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Fig. 3. The 18304 locus encodes Drosophila Pka-R1. (A) Diagram of the
genomic region defined by the overlap of two deficiencies that fail to
complement the 18304 maternal-effect phenotype. The position of the proximal
breakpoint of Df(3L)ri-XT106 and the distal breakpoint of
Df(3L)ME107 (shown in brackets) were identified by RFLPs specific for
the 18304 chromosome (see Materials and methods). Six gene candidates for the
18304 locus map to this region: Pka-R1 (four alternative transcripts
are shown), CG3288, CG13255, CSN3, CG11456 and CG32432
(Berkeley Drosophila Genome Project). (B) Domain organization of
Drosophila PKA regulatory subunit type 1. One isoform (RA) comprises
a dimerization domain at the N terminus, followed by an inhibitory domain and
two cAMP-binding domains. The other two isoforms (RB and RD) lack the
dimerization domain. (C) Western blot analysis using human PKA-R1ß
antibody to reveal the expression of PKA-R1 in the Drosophila ovary.
A single band of 50 kD is detected in ovarian extracts and corresponds to the
size of the RA isoform. (D) Alignment of D. melanogaster, H. sapiens, M.
musculus and C. elegans PKA-R1. In 18304, a conserved arginine
residue in the inhibitory domain is mutated to glutamine and, in E1, a
conserved glycine residue is mutated to aspartic acid. (E-G) Subcellular
localization of PKA-R1 protein during oogenesis revealed by anti-PKA-R1
immunostaining (E). PKA-R1 is detected in the cytoplasm, with an accumulation
at the cell membrane. (F) Rhodamine-conjugated phalloidin reveals the
subcortical actin cytoskeleton in the oocyte, nurse cells and follicle cells.
(G) Merged image of E and F showing co-localization of PKA-R1 and actin at the
cell cortex.
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