Estrogen-dependent expression and subcellular localization of the tight junction protein claudin-4 in HEC-1A endometrial cancer cells

Abstract

Abstract. Endometrial cancer is the most common female reproductive cancer in the United States and is associated with deregulated tight junction protein expression. Given the highly estrogen-responsive nature of this tissue, we investigated the effects of estrogen and its agonist, 4-OH TAM, on the expression and subcellular localization of the tight junction protein claudin-4 (CLDN-4), in HEC-1A endometrial cancer cells. In untreated HEC-1A cells, we observed dramatic overexpression of claudin-4 protein. In addition, differential detergent extraction analysis indicated that claudin-4 was localized primarily in the membrane but also found in the cytosolic, nuclear and cytoskeletal fractions. Upon exposure of HEC-1A to estradiol (E 2 ), we observed a biphasic effect both on the overall expression of claudin-4 protein and on its cytosolic and cytoskeletal presence as demonstrated by immunoblot analysis. Immunofluorescence analysis also revealed a biphasic effect of E 2 on claudin-4 expression. In contrast, we observed no changes in expression levels nor in the subcellular distribution patterns of claudin-4 in HEC-1A cells treated with different concentrations of 4-OH TAM. The intracellular presence of CLDN-4 coupled with the biphasic effects of E 2 on CLDN-4 expression in the cytoskeleton suggest that this protein may be involved in cell signaling to and from TJs.

Claudins: New Players in Human Fertility and Reproductive System Cancers

“…Moreover, claudin-2 was observed in the nucleus in human lung adenocarcinoma cells [83], and claudin-3 was reported to be present in nuclei in breast cancer cell lines [84] and human colorectal cancer [85]. Changes in claudin-4 location in endometrial cancer cells were also observed [86]. Interestingly, relocation of claudin-1 is not associated with protein modification, based on not-observed changes in the mass of this protein [80], whereas claudin-2 was reported to be dephosphorylated when located in nuclei of cells [83].…”

“…Cuevas et al showed that claudin-4 expression in HEC-1A cells exposed to physiological concentrations of estradiol (100 nM) was modulated, and its localization was changed. Cells without hormonal treatment presented most of the claudin-4 localization in cytoplasm and membrane, whereas 50 nM of estradiol caused an increase in the expression and membrane localization [86]. Claudin-11 is known to be regulated hormonally, and its knockout models result in infertility [87].…”

Tight Junction Proteins and the Biology of Hepatobiliary Disease

“…Nuclear localization has been reported for ZO1/ZO2 [69,70] and CLDN1-4 [71][72][73][74] in several types of cancer cells. The conditions or inducers under which these TJ proteins localize in the nucleus are poorly understood.…”

TGF-β1 increases permeability of ciliated airway epithelia via redistribution of claudin 3 from tight junction into cell nuclei

“…They might serve as a claudin pool for tight junction protein recycling [70] or being part of the cellular interface to the extracellular matrix [14,36]. Also nuclear localization was reported for claudin 1, 2, 3 and 4 especially in tumour cells [11,24,34,65] suggesting that claudins may act as modulators of gene transcription, cell proliferation and migration.…”