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Fig. 2. Assays of Shroom2 activity. (A-C) Shroom2 activity was
measured by blind-scoring of pigment accumulation in injected embryos. Panels
A-C show representative embryos displaying normal, mild and severe phenotypes
for the scoring shown in D-F. (D) Shroom2 (576-882) blocked the ability
of ectopic Shroom2 to induce apical pigment accumulation in a dose-dependent
manner. Shroom2 (0.4 ng) was injected alone or in combination with 0.8 ng or
1.6 ng of Shroom2(576-882) in one blastomeres at the four-cell stage.
(E) Shroom2 does not function through the Rap1B GTPase. Shroom2 (0.75
ng) was injected alone or in combination with 2 ng of Rap1B-N17 or RapGAP into
one blastomere at the four-cell stage. (F) Shroom2 acts upstream of
Rab27a as co-expression of either dominant-negative Rab27a-L130P or
Rab27a-A152P inhibited Shroom2 activity. Shroom2 (0.4 ng) was injected alone
or in combination with 1.2 ng of Rab27a-L130P or Rab27a-A152P into one
blastomere at the four-cell stage.
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