Ankyrin and beta-spectrin accumulate independently of alpha-spectrin in Drosophila.
We report the identification and initial characterization of Drosophila melanogaster ankyrin. Oligonucleotide primers based on the spectrin-binding domain of human brain ankyrin were used to amplify Drosophila genomic DNA. A cloned 184-bp PCR product was used to isolate Drosophila ankyrin cDNAs. Ankyrin cDNA probes detected a 5.5-kb transcript from embryonic poly(A)+ RNA and a single polytene chromosome locus (101F-102A). The cDNA sequence encodes a 170-kDa protein that is 53% identical to human brain ankyrin (Ank2). Antibodies directed against a recombinant fragment of Drosophila ankyrin reacted with a 170-kDa polypeptide from Drosophila embryos, larvae, S2 cells, and adult flies. The ankyrin antibody coimmunoprecipitated alpha- and beta-spectrin with ankyrin in detergent extracts of Drosophila embryo membranes. Antibodies against Drosophila ankyrin, alpha-spectrin and beta-spectrin were used to detect these proteins in wild-type and alpha-spectrin-mutant larvae. alpha-Spectrin levels were greatly diminished in mutant larvae, but levels of ankyrin and beta-spectrin were indistinguishable from wild type. The persistence of ankyrin and beta-spectrin may explain the relatively mild phenotype of alpha-spectrin mutants during early Drosophila development.
Dubreuil
R R
RR
Department of Pharmacological and Physiological Sciences, University of Chicago, IL 60637.
Yu
J
J
D000595:Amino Acid Sequence; D000818:Animals; D017487:Ankyrins / Q000096:biosynthesis* / Q000276:immunology / Q000302:isolation & purification; D000906:Antibodies; D001483:Base Sequence; D015139:Blotting, Southern; D001921:Brain / Q000378:metabolism; D017931:DNA Primers; D015342:DNA Probes; D018076:DNA, Complementary / Q000302:isolation & purification; D004331:Drosophila melanogaster / Q000378:metabolism*; D006801:Humans; D015151:Immunoblotting; D007814:Larva; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000276:immunology / Q000302:isolation & purification; D015183:Restriction Mapping; D013049:Spectrin / Q000096:biosynthesis*
D017487:Ankyrins; D000906:Antibodies; D017931:DNA Primers; D015342:DNA Probes; D018076:DNA, Complementary; D011994:Recombinant Proteins; D013049:Spectrin