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| 2: J Cell Sci. 2003 Jun 15;116(Pt 12):2483-94. |
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- Biochemical characterization, distribution and
phylogenetic analysis of Drosophila melanogaster ryanodine and IP3 receptors,
and thapsigargin-sensitive Ca2+ ATPase.
Vazquez-Martinez O, Canedo-Merino R, Diaz-Munoz M, Riesgo-Escovar JR.
Department of Molecular and Cellular Neurobiology, Neurobiology Institute,
Campus UNAM-Juriquilla, Universidad Nacional Autonoma de Mexico, Queretaro
76230, Mexico.
We characterized the biochemistry, distribution and phylogeny of Drosophila
ryanodine (RyR) and inositol triphosphate (IP3R) receptors and the endoplasmic
reticulum Ca2+-ATPase (SERCA) by using binding and enzymatic assays, confocal
microscopy and amino acid sequence analysis. [3H]-ryanodine binding in total
membranes was enhanced by AMP-PCP, caffeine and xanthine, whereas Mg2+,
Ruthenium Red and dantrolene were inhibitors. [3H]-ryanodine binding showed a
bell-shaped curve with increasing free [Ca2+], without complete inhibition at
millimolar levels of [Ca2+]. [3H]-IP3 binding was inhibited by heparin, 2-APB
and xestospongin C. Microsomal Ca2+-ATPase activity was inhibited by
thapsigargin. Confocal microscopy demonstrated abundant expression of
ryanodine and inositol triphosphate receptors and abundant Ca2+-ATPase in
Drosophila embryos and adults. Ryanodine receptor was expressed mainly in the
digestive tract and parts of the nervous system. Maximum parsimony and
Neighbour Joining were used to generate a phylogenetic classification of
Drosophila ryanodine and insitol triphosphate receptors and Ca2+-ATPase based
on 48 invertebrate and vertebrate complete sequences. The consensus trees
indicated that Drosophila proteins grouped with proteins from other
invertebrates, separately from vertebrate counterparts. Despite evolutionary
distances, our functional results demonstrate that Drosophila ryanodine and
inositol triphosphate receptors and Ca2+-ATPase are reasonably similar to
vertebrate counterparts. Our protein expression data are consistent with the
known functions of these proteins in the Drosophila digestive tract and
nervous system. Overall, results show Drosophila as a valuable tool for
intracellular Ca2+ dynamics studies in eukaryotes.
PMID: 12766186 [PubMed - in process]
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