Deletion of NH2− and COOH‐terminal sequences destroys function of the Ca2+ ATPase of rabbit fast‐twitch skeletal muscle sarcoplasmic reticulum

Skerjanc, Ilona S.; Clarke, David M.; Loo, Tip W.; MacLennan, David H.

doi:10.1016/0014-5793(93)81633-b

Cited by 18 publications

(8 citation statements)

References 17 publications

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“…Deletion of the N-terminal domain of SERCA1 reduced protein expression (13). Accordingly, mutation of SERCA2b Asn-39 to Asp or Thr reduced protein expression, suggesting that the integrity of the N terminus is important for pump expression.…”

Section: Protein Levels Of Serca2bmentioning

confidence: 99%

Multiple Effects of SERCA2b Mutations Associated with Darier's Disease

Ahn

Lee

Kim

et al. 2003

Journal of Biological Chemistry

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Darier's disease (DD) is an autosomal dominant disorder caused by mutations in the ATP2A2 gene, encoding sarco/endoplasmic reticulum Ca 2؉ -ATPase pump type 2b isoform (SERCA2b). Although >100 mutations in the ATP2A2 gene were identified, no apparent relation between genotype/phenotype emerged. In this work, we analyzed 12 DD-associated mutations from all of the regions of SERCA2b to study the underlying pathologic mechanism of DD and to elucidate the role of dimerization in SERCA2b activity. Most mutations markedly affected protein expression, partially because of enhanced proteasome-mediated degradation. All of the mutants showed lower activity than the wild type pump. Notably, several mutants that cause relatively severe phenotype of DD inhibited the activity of the endogenous and the co-expressed wild type SERCA2b. Importantly, these effects were not attributed to changes in passive Ca 2؉ leak, inositol 1,4,5-trisphosphate receptor activity, or sensitivity to inositol 1,4,5-trisphosphate. Rather, co-immunoprecipitation experiments showed that SERCA2b monomers interact to influence the activity of each other. These findings reveal multiple molecular mechanisms to account for the plethora of pathologic states observed in DD and provide the first evidence for the importance of SERCA2b dimerization in pump function in vivo.Darier's disease (DD, 1 OMIM 124200), also known as keratosis follicularis, is an autosomal dominant skin disorder characterized by loss of adhesion between epidermal cells (acantholysis) and abnormal keratinization (1). DD is also associated with an increased prevalence of neuropsychiatric disorders including bipolar disorders, schizophrenia, epilepsy, and mental retardation (1). Recently, mutations in the ATP2A2 gene encoding the sarco/endoplasmic reticulum Ca 2ϩ -ATPase pump type 2b isoform (SERCA2b) have been identified as a cause of DD (2). SERCA2b is a housekeeping protein, which is expressed ubiquitously and is responsible for Ca 2ϩ uptake into the ER, including the agonist-mobilizable Ca 2ϩ pool (3, 4). By determining ER Ca 2ϩ load, the SERCA2 pumps modulate several parameters of the Ca 2ϩ signal. In resting cells, the SERCA2 pump provides the bulk of the dynamic Ca 2ϩ -buffering capacity to prevent large fluctuation in [Ca 2ϩ ] i (5). In stimulated cells, SERCA2 controls the activity of the store-operated Ca 2ϩ influx channel by preventing accumulation of Ca 2ϩ at the mouth of the channels and their inhibition by [Ca 2ϩ ] i (6). SERCA2b reloads the ER with Ca 2ϩ at the end of cell stimulation (5) and between Ca 2ϩ spikes to determine the frequency of Ca 2ϩ oscillations (7), and to control the diverse cellular functions regulated by [Ca 2ϩ ] i and Ca 2ϩ oscillations (8, 9). Alteration in SERCA2 pump function by overexpression in Chinese hamster ovary cells (10) or partial deletion in mice (7) lead to adaptation of the Ca 2ϩ signaling machinery and Ca 2ϩ -regulated cell functions that translates to nearly normal physiological response in the majority of organs and tissues. This co...

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Section: Protein Levels Of Serca2bmentioning

confidence: 99%

Multiple Effects of SERCA2b Mutations Associated with Darier's Disease

Ahn

Lee

Kim

et al. 2003

Journal of Biological Chemistry

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“…Tryptic digestion of this ATPase provided experimental evidence for the first eight transmembrane segments that had been predicted, but an unexpected cleavage between H9 and H10 was observed (12). Mutagenesis to generate Ca 2ϩ transport mutants provided evidence that H4, H5, H6, and H8 were in the membrane domain (15)(16)(17)(18). The crystal structure of the SR Ca 2ϩ -ATPase at 14-Å resolution shows that the membrane domain can accommodate 10 membrane-spanning segments (23).…”

Section: ؉mentioning

confidence: 99%

The Membrane Topology of the Rat Sarcoplasmic and Endoplasmic Reticulum Calcium ATPases by in Vitro Translation Scanning

Bayle

Weeks

Sachs

1995

Journal of Biological Chemistry

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The membrane topology of the rat endoplasmic reticulum (ER) and sarcoplasmic reticulum (SR) Ca 2؉ATPases were investigated using in vitro transcription/ translation of fusion vectors containing DNA sequences encoding putative membrane-spanning domains. The sequences of these Ca A construct containing the fifth predicted transmembrane segment was able to act only as a stop transfer sequence. The sixth transmembrane segment did not insert cotranslationally into the membrane. The seventh was able to act as both a signal anchor and stop transfer sequence, and the eighth showed stop transfer ability in the M1 vector. The ninth transmembrane segment had both signal anchor and stop transfer capacity, whereas the tenth transmembrane segment showed only stop transfer sequence properties. The eleventh transmembrane sequence, unique to the ER Ca 2؉ ATPase, had both signal anchor and stop transfer properties. These translation data provide direct experimental evidence for 8 or 9 of the 10 or 11 predicted transmembrane sequences in the current topological models for the SR or ER Ca 2؉ ATPases, respectively.

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“…These results show that the amino acid residues in the Ala 3 -Ser 6 region are not essential for Ca 2ϩ transport function. Previously, Skerjanc et al (16) reported that there is no indication from site-directed mutagenesis that specific residues in the Glu 2 -His 32 region are crucial for enzymatic activity. This is consistent with our above conclusion.…”

Section: Effects Of Deletions and Substitutions Of Residues In The Nhmentioning

confidence: 99%

“…It was previously shown (16) that deletion of most of the residues (Glu 2 -His 32 ) in the NH 2 -terminal domain results in greatly reduced expression in COS-1 cells and inactivation of the enzyme. This raises the possibility that the NH 2 -terminal domain has a region sensitive to the endoplasmic reticulum (ER)-mediated quality control, the machinery of which recognizes and rapidly degrades misfolded proteins (this misfolding can be induced by mutations) and denatured proteins to suppress their cellular expression or accumulation (17,18).…”

mentioning

confidence: 99%