, we show that the ratio
of the distribution of free to membrane-bound ribosomes in a nonsecret
ory cell line (HeLa) was 3.3, while this ratio in a secretory cell lin
e (AR42J) was 1.2, indicating that cells active in secretion contain m
ore RER. We suggest that this biochemical technique provides a simpler
assay to detect changes in the size of the RER.