A Glial Variant of the Vesicular Monoamine Transporter Is Required To Store Histamine in the Drosophila Visual System
Figure 1
Mutant alleles of dVMAT reduce expression of DVMAT-A and -B.
(A) dVMAT-A and -B share a common translational start site (indicated as “A, B start”) and a common N-terminus, but diverge at their C-termini. Coding exons in the dVMAT gene that are common to both dVMAT-A and-B are shown as gray boxes, introns as black lines. To generate alternative carboxy termini, the indicated genomic sequence (magenta box) is spliced out from dVMAT-A and retained in dVMAT-B. The in situ probe for dVMAT-B contains the first 260 nucleotides of this sequence. The P element insert in the dVMAT mutant allele dVMATP1 (black arrowhead) disrupts the coding sequence of both dVMAT-A and -B. (B,C) In situ hybridization of head sections shows transcription of the dVMAT-B gene in a layer between the retina and lamina (B). Magnified view of (B) shown in (C). (D) Cartoons showing the predicted topology of DVMAT with lumenal domains above and cytoplasmic domains below the parallel gray lines representing the vesicle membrane. Open circles indicate domains shared by DVMAT-A and -B. Filled, magenta circles indicate the C-terminal domain specific for DVMAT-B. The P element insertion site in the last exon of the dVMATP1 (marked with “P” and a black arrowhead) functionally deletes transmembrane domains 11 and 12 and the C-terminus (shaded gray). The imprecise excision allele dVMATΔ14 results in an insertion of 51 base pairs, and 17 amino acids in-frame with the original downstream codons (marked as “Δ14” with a blue arrowhead, with blue circles indicating the inserted residues). (E) Western blot using the N-terminus antibody directed against both DVMAT-A and -B splice variants shows an absence of DVMAT protein in dVMATP1 homozygotes (P/P), and dramatically reduced levels in dVMATΔ14 (Δ14/Δ14), compared with heterozygous controls (P/CyO and Δ14/CyO). The plasma membrane associated protein Late Bloomer (indicated as “lb”) was used as a loading control. Re, retina; La, lamina; Me, medulla. Bars: (B) 50 microns, (C) 20 microns.