Hypertonic upregulation of betaine transport in renal cells is blocked by a proteasome inhibitor

Abstract

The renal betaine transporter (BGT1) protects cells in the hypertonic medulla by mediating uptake and accumulation of the osmolyte betaine. Transcription plays an essential role in upregulating BGT1 transport in MDCK cells subjected to hypertonic stress. During hypertonic stress, the abundance of the transcription factor TonEBP increases and it shifts from the cytoplasm to the nucleus where it activates transcription of the BGT1 gene. Little is known about post-transcriptional regulation of BGT1 protein. In the presence of the proteasome inhibitor MG-132, which blocked nuclear translocation of TonEBP, the hypertonic upregulation of BGT1 protein and transport was prevented and cell viability in hypertonic medium was impaired over 24 h. Urea also prevented the hypertonic upregulation of BGT1 protein and transport, but did not interfere with TonEBP translocation and cell viability. Shorter treatments of hypertonic cells with MG-132 avoided viability problems and produced dose-dependent inhibition of translocation and transport. When stably transfected MDCK cells that over-expressed BGT1 were treated for 6 h with hypertonic medium containing 3 microM MG-132, there was 43% inhibition of nuclear translocation, 83% inhibition of BGT1 transport, and no change in viability. While other proteasome functions may be involved, these data are consistent with a critical role for nuclear translocation of TonEBP in upregulation and membrane insertion of BGT1 protein.

Functional Expression of γ–Amino Butyric Acid Transporter 2 in Human and Guinea Pig Airway Epithelium and Smooth Muscle

“…It is possible that GAT4/BGT1, although present in these cells, either has a different IC 50 value in airway epithelial and smooth muscle cells than in neuronal cells, or serves a different function, such as transport of betaine. In renal cells, the transport of betaine is implicated in maintenance of intracellular osmotic pressure, and it may play a similar role in astrocytes (4,5,26). The role of GAT4/BGT1 in airway epithelial and smooth muscle cells is not clear.…”

Osmoregulatory inositol transporter SMIT1 modulates electrical activity by adjusting PI(4,5)P2levels

“…Hypertonic treatment speeded return of the PH-PLCδ1-RFP probe to the plasma membrane after Oxo-M; a proteasome inhibitor MG-132, which attenuates the translocation of TonEBP from the cytosolic region to the nucleus (59,60), diminished this effect (Fig. 7 I and J).…”

Effects of hyperosmolarity on the Na+-myo-inositol cotransporter SMIT2 stably transfected in the Madin-Darby canine kidney cell line

“…More recently, the transcriptional activator, tonicity-responsive enhancer binding protein (TonEBP), expressed in kidney (24,37) among other tissues, has been shown to be a crucial mediator involved in the activation of many of these transporters (23,42,46). In response to hyperosmotic challenge, TonEBP levels are increased, and the protein is mobilized to the nucleus to increase synthesis of target proteins, such as those mentioned above (10,17,18,32,51). Furthermore, by means of specific inhibitors, modulation of TonEBP has been linked to p38 (27,44).…”

“…Although the transfected SMIT2 vector does not contain a known tonicityresponsive enhancer (TonE) sequence, we examined the possible involvement of TonEBP in the stimulation of SMIT2 by using the proteasome inhibitor MG-132, which also inhibits TonEBP activity (32,52). As shown in Fig.…”