Calcium Pumps and Keratinocytes: Lessons From Darier's Disease and Hailey-Hailey Disease

J. Dhitavat; R.J. Fairclough; A. Hovnanian; S.M. Burge


The British Journal of Dermatology. 2004;150(5) 

In This Article

Calcium Pumps and Desmosome Adhesion

The formation and stability of desmosomes depend on interactions between transmembrane desmosomal cadherins (desmogleins, desmocollins), submembranous plaque proteins (plakoglobin, plakophilin, desmoplakins) and the cytokeratin filaments. Adhesion is mediated by Ca2+-dependent interactions between the extracellular domains of desmosomal cadherins, but the adhesion function of cadherins is moderated by complex interactions between the cytoplasmic domains of cadherins and plaque proteins such as plakoglobin. Interactions between other plaque proteins (plakophilin and desmoplakin) and the cytokeratin filaments both stabilize desmosomes and indirectly link the desmosomal cadherins to the keratin intermediate filament network.[51,52]

The assembly of desmosomes has been studied in vitro in the presence of both high and low levels of extracellular Ca2+. In low Ca2+ conditions (< 0.1 mmol L-1), desmosomes are not assembled, but constituent proteins continue to be synthesized and stored in the endoplasmic reticulum where they are folded and processed prior to transport to the cell membrane. Following a Ca2+ shift to physiological levels (1.2 mmol L-1), desmosomal proteins are gradually targeted to the cell membrane, where they form mature desmosomes.[53] Studies of desmosome formation in Madin-Darby canine kidney cells have shown that the trafficking of desmocollins to the cell surface is followed by the transport of desmogleins, plakoglobin and finally by desmoplakins.[54]

The translation, translocation, folding, processing and maturation of secreted proteins are regulated by the levels of luminal Ca2+ and Mn2+ in the endoplasmic reticulum and the Golgi apparatus[55] (Fig. 6). In keratinocytes, intraluminal Ca2+ levels in the endoplasmic reticulum are maintained by SERCA2b, whereas SPCA1 pumps mediate Mn2+ transport as well as Ca2+ into the Golgi apparatus.[56,57] These pumps also play a crucial role in setting up the cytosolic Ca2+ oscillations known to control many vital cell functions, through cycles of release and re-uptake of internal Ca2+ stores.[47,58]

Mutations in ATP2A2 affect the expression, degradation and activity of SERCA pumps. The SERCA pumps were inactive in 11 Darier's disease-associated mutations and were reduced by 50% in one mutation.[59] We studied four mutations: two found in segmental Darier's disease (unpublished data), one from acrokeratosis verruciformis and one from classical Darier's disease. In each case, the mutation resulted in complete loss of Ca2+ transport activity.[20] Most mutations markedly reduce protein expression by enhancing proteasome-mediated degradation.[59] Ahn et al. found that some mutants reduced the activity of wild-type SERCA2b pumps. Co-immunoprecipitation experiments indicated that interactions between SERCA2b monomers may influence pump activity, suggesting that SERCA2b dimerization may be important for pump function in vivo.[59]

Thapsigargin is a specific and potent inhibitor of all SERCA pumps. Early studies in keratinocytes suggested that inhibition of SERCA with thapsigargin disturbed the 'structural organization' of desmoplakin as well as that of involucrin (a marker of differentiation).[60] The authors suggested that thapsigargin might interfere with intracellular transport.[60] In Madin-Darby canine kidney epithelial cells, inhibition of SERCA with thapsigargin disrupts the assembly of both desmosomes and tight junctions. The sorting of desmoplakin (the only desmosomal component tracked in this study) to the plasma membrane was delayed and desmosomes were unstable, because plaque proteins did not associate appropriately with the cytoskeleton.[61] We used thapsigargin to inhibit all SERCA activity in normal keratinocytes and showed that the sorting of desmosomal cadherins as well as desmoplakins was impaired. The desmosomal proteins were retained in the endoplasmic reticulum as insoluble aggregates, probably secondary to misfolding.[62,63]

We also studied the sorting of the major desmosomal components in Darier's disease keratinocytes. Despite reduced SERCA2 activity, the desmosomal cadherins (desmoglein and desmocollin) were still efficiently transported to the cell surface, but desmoplakin formed insoluble aggregates. These results suggest that the key to acantholysis and dyskeratosis in Darier's disease may lie in the influence of SERCA2 on the trafficking of desmoplakin to the desmosomal plaque.[64]

The function of the human secretory pathway Ca2+/Mn2+-ATPase, SPCA1, is more difficult to study, because no inhibitors of pump activity have been identified. We have provided evidence that the ATP2C1 mutations found in Hailey-Hailey disease result in specific defects in Ca2+ and/or Mn2+ transport and protein expression.[57] The regulation of cytoplasmic Ca2+ is impaired in cultured keratinocytes from Hailey-Hailey disease and the normal epidermal Ca2+ gradient is attenuated in vivo in Hailey-Hailey disease.[13] We also know that the concentration of cellular ATP is reduced in cultured Hailey-Hailey disease keratinocytes and that this may lead to impairment of the Ca2+-induced reorganization of actin filaments.[65] Mutations in ATP2C1 may alter Ca2+ signalling in keratinocytes in Hailey-Hailey disease: HeLa cells lacking SPCA1 have less frequent baseline Ca2+ oscillations than control cells, suggesting that SPCA1 plays a part in shaping the cytosolic Ca2+ signals in these epithelial cells.[47] Studies of the yeast SPCA1 homologue (PMR1) have shown that protein sorting and glycosylation are impaired in null mutants.[55,56,66,67] By analogy, reduced SPCA1 activity in Hailey-Hailey disease might affect Ca2+ signalling in keratinocytes, altering the sorting of desmosomal proteins or their glycosylation.