Cloning of a novel alpha 1-subunit of the voltage-dependent calcium channel from the beta-cell.
To study the molecular regulation of voltage-dependent Ca2+ channels (VDCCs) in the beta-cell, we have cloned a cDNA for the alpha 1-subunit from a hamster insulin-secreting cell line (HIT-T15). The cDNA (HCa3a) encodes a 1610-amino acid protein with four repeating membrane domains and an overall structure characteristic of other alpha 1-subunits. Although the cDNA shows a high degree of sequence homology (97%) with a rat brain alpha 1-subunit (RB alpha 1), the C-terminal 15 amino acids of HCa3a share no similarity with any cloned alpha 1 protein. High stringency Northern blot analysis revealed a single transcript of approximately 8.6 kilobases in HIT cells and hamster pancreas. A similarly sized species was detected in hamster brain, heart, and skeletal muscle. Using polymerase chain reaction and a primer set unique to HCa3a, this alpha 1 isoform was found to be expressed in islet cell lines derived from rat, mouse, and hamster. The HIT cell alpha 1-subunit is also expressed in discrete regions of the rat central nervous system, including the cortex, cerebellum, hypothalamus, and brain stem. The expression of two alpha 1 isoforms (HCa3a and cardiac) in the HIT cell underscores the possible complexity of VDCCs in the regulation of beta-cell signal transduction. With its widespread tissue distribution, HCa3a does not conform to the current classification system used for L-type VDCCs; this suggests that an alternative system of classification is required.
Yaney
G C
GC
Department of Medicine, Baylor College of Medicine, Houston, Texas 77030.
Wheeler
M B
MB
Wei
X
X
Perez-Reyes
E
E
Birnbaumer
L
L
Boyd
A E
AE
Moss
L G
LG
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001923:Brain Chemistry; D015220:Calcium Channels / Q000235:genetics*; D006224:Cricetinae; D004247:DNA / Q000235:genetics; D007340:Insulinoma; D007515:Islets of Langerhans / Q000378:metabolism*; D008647:Mesocricetus; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000235:genetics; D009419:Nerve Tissue Proteins / Q000235:genetics; D009928:Organ Specificity; D016133:Polymerase Chain Reaction; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014407:Tumor Cells, Cultured
D015220:Calcium Channels; D009124:Muscle Proteins; D009419:Nerve Tissue Proteins; D004247:DNA