In muscle cells, the excitation-contraction cycle is triggered by an increase in the concentration of free cytoplasmic Ca(2+). The Ca(2+)-ATPase present in the membrane of the sarcoplasmic reticulum (SR) pumps Ca(2+) from the cytosol into this intracellular compartment, thus promoting muscle relaxation. The microsomal fraction derived from the longitudinal smooth muscle of the body wall from the sea cucumber Ludwigothurea grisea retains a membrane-bound Ca(2+)-ATPase that is able to transport Ca(2+) mediated by ATP hydrolysis. Immunological analyses reveal that monoclonal antibodies against sarco-endoplasmic reticulum Ca(2+)-ATPase (SERCA1 and SERCA2a) cross-react with a 110 kDa band, indicating that the sea cucumber Ca(2+)-ATPase is a SERCA-type ATPase. Like the mammalian Ca(2+)-ATPase isoforms so far described, the enzyme also shows a high affinity for Ca(2+) and ATP, has an optimum pH of approximately 7.0 and is sensitive to thapsigargin and cyclopiazonic acid, specific inhibitors of the SERCA pumps. However, unlike the mammalian SERCA isoforms, concentrations of ATP above 2 mmol l(−1) inhibit Ca(2+) transport, but not ATP hydrolysis, in sea cucumber vesicles, suggesting that high ATP concentrations uncouple the Ca(2+)-ATPase. Another unique feature observed with the sea cucumber Ca(2+)-ATPase is the high dependence of maximal activity on K(+) or Na(+). Similar activation promoted by these cations was observed with various mammalian Ca(2+)-ATPase preparations when they were incubated in the presence of low concentrations of sulphated polysaccharides. In control experiments, K(+) and Na(+) have almost no effect on Ca(2+) transport, but in the presence of heparin or fucosylated chondroitin sulphate, the activity of the different mammalian Ca(2+)-ATPases is inhibited and they are activated by either K(+) or Na(+) in a manner similar to the native sea cucumber ATPase. These results led us to investigate the possible occurrence of a highly sulphated polysaccharide on vesicles from the SR of sea cucumber smooth muscle that could act as an ‘endogenous’ Ca(2+)-ATPase inhibitor. In fact, SR vesicles derived from the sea cucumber, but not from rabbit muscle, contain a highly sulphated polysaccharide. After extraction and purification of these polysaccharide molecules, their effect was tested on vesicles obtained from rabbit muscle. This compound inhibited Ca(2+) uptake in rabbit SR vesicles, at concentrations lower than heparin, and restored the dependence on monovalent cations. These results strongly suggest that the sea cucumber Ca(2+)-ATPase is activated by monovalent cations because of the presence of endogenous sulphated polysaccharides.

REFERENCES

Brandl
C. J.
,
de Leon
S.
,
Martin
D. R.
,
MacLennan
D. H.
(
1987
).
Adult forms of the Ca2+-ATPase of sarcoplasmic reticulum.
J. Biol. Chem
262
,
3768
–.
Burk
S. E.
,
Lytton
J.
,
MacLennan
D. H.
,
Shull
G. E.
(
1989
).
cDNA cloning, functional expression and mRNA tissue distribution of a third organellar Ca2+pump.
J. Biol. Chem
264
,
18
–.
Cario
C.
,
Malavar
L.
,
Hernandez-Nicase
M. L.
(
1996
).
Two distinct distribution patterns of sarcoplasmic reticulum in two functionally different giant smooth muscle cells of Beroeovata.
Cell Tissue Res
282
,
435
–.
Castellani
L.
,
Harwicke
P. M. D.
,
Franzini-Armstrong
C.
(
1989
).
Effect of Ca2+on the dimeric structure of scallop sarcoplasmic reticulum.
J. Cell Biol
108
,
511
–.
Chaloub
R. M.
,
de Meis
L.
(
1980
).
Effect of K+on phosphorylation of the sarcoplasmic reticulum ATPase by either Pior ATP.
J. Biol. Chem
255
,
6168
–.
Champeil
P.
,
Henao
F.
,
de Foresta
B.
(
1997
).
Dissociation of Ca2+from sarcoplasmic reticulum Ca2+-ATPase and changes in fluorescence of optically selected Trp residues. Effect of KCl and NaCl and implications for substeps in Ca2+dissociation.
Biochemistry
36
,
12383
–.
Chen
C.
(
1986
).
Contractions of holothurian longitudinal body wall muscle.
Chin. J. Physiol
29
,
311
–.
de Meis
L.
(
1971
).
Allosteric inhibition by alkali ions of the Ca2+uptake and adenosina triphosphatase activity of skeletal muscle microsomes.
J. Biol. Chem
246
,
4764
–.
de Meis
L.
,
Hasselbach
W.
(
1971
).
Acetyl phosphate as920substrate for Ca2+uptake in skeletal muscle microsomes.
J. Biol. Chem
246
,
4759
–.
de Meis
L.
,
Hasselbach
W.
,
Machado
R. D.
(
1974
).
Characterization of calcium oxalate and calcium phosphate deposits in sarcoplasmic reticulum vesicles.
J. Cell Biol
62
,
505
–.
de Meis
L.
,
Sorenson
M. M.
(
1989
).
ATP regulation of calcium transport in back inhibited sarcoplasmic reticulum vesicles.
Biochim. Biophys. Acta
984
,
373
–.
de Meis
L.
,
Suzano
V.
(
1994
).
Uncoupling of muscle and blood platelets Ca2+-transport ATPases by heparin.
J. Biol. Chem
269
,
14525
–.
de Meis
L.
,
Vianna
A. L.
(
1979
).
Energy interconvertion by the Ca2+transport ATPase of sarcoplasmic reticulum.
Annu. Rev. Biochem
48
,
275
–.
Devlin
C. L.
(
1993
).
Acetylcholine-induced contractions in a holothurian (Isostichopus badionotus) smooth muscle are blocked by the calcium antagonists, diltiazem and verapamil.
Comp. Biochem. Physiol
106
,
573
–.
Duggan
P. F.
(
1977
).
Calcium uptake and associated adenosine triphosphatase activity in fragmented sarcoplasmic reticulum.
J. Biol. Chem
252
,
1620
–.
Engelender
S.
,
de Meis
L.
(
1996
).
Pharmacological differentiation between intracellular calcium pumps isoforms.
Mol. Pharmacol
50
,
1243
–.
Engelender
S.
,
Wolosker
H.
,
de Meis
L.
(
1995
).
The Ca2+-ATPase isoforms of platelets are located in distinct functional Ca2+pools and are uncoupled by a mechanism different from thatof skeletal muscle Ca2+-ATPase.
J. Biol. Chem
270
,
21050
–.
Eylers
J. P.
(
1982
).
Ion-dependent viscosity of holothurian body wall and its implications for the functional morphology of echinoderms.
J. Exp. Biol
99
,
1
–.
Hill
R. B.
,
Sanger
J. W.
,
Yantorno
R. E.
,
Deutsch
C.
(
1978
).
Contraction in a muscle with negligible sarcoplasmic reticulum: The longitudinal retractor of the sea cucumber Isostichopus badionotus (Selenka), holothuria aspidochirota.
J. Exp. Zool
206
,
137
–.
Inesi
G.
(
1985
).
Mechanism of Ca2+transport.
Annu. Rev. Physiol
47
,
573
–.
Kalabokis
V. N.
,
Bozzola
J. J.
,
Castellani
L.
,
Harwicke
P. M. D.
(
1991
).
A possible role for the dimmer ribbon state of scallop sarcoplasmic reticulum.
J. Biol. Chem
266
,
22044
–.
Landeira-Fernandez
A. M.
,
Aiello
K. R. M.
,
Aquino
R. S. A.
,
Silva
L. C. F.
,
de Meis
L.
,
Mourão
P. A. S.
(
2000
).
A sulphated polysaccharide from the sarcoplasmic reticulum of sea cucumber smooth muscle is an endogenous inhibitor of the Ca2+-ATPase.
Glycobiol
10
,
773
–.
Landeira-Fernandez
A. M.
,
Costa
M. S.
,
de Meis
L.
(
1996
).
Modulation of maize roots H+-ATPase by sulphated polysaccharides.
Biosci. Rep
16
,
439
–.
Landeira-Fernandez
A. M.
,
Galina
A.
,
Jennings
P.
,
Montero-Lommeli
M.
,
de Meis
L.
(
2000
).
Sarcoplasmic reticulum Ca2+-ATPase of sea cucumber smooth muscle: Regulation by K+and ATP.
Comp. Biochem. Physiol
126
,
263
–.
Lehman
W.
,
Kendrick-Jones
J.
,
Szent-György
A.G.
(
1973
).
Myosin-linked regulatory systems: Comparative studies.
Cold Spring Harbor Symp. Quant. Biol
37
,
319
–.
Lytton
J.
,
MacLennan
D. H.
(
1988
).
Molecular cloning of cDNAs from human kidney coding for two alternatively spliced products of the cardiac Ca2+-ATPase gen.
J. Biol. Chem
263
,
15024
–.
Lytton
J.
,
Westlin
M.
,
Burk
S. E.
,
Shull
G. E.
,
MacLennan
D. H.
(
1992
).
Functional comparisons between isoforms of the sarcoplasmic reticulum family of calcium pumps.
J. Biol. Chem
267
,
14483
–.
Lytton
J.
,
Westlin
M.
,
Hanley
M. R.
(
1991
).
Thapsigargin inhibits the sarcoplasmic or endoplasmic reticulum Ca2+-ATPase family of Ca2+pumps.
J. Biol. Chem
266
,
17067
–.
Lytton
J.
,
Zarain-Herzberg
A.
,
Periasamy
M.
,
MacLennan
D. H.
(
1989
).
Molecular cloning of the mammalian smooth muscle sarco(endo)plasmic reticulum Ca2+-ATPase.
J. Biol. Chem
264
,
7059
–.
MacLennan
D. H.
(
1985
).
Amino-acid sequence of a Ca2+-Mg2+dependent ATPase from rabbit muscle sarcoplasmic reticulum, deduced from its complementary DNA sequence.
Nature
316
,
696
–.
Mattai
J.
,
Kwak
J. C. T.
(
1981
).
Mg and Ca binding to heparin in the presence of added univalent salt.
Biochim. Biophys. Acta
276
,
303
–.
Meissner
G.
(
1994
).
Ryanodine receptor/Ca2+release channels and their regulation by endogenous effectors.
Annu. Rev. Physiol
56
,
485
–.
Mintz
E.
,
Guillain
F.
(
1997
).
Ca2+transport by the sarcoplasmic reticulum ATPase.
Biochim. Biophys. Acta
1318
,
52
–.
Mitidieri
F.
,
de Meis
L.
(
1999
).
Ca2+release and heat production by the endoplasmic reticulum Ca2+-ATPase of blood platelets.
J. Biol. Chem
274
,
28344
–.
Morris
S. C.
(
1993
).
The fossil record and the early evolution of the metazoa.
Nature
361
,
219
–.
Motokawa
T.
(
1982
).
Factors regulating the mechanical properties of holothurian dermis.
J. Exp. Biol
99
,
29
–.
Motokawa
T.
(
1984
).
Connective tissue catch in echinoderms.
Biol. Rev
59
,
255
–.
Mourão
P. A. S.
,
Pereira
M. S.
,
Pavão
M. S. G.
,
Mulloy
B.
,
Tollefsen
D. M.
,
Mowinkel
M. C.
,
Abildgaard
U.
(
1996
).
Structure and anticoagulant activity of a fucosylated chondroitin sulphate from echinoderm.
J. Biol. Chem
271
,
23973
–.
Moutin
M.-J.
,
Dupont
Y.
(
1991
).
Interaction of potassium and magnesium with the high affinity calcium binding sites of the sarcoplasmic reticulum calcium-ATPase.
J. Biol. Chem
266
,
5580
–.
Nolan
D. P.
,
Reverald
P.
,
Pays
E.
(
1994
).
Overexpression and characterization of a gene for a Ca2+-ATPase of the endoplasmic reticulum in Trypanosoma brucei.
J. Biol. Chem
269
,
26045
–.
Ozawa
T.
(
1999
).
Ryanodine-sensitive Ca2+release mechanism of a rat prancreatic acinar cells is modulated by calmodulin.
Biochim. Biophys. Acta
1452
,
254
–.
Palmero
I.
,
Sastre
L.
(
1989
).
Complementary DNA cloning of a protein highly homologous to mammalian sarcoplasmic reticulum Ca2+-ATPase from the crustacean Artemia.
J. Mol. Biol
210
,
737
–.
Pedersen
P. L.
,
Carafoli
E.
(
1987
).
Ion Motive ATPases. I. Ubiquity, properties and significance to cell function.
Trends Biochem. Sci
12
,
146
–.
Pedersen
P. L.
,
Carafoli
E.
(
1987
).
Ion Motive ATPases. II. Energy coupling and work output.
Trends Biochem. Sci
12
,
186
–.
Pick
U.
(
1982
).
The interaction of vanadate ions with the Ca2+-ATPase from sarcoplasmic reticulum.
J. Biol. Chem
257
,
6111
–.
Prosser
C. L.
,
Mackie
G. O.
(
1980
).
Contractions of holothurian muscles
.
J. Comp. Physiol
.
136
,
103
112
.
Ribeiro
J. M. C.
,
Vianna
L. A.
(
1978
).
Allosteric modification by K+of the (Ca2+Mg2+)-dependent ATPase of sarcoplasmic reticulum.
J. Biol. Chem
253
,
3153
3157
.
Rocha
J. B. T.
,
Landeira-Fernandez
A. M.
,
de Meis
L.
(
1998
).
Modification of the pH dependence of animal and plant transport ATPases by sulphated polysaccharides.
Biochem. Biophys. Res. Commun
244
,
720
–.
Rocha
J. B. T.
,
Wolosker
H.
,
Souza
D. S.
,
de Meis
L.
(
1996
).
Alteration of Ca2+fluxes in brain microsomes by K+and Na+: Modulation by sulphated polysaccharides and trifluoperazine.
J. Neurochem
66
,
772
–.
Ruggiero
J.
,
Vieira
R. P.
,
Mourão
P. A. S.
(
1994
).
Increased calcium affinity of a fucosylated chondroitin sulphate from sea cucumber.
Carbohydr. Res
256
,
275
–.
Shigekawa
M.
,
Pearl
L. J.
(
1976
).
Activation of calcium transport in skeletal muscle sarcoplasmic reticulum by monovalent cations.
J. Biol. Chem
251
,
6947
–.
Suzuki
S.
(
1982
).
Physiological and cytochemical studies on activator calcium in contraction by smooth muscle of a sea cucumber, Isotichopus badionotus.
Cell Tissue Res
222
,
11
–.
Trotter
J. A.
,
Koob
T. J.
(
1989
).
Collagen and proteoglycan in sea urchin ligament with mutable mechanical properties.
Cell Tissue Res
258
,
527
–.
Trotter
J. A.
,
Lyons-Levy
G.
,
Thurmond
F. A.
,
Kroob
T. J.
(
1995
).
Covalent composition of collagen fibrils from the dermis of the sea cucumber, Cucumaria frondosa, a tissue with mutable mechanical properties.
Comp. Biochem. Physiol
112
,
463
–.
Wolosker
H.
,
Rocha
J. B. T.
,
Engelender
S.
,
Panizzutti
R.
,
de Miranda
J.
,
de Meis
L.
(
1997
).
Sarco/endoplasmic reticulum Ca2+-ATPase isoforms: Diverse responses to acidosis.
Biochem. J
321
,
545
–.
Xu
L.
,
Mann
G.
,
Meissner
G.
(
1996
).
Regulation of cardiac Ca2+release channel (ryanodine receptor) by Ca2+, H+, Mg2+and adenine nucleotides under normal and simulated ischemic conditions.
Circ. Res
79
,
1100
–.
Xu
L.
,
Tripathy
A.
,
Pasek
D. A.
,
Meissner
G.
(
1999
).
Ruthenium red modifies the cardiac and skeletal muscle Ca2+release channels. Regulation of cardiac Ca2+release channel (ryanodine receptors) by multiple mechanisms.
J. Biol. Chem
274
,
32680
–.
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