Phorbol esters downregulate expression of the sodium/calcium exchanger in renal epithelial cells

Abstract

The Na+/Ca2+ exchanger (NCE) contributes to Ca2+ reabsorption by connecting tubules of the nephron. A line of renal epithelial cells from monkey kidney (LLC-MK2) was used to investigate the regulation of NCE expression. After the activation of protein kinase C (PKC) by phorbol myristate acetate (PMA), NCE activity decreased exponentially by 75% in 48 h (half time approximately 19 h). PMA decreased NCE mRNA by 85% in 24 h. The decrease in NCE transcript preceded the downregulation of NCE activity. NCE protein was quantified with a monoclonal antibody to cardiac NCE. PMA decreased the binding of 3H-labeled antibody to cell sonicates by 40% in 24 h. Immunoblots show that PMA produced a marked and extended increase in membrane-associated PKC-alpha, although PMA depleted total PKC-alpha by 65% in 24 h. In vivo 32P labeling of myristolated alanine-rich C kinase substrate, a specific PKC substrate, confirmed that PMA produced a rapid and extended activation of PKC. 4 alpha-PMA, a stereoisomer of PMA that neither binds nor activates PKC, had no effect on NCE activity or transcript. These findings indicate that activation of PKC with phorbol esters downregulates NCE mRNA, protein, and activity in renal epithelial cells.

Immunohistochemical Detection of the Sodium‐Calcium Exchanger in Rat Hippocampus Cultures Using Subtype‐Specific Antibodies

“…Hence, a lack of glia labeling could result if this change would cause a significant decrease in the affinity of mAb R3F1. However, this explanation was judged less likely since the antibody recognizes so well the kidney isoforms of NCX114 that are known to belong to the B family. (2) Alternatively, the expression of NCX1 in glial cells might depend on culture conditions or on the developmental stage of the cultivated cells.…”

Intracellular protein phosphorylation in adherent U937 monocytes mediated by various culture conditions and fibronectin-derived surface ligands

“…Cross-referencing with the Swiss-Prot ® database and TrEMBL ® (http:// us.expasy.org), several potential adhesion-related phosphotyrosine proteins matching the molecular weight range are likely candidate targets of PMA in U937. Proteins that could be upregulated include: non-receptor tyrosine protein kinase TYK2 (134 kDa) [18], adapterrelated protein complex 2 alpha 2 subunit (104 kDa), epidermal growth factor receptor substrate 15 (99 kDa) [19], CD97 antigen (82 kDa) [20], mitogen-activated protein kinase 6 (83 kDa) [21], paxillin (65 kDa) [22], transcription factor NF-E2 45 kDa subunit (41 kDa) [23], mitogen-activated protein kinase 1 (41 kDa) [24], mitogen-activated protein kinase 3 (43 kDa), and myristoylated alanine-rich C-kinase substrate (31 kDa) [25]. Plausible downregulated proteins include: mitogen-activated protein kinase 4 (63 kDa) [24] and nuclear transcription factor Y subunit beta (23 kDa) [26], hinting that these phosphotyrosine proteins might serve as constraints in cell adhesion and/or cell spreading.…”

Calcineurin Controls the Transcription of Na+/Ca2+ Exchanger Isoforms in Developing Cerebellar Neurons

“…Although ␣-adrenergic stimulation led to the increase of NCX1 mRNA in cultured cardiac myocytes (25), glucocorticoids and protein kinase A down-regulated it in vascular smooth muscle cells. Protein kinase C had the same effect in endothelial cells (26,27). Changes in the expression of the NCX1 gene have also been observed during cardiac development (28) and pressure overload (29).…”