Subcellular distribution of native estrogen receptor α and β subtypes in cultured human lens epithelial cells

The expression and intracellular localization of the estrogen receptor (ER) subtypes, ERα and ERβ were examined in cultured human lens epithelial cells (HLE-B3). ERα and ERβ mRNA expression was evaluated by coupled reverse transcription-polymerase chain reaction (RT-PCR) and Southern blot analysis. Subcellular localization of ERα and ERβ was determined on formaldehyde-fixed, Saponin-permeabilized cells using conventional immunofluorescence techniques, as well as immunodetection of differential cellular components after subjecting the cultured cells to fractionation by sucrose gradient centrifugation. Using RT-PCR, ER species specific primers distinguished mRNA from total RNA extracted from HLE-B3 cells, as well as from human breast adenocarcinoma cells (MCF-7), which provided a positive control. The 286-bp (ERα) and 167-bp (ERβ) PCR products were verified by sequence analysis. Southern blot analysis using internal oligonucleotides directed to specific primer pairs for ERα and ERβ, respectively, further confirmed the authenticity of the PCR products. HLE-B3 cells expressed ERα and ERβ in association with the nucleus and ERβ in the mitochondria. That the mitochondrial-enriched subfraction correlated with the presence of the ERβ subtype was confirmed by Western blot analysis. The differential subcellular partitioning of ERα and ERβ subtypes lends a new aspect to the estrogen signalling system wherein mitochondrial stabilization, possibly through ER binding, may play a causal role in the maintenance of cellular integrity. The occurrence of ER subtypes in human lens epithelial cells suggests that estrogen plays a role in the physiology of the lens. The comparative nuclear and mitochondrial distribution of ERα and ERβ raises the interesting probability that the two receptors, dependent upon subcellular localization, may have differential cytoprotective potential, and by inference, disparity in their mechanisms of action.