KCl cotransport activation in human erythrocytes by high hydrostatic pressure.

Godart, H; Ellory, J. Clive

doi:10.1113/jphysiol.1996.sp021226

Cited by 20 publications

(10 citation statements)

References 36 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In KCC3-transfected HEK-293 cells, 5 M staurosporine gave a 3-fold stimulation in KCC3 activity (8). Compared with KCC1 in human erythrocytes (23) and KCC3 in HEK-293 cells (8), the staurosporine effect was much attenuated in our model. The present study also shows that tyrosine phosphorylation is involved in the regulation of KCC3 activity.…”

Section: Discussionmentioning

confidence: 71%

“…Staurosporine, a protein kinase inhibitor of high potency and broad specificity, is reported to stimulate KCC activity in various cell types (8,22). For example, the half-maximum stimulatory concentration of staurosporine is about 0.5 M to activate KCC1 in human erythrocytes (23). However, 2 M staurosporine did not activate KCC1 activity in HEK-293 cells transfected with full-length rabbit KCC1 (24).…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

The KCl cotransporter isoform KCC3 can play an important role in cell growth regulation

Shen

Chou

Hsu

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

The KCl cotransporter (KCC) plays a significant role in the ionic and osmotic homeostasis of many cell types. Four KCC isoforms have been cloned. KCC1 and KCC4 activity is osmolality-sensitive and involved in volume regulation. KCC2, a neuronal-specific isoform, can lower intracellular Cl ؊ and is critical for inhibitory GABA responses in the mature central nervous system. KCC3, initially cloned from vascular endothelial cells, is widely but not universally distributed and has an unknown physiological significance. Here we show a tight link between the expression and activity of KCC3 and cell growth by a NIH͞3T3 fibroblast expression system. KCC3 activity is sensitive to [(dihydroindenyl)oxy] alkanoic acid (DIOA) and N-ethylmaleimide and is regulated by tyrosine phosphorylation. Osmotic swelling does not activate KCC3, and the process of regulatory volume decrease is refractory to DIOA, indicating that KCC3 is not involved in volume regulation. KCC3 expression enhances cell proliferation, and this growth advantage can be abolished by the inhibition of KCC3 by DIOA. Fluorescence-activated cell sorting measurements and Western blot analysis show DIOA caused a significant reduction of the cell fraction in proliferative phase and a change in phosphorylation of retinoblastoma protein (Rb) and cdc2, suggesting that KCC3 activity is important for cell cycle progression. Insulin-like growth factor-1 up-regulates KCC3 expression and stimulates cell growth. Tumor necrotic factor-␣ down-regulates KCC3 expression and causes growth arrest. These data indicate that KCC3 is an important KCC isoform that may be involved in cell proliferation. The KCl cotransporter (KCC) has been implicated not only in regulatory volume decrease but also in transepithelial salt absorption, renal K ϩ secretion, myocardial K ϩ loss during ischemia, and regulation of neuronal Cl Ϫ concentrations (1-3). A major advance in the understanding of KCC has been the recent identification of genes that encode four KCC isoforms. KCC1 is a ''housekeeping'' isoform for cell volume regulation (4). KCC2, a neuronal-specific isoform, by lowering intracellular Cl Ϫ , is critical for inhibitory GABA responses in the mature central nervous system (5, 6). The KCC3 gene is located on chromosome 15q13, which colocalizes with the gene for myoclonal epilepsy, and the mRNA of KCC3 has been found in vascular endothelial cells, heart, kidney, brain, placenta, liver, and lung (7-9). Despite the wide distribution of KCC3, next to nothing is known about the functional and regulatory properties of this isoform. KCC4, predominantly found in heart and kidney, is volume-sensitive and also contributes to volume regulation (9, 10).The present study was aimed at characterizing the regulation and function of the recently cloned human KCC3 by using the NIH͞3T3 fibroblast expression system. NIH͞3T3 cells, which, unlike many other cell types [such as human embryonic kidney (HEK)-293 cells and Xenopus oocytes], do not have endogenous KCC activity and therefore provide a suitable system fo...

show abstract

Section: Discussionmentioning

confidence: 71%

Section: Discussionmentioning

confidence: 99%

The KCl cotransporter isoform KCC3 can play an important role in cell growth regulation

Shen

Chou

Hsu

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

show abstract

“…4 at 37ЊC with either HCl or Tris base (monitored before and after the experimental incubation period). For experimental convenience, KCl cotransport activity was measured as unidirectional K + influx with Rubidium 86 ( 86 Rb + ) as a K + congener, at a final activity of 37 kBq/ml (Godart & Ellory, 1996). In all experiments the final K + concentration was 7 .…”

Section: Methodsmentioning

confidence: 99%

Do HbSS erythrocytes lose KCl in physiological conditions?

Dormandy¹,

Ellory²

1997

Br J Haematol

View full text Add to dashboard Cite

Summary. KCl cotransporter activity in sickle (HbSS) red blood cells (RBCs) was measured in cells suspended in 'simple' physiological saline, saline augmented with inorganic salts, and autologous plasma. Our results showed that the transporter was only functioning at 20% of the level of cells in saline when cells were resuspended in autologous plasma. Kinetic analysis of the data showed that plasma decreased both V max and K m for K + of the transporter. The plasma factor(s) responsible was heat-stable and dialysable (i.e. size < 10 kD).Adding magnesium, calcium, inorganic phosphate or bicarbonate to 'simple' saline to mimic the effect of plasma revealed that Mg 2+ and Ca 2+ had no significant effect at physiological concentrations. P i was not effective at 1 . 1 mM , but did inhibit significantly (42Ϯ2%Þ at 5 . 6 mM . HCO ¹ 3 had a major inhibitory effect on K + influx when added to saline, and was identified as the principal candidate for the plasma effect.We suggest bicarbonate may play a significant role in modifying KCl cotransport, and hence HbSS cell volume in vivo. It acts by altering the set point of the transporter via the signalling systems involved in its regulation.

show abstract

“…Figure 6 shows the dose-response curve of calyculin A and okadaic acid, two potent PP inhibitors, for the inhibition of hypotonicity-induced 86 Rb + efflux. These two inhibitors have been reported to affect transport after a delay [19]. Therefore, cervical cancer SiHa cells were preincubated for 20 min at 37°C with various concentrations of inhibitors before the efflux assays.…”

Section: Role Of Protein Kinase/phosphatase In Regulating the Kcc In mentioning

confidence: 99%

Volume-sensitive KCl cotransport associated with human cervical carcinogenesis

Shen

Chou

Ellory

2000

Pflugers Arch - Eur J Physiol

View full text Add to dashboard Cite

This study investigates the volume-sensitive KCI cotransporter (KCC) in various types of human cervical epithelial cell, testing the hypothesis that cervical malignancy is accompanied by differential expression of volume-sensitive KCC. Normal human cervical epithelial cells have KCCs which are quiescent in normal physiological conditions and are relatively refractory to hypotonic stress. By contrast, cervical cancer cells have KCCs which are also nearly quiescent in normal physiological conditions but high transport rates are observed in response to hypotonic challenge. Using isoform-specific primers, mRNA transcripts of KCC1, KCC3 and KCC4 were identified by reverse transcriptase polymerase chain reaction (RT-PCR) in several types of cervical cell, and confirmed by digestion with specific restriction endonucleases. By semiquantitative RT-PCR with beta-actin as the internal standard, the results indicate that cervical carcinogenesis is accompanied by the up-regulation of mRNA transcripts in KCC1, KCC3 and KCC4. [(Dihydroindenyl)oxy] alkanoic acid (DIOA), a KCC inhibitor, blocked both the regulatory volume decrease (RVD) process and volume-sensitive 86Rb+ efflux from cervical cancer cells in a dose-dependent manner. The volume-sensitive 86Rb+ efflux from cervical cancer cells was also blocked by two protein phosphatase inhibitors, calyculin A and okadaic acid, with IC50 values of 0.8 and 6 nM, respectively. Conversely, protein kinase inhibitors, chelerythrine and staurosporine, increased Cl- dependent 86Rb+ efflux. NEM (1 mM) led to a fivefold stimulation of 86Rb+ efflux which was totally Cl- dependent in cervical cancer cells. Hypotonicity could not stimulate any further 86Rb+ efflux after NEM treatment. These results indicate that the volume-sensitive KCC in cervical cancer cells plays a significant role in volume regulation and that the activities are modulated by a phosphorylation cascade. Taken together with our previous studies, we suggest the volume-regulatory ion channels and the co-transport systems work synergistically for volume regulation in human cervical cancer cells.

show abstract

KCl cotransport activation in human erythrocytes by high hydrostatic pressure.

Cited by 20 publications

References 36 publications

The KCl cotransporter isoform KCC3 can play an important role in cell growth regulation

The KCl cotransporter isoform KCC3 can play an important role in cell growth regulation

Do HbSS erythrocytes lose KCl in physiological conditions?

Volume-sensitive KCl cotransport associated with human cervical carcinogenesis

Contact Info

Product

Resources

About