Regulatory control of the Na-Cl co-transporter NCC and its therapeutic potential for hypertension

Abstract

Hypertension is the largest risk factor for cardiovascular disease, the leading cause of mortality worldwide. As blood pressure regulation is influenced by multiple physiological systems, hypertension cannot be attributed to a single identifiable etiology. Three decades of research into Mendelian forms of hypertension implicated alterations in the renal tubular sodium handling, particularly the distal convoluted tubule (DCT)-native, thiazide-sensitive Na-Cl cotransporter (NCC). Altered functions of the NCC have shown to have profound effects on blood pressure regulation as illustrated by the over activation and inactivation of the NCC in Gordon's and Gitelman syndromes respectively. Substantial progress has uncovered multiple factors that affect the expression and activity of the NCC. In particular, NCC activity is controlled by phosphorylation/dephosphorylation, and NCC expression is facilitated by glycosylation and negatively regulated by ubiquitination. Studies have even found parvalbumin to be an unexpected regulator of the NCC. In recent years, there have been considerable advances in our understanding of NCC control mechanisms, particularly via the pathway containing the with-no-lysine [K] (WNK) and its downstream target kinases, SPS/Ste20-related proline-alanine-rich kinase (SPAK) and oxidative stress responsive 1 (OSR1), which has led to the discovery of novel inhibitory molecules. This review summarizes the currently reported regulatory mechanisms of the NCC and discusses their potential as therapeutic targets for treating hypertension.

Keywords: ATP, adenosine triphosphate; Blood pressure regulation; CCC, cation-coupled chloride cotransporters; CCT, conserved carboxy-terminal; CNI, calcineurin inhibitors; CUL3, cullin 3; CUL3/KLHL3-WNK-SPAK/OSR1; Ca2+, calcium ion; Cardiovascular disease; DAG, diacylglycerol; DCT, distal convoluted tubule; DUSP, dual specificity phosphatases; ECF, extracellular fluid; ELISA, enzyme-bound immunosorbent analysis; ERK, extracellular signal-regulated kinases; EnaC, epithelial sodium channels; GABA, gamma-aminobutyric acid; HEK293, human embryonic kidney 293; Hypertension; I1, inhibitor 1; K+, potassium ion; KCC, potassium-chloride-cotransporters; KLHL3, kelch-like 3; KS-WNK1, kidney specific-WNK1; Kinase inhibitors; MAPK, mitogen-activated protein kinase; MO25, mouse protein-25; Membrane trafficking; NCC, sodium–chloride cotransporters; NKCC, sodium–potassium–chloride-cotransporter; Na+, sodium ion; NaCl, sodium chloride; NaCl-cotransporter NCC; OSR1, oxidative stress-responsive gene 1; PCT, proximal convoluted tubule; PHAII, pseudohypoaldosteronism type II; PP, protein phosphatase; PV, parvalbumin; ROMK, renal outer medullary potassium; RasGRP1, RAS guanyl-releasing protein 1; SLC12, solute carrier 12; SPAK, Ste20-related proline-alanine-rich-kinase; TAL, thick ascending limb; Therapeutic targets; WNK, with-no-lysine kinases; mDCT, mammalian DCT; mRNA, messenger RNA.

Graphical abstract

Figure 1

Sodium handling in the distal nephron. The thick ascending limb (TAL) is a region responsible for 20%–30% of sodium (Na⁺) reabsorption. The predominant mechanism of transport in the TAL is the Na–K–Cl cotransporter 2 (NKCC2). The distal convoluted tubule (DCT) is responsible for 5%–10% of Na⁺ reabsorption. The major Na⁺ transport in the DCT is the Na–Cl co-transporter (NCC). Other ion transport mechanisms include the renal outer medullary potassium channel (ROMK), the sodium potassium pump (Na⁺/K⁺ ATPase) and the chloride channel Kb (CLC-Kb).

Figure 2

Proposed integrated model of NCC regulation. The Na⁺–Cl⁻ co-transporter (NCC) is the principal salt absorptive pathway in the distal convoluted tubule (DCT). The NCC is activated by kinase-induced phosphorylation (P) via the with-no-lysine [K] (WNK) and its downstream target kinases, SPS/Ste20-related proline–alanine-rich kinase (SPAK) and oxidative stress responsive (OSR) and deactivated by phosphatase-induced dephosphorylation via protein phosphatase 1-3-4 (PP1-3-4). Activation of SPAK is enhanced via attachment of the mouse protein-25 (MO25). The expression of NCC is directly regulated via ubiquitination and subsequent endocytosis of NCC via the RAS guanyl nucleotide-releasing protein (RasGRP) and its downstream target extracellular signal-regulated protein kinase (ERK) 1/2 and NEDD4-2 or indirectly via ubiquitination and suppression of WNK by the cullin3 (CUL3)/kelch-like-3 (KLHL3) ubiquitin ligase complex. ERK1/2 is inhibited by DUSP6 and the phosphatases are inhibited by an endogenous inhibitor 1 (I1). Further studies are required to evaluate the NCC regulatory effects of parvalbumin (PV) via modulations of the ATP-induced Ca²⁺ signaling. The modulators of these regulatory events include parathyroid hormone (PTH), aldosterone, purine receptors and cAMP elevating hormones.

Figure 3

Summary of therapeutic strategies to target the NCC via its regulatory pathway. Over activation of the Na⁺–Cl^– co-transporter (NCC) leads to salt retention and hypertension. The CUL3/KLHL3–WNK–SPAK/OSR1 regulatory pathway of NCC (black arrows) presents 6 points of interventions. Red arrows represent the therapeutic inhibition of 1) NCC, 2) WNK, 3) SPAK, 4) MO25, and 5) WNK–SPAK/OSR1 interactions which would all suppress NCC activation. The green arrow represents 6) the stabilisation of CUL3/KLHL3 interactions which would increase degradation of WNK and thus suppress NCC activation by the WNK–SPAK/OSR1 pathway. Glycosylation of the NCC is critical for the function and trafficking of NCC to the plasma membrane and thus (7) impairment of glycosylation could also suppress NCC activity and potentially be therapeutic.