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Identity of Lysosomal NAADP‐Sensitive Ca2+ Release Channels is TRP‐ML1 in Coronary Arterial Smooth Muscle Cells
Author(s) -
Zhang Fan,
Yi Fan,
Chen QiZheng,
Dobson Crystal S.,
Li PinLan
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.5.a548-a
Subject(s) - transient receptor potential channel , lysosome , chemistry , microbiology and biotechnology , transfection , biophysics , biochemistry , receptor , biology , gene , enzyme
Recently, we characterized lysosomal nicotinic acid adenine dinucleotide phosphate (NAADP)‐sensitive Ca 2+ release channels in coronary arterial smooth muscle cells (CASMCs). The present study was designed to further confirm the identity of these lysosomal Ca 2+ release channels that possibly relate to transient receptor potential‐mucolipin‐1 (TRP‐ML1) protein, a non‐selective Ca 2+ permeable cation channel. Western blot analysis demonstrated the existence of TRP‐ML1 protein in purified lysosomes from CASMCs. By lipid bilayer reconstitution of these purified CASMCs lysosomes, this NAADP‐activated channel activity in TRP‐ML1‐deprived lysosomal preparations was significantly reduced. In intact CASMCs, endothelin‐1 (a lysosomal Ca 2+ release agonist) induced a large lysosome‐associated Ca 2+ release, which was significantly attenuated by transfection of TRP‐ML1 siRNA in the cells. However, oxotremorine‐induced Ca 2+ release in CASMCs was not altered by TRP‐ML1 siRNA. In addition, ultrasound microbubble introduction of NAADP into TRP‐ML −/− human fibroblastcells did not produce any Ca 2+ release response. However, when these TRP‐ML −/− cells were transfected with a full‐length TRP‐ML1 gene, NAADP‐induced Ca 2+ release response was restored. We conclude that lysosomal TRP‐ML1 is functioning as NAADP‐sensitive Ca 2+ release channels (Supported by NIH grants HL057244‐09 and HL075316‐01).