Evidence for the involvement of Tiam1 in axon formation - PubMed (original) (raw)

A, Western blots showing the effect of Tiam1 antisense oligonucleotides on Tiam1, tubulin (Tub), and Rho protein levels. Lane 1, Control non-treated;lane 2, sense treated (ST1a); lane 3, sense treated (ST1b); lane 4, AST1a treated (0.5 μ

m

); lane 5, AST1a treated (1.25 μ

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); lane 6, AST1a treated (2.5 μ

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); lane 7, AST1b treated (2.5 μ

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). For this experiment the oligonucleotides were added 4 hr after plating and replenished 12 hr later. Cell extracts were obtained 1 d after plating. The blots were revealed with Tiam1, tyrosinated α-tubulin, and Rho antibodies; 20 μg of protein was loaded in each lane. Lanes 3 and 4 of the blot stained with the Rho antibody contain no protein. B–I, Tiam1 suppression prevents growth cone enlargement in stage 2 neurons. Double-immunofluorescence micrographs from non-treated (B, C), sense-treated (D, E), AST1a-treated (F, G), or AST1b-treated (H, I) cultures showing the morphology of stage 2 hippocampal pyramidal neurons. Note that all control or sense-treated neurons display a neurite with a large growth cone. This phenomenon is not observed in the antisense-treated neurons; all of the growth cones are of uniform and small size. For this experiment cultures were labeled with rhodamine phalloidin (B, D, F, H) and with an mAb against tyrosinated α-tubulin (C, E, G, I). Oligonucleotides were used at concentrations of 2.5 μ

m

. Scale bar, 10 μm.