Primary cilia and regulation of renal Na+transport. Focus on “Heightened epithelial Na+channel-mediated Na+absorption in a murine polycystic kidney disease model epithelium lacking apical monocilia”

Abstract

EDITORIAL FOCUSPrimary cilia and regulation of renal Na+ transport. Focus on “Heightened epithelial Na+ channel-mediated Na+ absorption in a murine polycystic kidney disease model epithelium lacking apical monocilia”M. A. GrayM. A. GrayPublished Online:01 Apr 2006https://doi.org/10.1152/ajpcell.00640.2005MoreSectionsPDF (50 KB)Download PDF ToolsExport citationAdd to favoritesGet permissionsTrack citations ShareShare onFacebookTwitterLinkedInEmailWeChat primary cilia (PC) are solitary, phallus-like structures that protrude into the extracellular environment from the surface of a surprisingly large number of different cell types (see Ref. 15 for a recent review), including most epithelial cells. They are generally nonmotile, apart from a specialized version known as nodal cilia, which are found in the node of the gastrulation stage embryo and which play an essential role in determining left-right axis patterning. Despite being identified >100 years ago, the precise function of the PC remains unclear in most mammalian cells. Because of their location, they are likely to act as some form of “sensor” that relays information about the extracellular environment to the cell body and thereby signal changes in processes as diverse as differentiation, proliferation, and membrane ion transport. However, exactly what they sense (chemicals, osmolarity, and flow) and how they transduce these sensations into a cellular response remain major unanswered questions in the ever-burgeoning “primary cilium” field.Primary cilia have certainly been in the spotlight in the last few years, mainly because of research that links them to diverse areas such as embryonic development, left-right patterning, and, in particular, to many forms of hereditary renal disease, such as polycystic kidney disease (PKD), the most common cause of end-stage renal failure in adults (19). Indeed, the majority of proteins linked to cystic disease (renal and extrarenal) appear to localize to the PC-basal body complex, making this structure a major focus of cystic disease research (see Ref. 3 for a recent review). In addition, two major cell signaling pathways, the canonical and noncanonical Wnt pathway (17) and the Hedgehog-based signaling system (2), have also very recently been linked to the PC, suggesting that this organelle, in addition to renal disease, orchestrates key events in embryonic development, limb formation, and possibly cancer.Recent studies have helped to shed new light on the role of the PC in renal tubules. Work from two groups (12, 15) indicates that PC function as mechanoreceptors that sense alterations in tubular flow (by bending) and transduce this into an increase in intracellular Ca2+ concentration ([Ca2+]i). Bending of the PC is linked to the opening of a plasma membrane Ca2+-permeable cation channel, which induces further Ca2+ release from internal stores through a process of Ca2+-induced Ca2+ release. The cation channel is actually a complex of two proteins, polycystin-1 and -2, the latter being a member of the transient receptor potential (TRP) family of Ca2+-influx channels (4). Importantly, mutations in either of the genes that code for polycystin 1 and 2 (PKD1 and PKD2) cause autosomal dominant PKD (ADPKD) (19). So, a current hypothesis is that flow-induced changes in [Ca2+]i are disrupted in patients with ADPKD, and this predisposes the kidney to cyst development (see Ref. 19 for more discussion on the pathogenesis of ADPKD). While this is an attractive hypothesis, how bending the PC is linked to opening of the polycystin-1/2 Ca2+ channel complex is not yet understood. Nor is it clear how alterations in Ca2+ signaling lead to the formation of numerous encapsulated, fluid-filled cysts along the nephron, as seen in ADPKD. That said, the intracellular levels of cAMP appear to be integral to the cystic phenotype in both ADPKD and autosomal recessive PKD (ARPKD) (1). Elevated cAMP levels in cystic cells may explain enhanced growth and fluid secretion via activation of the MAP kinase system and cystic fibrosis transmembrane conductance regulator (CFTR) chloride channels, respectively. Current ideas suggest that in normal renal cells, a flow-induced rise in [Ca2+]i leads to a decrease in cAMP levels in two ways: via activation of a Ca2+-sensitive cAMP-dependent phosphodiesterase, as well as by decreasing the activity of a Ca2+-inhibitable adenylate cyclase. Thus, in PKD, increases in [Ca2+]i would not occur and cytosolic cAMP levels would be raised. On the basis of these ideas, V2 receptor antagonists (which were predicted to lower cAMP levels in collecting duct cells) were tested and found to reduce cyst development in murine models of ARPKD (6).In the study by Olteanu et al. (Ref. 13, see p. C952 of this issue), the thinking about the “cystic phenotype” appears to need some radical rethinking. These authors have now shown that defects in the PC are linked to a significant change in the absorptive (not secretory) capacity of cultured renal cortical collecting duct cells derived from the orpk mouse. This mouse has a hypomorphic mutation in the Tg737 gene that encodes the PC-associated protein, polaris (see Ref. 8 for a recent review on murine models of PKD). This protein is required for proper ciliary assembly, and cells lacking polaris possess very short PC. These orpk mice show morphological features similar to those seen in human ARPKD, including enlarged collecting ducts that remain contiguous to the nephron, left-right axis patterning, and skeletal problems. Olteanu and co-workers (13) studied the transepithelial ion transport properties of mutant cortical collecting duct (CCD) principal cells and compared these to similar cells that had been genetically rescued by expression of the normal Tg737 gene. Ion transport was studied after growing the cells on semipermeable supports using classic electrophysiological techniques. The major finding of this work was the unexpected and very marked increase in Na+ absorption in the Tg737-deficient cells, with no effect on Cl−-secretory capacity. The authors used a pharmacological approach to clearly show that this raised Na+ absorption was due primarily to an increase in epithelial Na+ channel (ENaC)-mediated transport. These results were surprising on several counts. First, in ADPKD cells, enhanced/dysregulated Cl− secretion appears to be the general phenotype in the CD and probably involves aberrant stimulation of CFTR (7) through defects in Ca2+ homeostasis. The work of Olteanu et al. (13) shows no such increase in Cl− secretion in their monolayers and thus their data point to a fundamental difference in disease phenotype between ARPKD (or at least in this model of recessive murine cystic disease) and ADPKD. This is actually consistent with work by Nakanishi et al. (11), who found that the presence or absence of CFTR made no difference to the phenotype of bpk mice, another mouse model of ARPKD.Second, recent work from Veizis and Cotton's laboratory (18) showed that Na+ absorption was abnormally regulated by EGF in mouse CD cells derived from bpk mice. In these studies, cystic cells showed a pronounced decrease in ENaC-mediated absorption upon exposure to apical EGF, a result not observed in normal cells and probably explained by the mislocalization of the EGFR to the apical plasma membrane in these mutant cells. Thus a reduction (and not an increase) in Na+ absorption was touted as a possible contributory factor to ARPKD pathophysiology (18). Olteanu et al. (13) were very careful to exclude EGF from their culture media and, in doing so, were able to observe the ENaC-mediated hyperabsorption in their cystic cells. Of course, a key question now is how does this EGF work relate to what happens in human ARPKD? Olteanu et al. (13) argue that encapsulated cysts are not a feature of ARPKD, and thus EGF levels are unlikely to be raised in tubular fluid bathing ARPKD cells, in contrast to the situation in ADPKD. Furthermore, the enhanced Na+ absorption observed by Olteanu et al. (13) would provide a nice explanation for the hypertension that is observed in many ARPKD patients.Regardless of the role of EGF, critical questions that need to be answered are the mechanism underlying the hyperabsorption in cystic cells and the role of the PC in this process. Several possibilities exist, including the following: increased ENaC protein expression, enhanced activation of latent channels by secreted proteases, and altered ENaC regulation via a change in cytosolic/luminal pH (13). In this context, it is interesting that recent patch-clamp studies performed on isolated renal PC suggest that ENaC is functionally expressed in the ciliary membrane itself (16), which thus establishes a direct link between ENaC and the PC. In addition, recent work by Liu et al. (10) showed that flow-induced changes in [Ca2+]i are markedly blunted in the orpk mutant cells (10), a finding similar to what has been reported in cells lacking polycystin 1 or 2 (12). The experiments in the orpk mice used isolated collecting duct tubules and should be a good model of the in vivo situation. Dysregulated Ca2+ signaling therefore could be a contributing factor to the change in ENaC-mediated transport. However, it is not obvious how aberrant flow-mediated Ca2+ signaling can, on the one hand, lead to autonomous, fluid-filled cysts as seen in ADPKD, whereas on the other hand, it produces dilated tubules that hyperabsorb fluid, as suggested for ARPKD (13). To muddy the waters even more, recent [Ca2+]i experiments on microperfused mTAL tubules from mice appear to question the validity of the cilium-polycystin-2 Ca2+ hypothesis (9a). This work shows that flow-induced Ca2+ responses may actually be caused by a mechanically stimulated (stretching) release of ATP, followed by autocrine/paracrine activation of purinergic receptors and IP3-mediated release of Ca2+ from stores. Note, however, that a similar “mechanosensation” mechanism operates in monolayers of Madin-Darby canine kidney cells, but this does not appear to require functional PCs (14). Clearly, similar experiments now need to be repeated in collecting ducts from normal and cystic mice to investigate whether ATP signaling is disrupted in cystic cells and whether differences exist between ARPKD and ADPKD phenotypes. Extracellular ATP (and UTP) is known to downregulate ENaC function acutely in a variety of cells via phosphoinositide metabolites (9), and recently EGF and ATP were shown to inhibit amiloride-sensitive Na+ absorption in CD cells via a MAP kinase pathway (5). It is tempting to speculate that a lack of ATP release could underlie the marked elevation of amiloride-sensitive Na+ transport in the orpk cystic cells. Clearly, the role that the PC may play in nucleotide signaling is going to be a key area of research in the future.Overall, the work of Olteanu et al. (13) has highlighted a novel phenotypic characteristic of ARPKD, which is clearly an important issue for the future understanding of the disease process as well as for the development of new treatments for ARPKD, and possibly for related cystic diseases. An interesting by-product of all this research is that scientists from disparate fields (electrophysiology to molecular genetics, in my case!) have been brought together by this solitary, antenna-like structure. This is something that can only be encouraged, particularly in our current climate of reductionist science.REFERENCES1 Belibi FA, Reif G, Wallace DP, Yamaguchi T, Olsen L, Li H, Helmkamp GM Jr, and Grantham JJ. Cyclic AMP promotes growth and secretion in human polycystic kidney epithelial cells. Kidney Int 66: 964–973, 2004.Crossref | PubMed | ISI | Google Scholar2 Corbit KC, Aanstad P, Singla V, Norman AR, Stainier DY, and Reiter JF. Vertebrate smoothened functions at the primary cilium. 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Am J Physiol Renal Physiol 288: F474–F482, 2005.Link | ISI | Google Scholar19 Wilson PD. Polycystic kidney disease. N Engl J Med 350: 151–164, 2004.Crossref | PubMed | ISI | Google ScholarAUTHOR NOTESAddress for reprint requests and other correspondence: M. A. Gray, Institute for Cell and Molecular Biosciences, School of Biomedical Sciences, Univ. Medical School, Framlington Pl., Newcastle upon Tyne, NE2 4HH, UK (e-mail: [email protected]) Download PDF Back to Top Next FiguresReferencesRelatedInformationCited BySalt-deficient diet exacerbates cystogenesis in ARPKD via epithelial sodium channel (ENaC)EBioMedicine, Vol. 40NEDD4-family E3 ligase dysfunction due to PKHD1/Pkhd1 defects suggests a mechanistic model for ARPKD pathobiology10 August 2017 | Scientific Reports, Vol. 7, No. 1Enhancement of Renal Epithelial Cell Functions through Microfluidic-Based Coculture with Adipose-Derived Stem CellsTissue Engineering Part A, Vol. 19, No. 17-18Epidermal growth factor-mediated proliferation and sodium transport in normal and PKD epithelial cellsBiochimica et Biophysica Acta (BBA) - Molecular Basis of Disease, Vol. 1812, No. 10 More from this issue > Volume 290Issue 4April 2006Pages C947-C949 Copyright & PermissionsCopyright © 2006 the American Physiological Societyhttps://doi.org/10.1152/ajpcell.00640.2005PubMed16531571History Published online 1 April 2006 Published in print 1 April 2006 Metrics