Antibodies specific for distinct Kv subunits unveil a heterooligomeric basis for subtypes of alpha-dendrotoxin-sensitive K+ channels in bovine brain.
The authentic subunit compositions of neuronal K+ channels purified from bovine brain were analyzed using a monoclonal antibody (mAb 5), reactive exclusively with the Kv1.2 subunit of the latter and polyclonal antibodies specific for fusion proteins containing C-terminal regions of four mammalian Kv proteins. Western blotting of the K+ channels isolated from several brain regions, employing the selective blocker alpha-dendrotoxin (alpha-DTX), revealed the presence in each of four different Kvs. Variable amounts of Kv1.1 and 1.4 subunits were observed in the K+ channels purified from cerebellum, corpus striatum, hippocampus, cerebral cortex, and brain stem; on the other hand, contents of Kv1.6 and 1.2 subunits appeared uniform throughout. Each Kv-specific antibody precipitated a different proportion (anti-Kv1.2 > 1.1 >> 1.6 > 1.4) of the channels detectable with radioiodinated alpha-DTX in every brain region, consistent with a widespread distribution of these oligomeric subtypes. Such heterooligomeric combinations were further documented by the lack of additivity upon their precipitation with a mixture of antibodies to Kv1.1 and Kv1.2; moreover, cross-blotting of the multimers precipitated by mAb 5 showed that they contain all four Kv proteins. Collectively, these findings demonstrate that subtypes of alpha-DTX-susceptible K+ channels are prevalent throughout mammalian brain which are composed of different Kv proteins assembled in complexes, shown previously to also contain auxiliary beta-subunits [Parcej, D. N., Scott, V. E. S., & Dolly, J.O. (1992) Biochemistry 31, 11084-11088].
Scott
V E
VE
Department of Biochemistry, Imperial College, London, U.K.
Muniz
Z M
ZM
Sewing
S
S
Lichtinghagen
R
R
Parcej
D N
DN
Pongs
O
O
Dolly
J O
JO
D000818:Animals; D000911:Antibodies, Monoclonal; D001483:Base Sequence; D015153:Blotting, Western; D001923:Brain Chemistry; D001933:Brain Stem / Q000737:chemistry; D002417:Cattle; D002531:Cerebellum / Q000737:chemistry; D002540:Cerebral Cortex / Q000737:chemistry; D003342:Corpus Striatum / Q000737:chemistry; D018076:DNA, Complementary; D004546:Elapid Venoms / Q000494:pharmacology*; D006624:Hippocampus / Q000737:chemistry; D007163:Immunosorbent Techniques; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D015221:Potassium Channels / Q000737:chemistry* / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000276:immunology; D014018:Tissue Distribution
D000911:Antibodies, Monoclonal; D018076:DNA, Complementary; D004546:Elapid Venoms; D046911:Macromolecular Substances; D015221:Potassium Channels; D011993:Recombinant Fusion Proteins; C020273:dendrotoxin