Replacement of the glycoinositol phospholipid anchor of Drosophila acetylcholinesterase with a transmembrane domain does not alter sorting in neurons and epithelia but results in behavioral defects.
Drosophila has a single glycoinositol phospholipid (GPI)-anchored form of acetylcholinesterase (AChE) encoded by the Ace locus. To assess the role that GPI plays in the physiology, of AChE, we have replaced the wild-type GPI-AChE with a chimeric transmembrane form (TM-AChE) in the nervous system of the fly. Ace null alleles provided a genetic background completely lacking in endogenous GPI-AChE, and Ace minigene P transposon constructs were used to express both GPI- and TM-AChE forms in the tissues where AChE is normally expressed. Control experiments with the GPI-AChE minigene demonstrated a threshold between 9 and 12% of normal AChE activity for adult viability. Ace mutant flies were rescued by GPI-AChE minigene lines that expressed 12-40% of normal activity and were essentially unchanged from wild-type flies in behavior. TM-AChE minigene lines were able to rescue Ace null alleles, although with a slightly higher threshold than that for GPI-AChE. Although rescued flies expressing GPI-AChE at a level of 12% of normal activity were viable, flies expressing 13-16% of normal activity from the TM-AChE transgene died shortly after eclosion. Flies expressing TM-AChE at about 30% of normal levels were essentially unchanged from wild-type flies in gross behavior but had a reduced lifespan secondary to subtle coordination defects. These flies also showed reduced locomotor activity and performed poorly in a grooming assay. However, light level and electron microscopic immunocytochemistry showed no differences in the localization of GPI- and TM-AChE. Furthermore, endogenous and ectopic-induced expression of both AChEs in epithelial tissues of the adult and embryo, respectively, showed that they were sorted identically. Most epithelial cells sorted GPI- and TM-AChE to the apical surface, but cuticle-secreting epithelia sorted both proteins basolaterally. Our data suggest that rather than having a primary role in protein sorting, the GPI anchor or AChE plays some other more subtle cellular role in neuronal physiology.
Incardona
J P
JP
Department of Genetics, Case Western Reserve University, School of Medicine, Cleveland, Ohio 44106, USA.
Rosenberry
T L
TL
D000110:Acetylcholinesterase / Q000235:genetics* / Q000378:metabolism; D000483:Alleles; D000818:Animals; D004330:Drosophila / Q000196:embryology / Q000201:enzymology* / Q000235:genetics; D004847:Epithelial Cells; D017261:Glycosylphosphatidylinositols / Q000235:genetics / Q000502:physiology*; D008854:Microscopy, Electron; D009043:Motor Activity / Q000235:genetics; D009474:Neurons / Q000166:cytology* / Q000201:enzymology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015183:Restriction Mapping; D014018:Tissue Distribution
D017261:Glycosylphosphatidylinositols; D011993:Recombinant Fusion Proteins; D000110:Acetylcholinesterase