Ankyrin-B Regulates Cav2.1 and Cav2.2 Channel Expression and Targeting

• Published in: The Journal of biological chemistry Vol. 289; no. 8; pp. 5285 - 5295
• Main Authors: Kline, Crystal F., Scott, John, Curran, Jerry, Hund, Thomas J., Mohler, Peter J.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 21-02-2014; American Society for Biochemistry and Molecular Biology
• Subjects: Adaptor Proteins; Amino Acid Motifs; Amino Acid Sequence; Animals; Ankyrins - metabolism; Brain; Calcium Channels, N-Type - chemistry; Calcium Channels, N-Type - metabolism; Conserved Sequence; Cytoskeleton; HEK293 Cells; Humans; Immunoprecipitation; Ion Channels; Membrane Biology; Membrane Trafficking; Mice; Mice, Inbred C57BL; Mice, Knockout; Molecular Sequence Data; Protein Binding; Protein Sorting; Protein Targeting; Purkinje Cells - metabolism; Rats; Tyrosine - metabolism; Membrane Trafficking; Protein Sorting; Cytoskeleton; Ion Channels; Adaptor Proteins; Protein Targeting
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M113.523639

N-type and P/Q-type calcium channels are documented players in the regulation of synaptic function; however, the mechanisms underlying their expression and cellular targeting are poorly understood. Ankyrin polypeptides are essential for normal integral membrane protein expression in a number of cell types, including neurons, cardiomyocytes, epithelia, secretory cells, and erythrocytes. Ankyrin dysfunction has been linked to defects in integral protein expression, abnormal cellular function, and disease. Here, we demonstrate that ankyrin-B associates with Cav2.1 and Cav2.2 in cortex, cerebellum, and brain stem. Additionally, using in vitro and in vivo techniques, we demonstrate that ankyrin-B, via its membrane-binding domain, associates with a highly conserved motif in the DII/III loop domain of Cav2.1 and Cav2.2. Further, we demonstrate that this domain is necessary for proper targeting of Cav2.1 and Cav2.2 in a heterologous system. Finally, we demonstrate that mutation of a single conserved tyrosine residue in the ankyrin-binding motif of both Cav2.1 (Y797E) and Cav2.2 (Y788E) results in loss of association with ankyrin-B in vitro and in vivo. Collectively, our findings identify an interaction between ankyrin-B and both Cav2.1 and Cav2.2 at the amino acid level that is necessary for proper Cav2.1 and Cav2.2 targeting in vivo.
Calcium channels control membrane excitability. The mechanisms underlying Cav2.1/Cav2.2 targeting are not well understood.
Ankyrin-B associates with Cav2.1/Cav2.2. Loss of ankyrin-B results in reduced expression of Cav2.1/Cav2.2 in select brain regions.
Ankyrin-B plays a role in the expression of Cav2.1/Cav2.2.
Results identify pathway for membrane targeting of calcium channels and regulation of membrane excitability.