Demonstration of the mineralocorticoid hormone receptor and action in human leukemic cell lines.
We studied the expression of the mineralocorticoid receptor (MCR), and of the amiloride-sensitive sodium channel (ASSC) regulated by the MCR, in human leukemic cell lines. Cell extracts from TF1 (proerythroblastic), HEL (human erythroblastic leukemia) and U937 (myeloblastic) cell line were positive for the ASSC, as a 82 kDa band in Western blots developed with the aid of a polyclonal antibody raised against the peptide QGLGKGDKREEQGL, corresponding to the region 44-58 of the alpha subunit of the epithelial sodium channel (ENaC) cloned from rat colon, linked to KLH. The polyclonal antibody against the MCR revealed a single band of about 102 kDa in extracts from HEL and TF1 cells. The immunofluorescent labelling of the MCR in all cell lines showed a nucleocytoplasmic localization of the receptor but the ASSC was exclusively membrane-bound and these results were confirmed by confocal microscopy. The expression of the MCR in the HEL cells was evident as a predicted band of 843 bp (234 amino acids) in electrophoresis of the PCR product obtained after total RNA had been reverse transcribed and then amplified using the primers 5'-AGGCTACCACAGTCTCCCTG-3' and 5'-GCAGTGTAAAATCTCCAGTC-3' (sense and antisense, respectively). The ENaC was similarly evident with the aid of the primers 5'-CTGCCmATG GATGATGGT-3' (sense) and 5'-GTTCAGCTCGAAGAAGA-3' (antisense) as a predicted band of 520 bp. In both cases, 100% identity was observed between the sequences of the PCR products compared to those from known human sources. The multiplication of the HEL cells was influenced by antagonists (RU 26752, ZK 91587) targeted for specificity to the MCR and this was selectively reversed by the natural hormone aldosterone. These steroids also provoked chromatin condensation in the HEL population. These permit new and novel possibilities to understand the pathobiology of human leukemia and to delineate sodium-water homeostasis in nonepithelial cells.
Mirshahi
M
M
CNRS and Inserm E 9912, Paris, France.
Mirshahi
S
S
Golestaneh
N
N
Mishal
Z
Z
Nicolas
C
C
Hecquet
C
C
Agarwal
M K
MK
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D017931:DNA Primers; D006801:Humans; D007150:Immunohistochemistry; D007938:Leukemia / Q000378:metabolism* / Q000473:pathology; D018613:Microscopy, Confocal; D016133:Polymerase Chain Reaction; D051381:Rats; D018161:Receptors, Mineralocorticoid / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D017931:DNA Primers; D018161:Receptors, Mineralocorticoid