Calmodulin interacts with ATP binding cassette transporter A1 to protect from calpain-mediated degradation and upregulates high-density lipoprotein generation.
OBJECTIVE
To investigate the interaction of ATP-binding cassette transporter A1 (ABCA1) with calmodulin in relation to its calpain-mediated degradation because many calpain substrates bind calmodulin to regulate cellular functions.


RESULTS
The activity of ABCA1 is regulated through proteolysis by calpain. An immunoprecipitation and glutathione S-transferase pull-down assay revealed that ABCA1 directly binds calmodulin in a Ca(2+)-dependent manner. The cytoplasmic loop of ABCA1 contains a typical calmodulin binding sequence of 1-5-8-14 motifs (1245 to 1257 amino acids). The peptide of this region showed binding to calmodulin, and deletion of the 1-5-8-14 motif abolished this interaction. This motif is located near the ABCA1 Pro-Glu-Ser-Thr sequence, and the presence of calmodulin/Ca(2+) protected the peptides from proteolysis by calpain. The knockdown of calmodulin by a specific small and interfering RNA increased the degradation of ABCA1 and decreased ABCA1 protein and apolipoprotein A-I-mediated lipid release. Surprisingly, calmodulin inhibitor W7 increased calmodulin binding to ABCA1 and protected it from calpain-mediated degradation, consistent with our previous finding that this compound increased apolipoprotein A-I-mediated cell cholesterol release.


CONCLUSIONS
Calmodulin directly binds and stabilizes ABCA1 in the presence of Ca(2+) and increases the generation of high-density lipoprotein.
Iwamoto
Noriyuki
N
Biochemistry, Nagoya City University Graduate School of Medical Sciences, Nagoya 467-8601, Japan.
Lu
Rui
R
Tanaka
Nobukiyo
N
Abe-Dohmae
Sumiko
S
Yokoyama
Shinji
S
D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism*; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D016632:Apolipoprotein A-I / Q000378:metabolism; D041702:BALB 3T3 Cells; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D002154:Calpain / Q000378:metabolism*; D006801:Humans; D047468:Immunoprecipitation; D008075:Lipoproteins, HDL / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D025941:Protein Interaction Mapping; D011499:Protein Processing, Post-Translational; D055550:Protein Stability; D034622:RNA Interference; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013449:Sulfonamides / Q000494:pharmacology; D013997:Time Factors; D015854:Up-Regulation
C577836:ABCA1 protein, human; C505693:APOA1 protein, human; D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters; D016632:Apolipoprotein A-I; D002147:Calmodulin; D008075:Lipoproteins, HDL; D011993:Recombinant Fusion Proteins; D013449:Sulfonamides; C017967:W 7; D002154:Calpain; D002118:Calcium