Spectrin functions upstream of ankyrin in a spectrin cytoskeleton assembly pathway.
Prevailing models place spectrin downstream of ankyrin in a pathway of assembly and function in polarized cells. We used a transgene rescue strategy in Drosophila melanogaster to test contributions of four specific functional sites in beta spectrin to its assembly and function. (1) Removal of the pleckstrin homology domain blocked polarized spectrin assembly in midgut epithelial cells and was usually lethal. (2) A point mutation in the tetramer formation site, modeled after a hereditary elliptocytosis mutation in human erythrocyte spectrin, had no detectable effect on function. (3) Replacement of repetitive segments 4-11 of beta spectrin with repeats 2-9 of alpha spectrin abolished function but did not prevent polarized assembly. (4) Removal of the putative ankyrin-binding site had an unexpectedly mild phenotype with no detectable effect on spectrin targeting to the plasma membrane. The results suggest an alternate pathway in which spectrin directs ankyrin assembly and in which some important functions of spectrin are independent of ankyrin.
Das
Amlan
A
Program in Cell & Developmental Biology, Department of Biological Sciences, University of Illinois at Chicago, Chicago, IL 60607, USA.
Base
Christine
C
Dhulipala
Srilakshmi
S
Dubreuil
Ronald R
RR
D000818:Animals; D017487:Ankyrins / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D002462:Cell Membrane; D003433:Crosses, Genetic; D003599:Cytoskeleton / Q000378:metabolism*; D004330:Drosophila / Q000196:embryology / Q000235:genetics / Q000378:metabolism*; D029721:Drosophila Proteins / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism; D004912:Erythrocytes / Q000166:cytology / Q000378:metabolism; D005260:Female; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D008297:Male; D009154:Mutation; D010641:Phenotype; D015398:Signal Transduction; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism; D013049:Spectrin / Q000502:physiology*; D019076:Transgenes
D017487:Ankyrins; D029721:Drosophila Proteins; C485184:enhanced green fluorescent protein; D013049:Spectrin; D049452:Green Fluorescent Proteins; C472021:ATPalpha protein, Drosophila; D000254:Sodium-Potassium-Exchanging ATPase