Figure 2. RIC-8 Distribution in Early C. elegans Embryos. (A-G) Wild-type (A, late telophase of one-cell stage; B, four-cell stage),
ric-8(RNAi) (C, four-cell stage),
ric-8(
md1909) (D, four-cell stage),
goa-1/gpa-16(RNAi) (E, four-cell stage),
gpr-1/2(RNAi) (F, four-cell stage), or GFP-RIC-8 (H, four-cell stage) embryos stained with RIC-8 (A-F) or GFP (G) antibodies, as well as with alpha-tubulin antibodies. Left panels show RIC-8 or GFP signal alone, right panels the merge of RIC-8 or GFP (red), alpha-tubulin (green), and DNA (blue). Arrows in (A)-(B) and (D)-(F) point to enrichment at the cell cortex, arrowhead in (A) to enrichment in the vicinity of the microtubule aster. Scale bar = 10 μm.(H) Western blot analysis using RIC-8 antibodies on embryonic extracts of the indicated genotypes (top); the blot was re-probed with antibodies against alpha-tubulin as a loading control (bottom). Note that RIC-8 antibodies detect a major species at the expected size (arrowhead), which is significantly decreased in
ric-8(RNAi) extracts, as well as a minor species (star), which is not, and that is thus likely nonspecific. Note also that RIC-8 in
ric-8(
md1909) co-migrates with this minor species, due to an internal in-frame deletion.