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Fig. 5 | BMC Molecular and Cell Biology

Fig. 5

From: Aurora A site specific TACC3 phosphorylation regulates astral microtubule assembly by stabilizing γ-tubulin ring complex

Fig. 5

Phosphorylation of TACC3 at Ser558 is involved in the assembly of γ-TuRC from γ-TuSC. Sedimentation of γ-TuSC, and γ-TuRC along with GFP-TACC3 S558A or S558D from a. GFP-TACC3KDP(S558A)-TACC3 shRNA vs. b. GFP-TACC3KDP(S558D)-TACC3 shRNA transfected (48 h) HeLa cells. Cell lysates were centrifuged through 5–40% sucrose gradient at 150,000 X g, and the fractions collected from different layers of the gradient were then run through 10% SDS-PAGE followed by immunoblotting against γ-tubulin, GCP3, GCP4 and GFP-TACC3. Sedimentation coefficient (S) values of the protein complexes were assessed by calibrating against the known markers, BSA (66 kDa, 4.4 S), β-amylase (200 kDa, 8.9 S) and Thyroglobulin (669 kDa, 19.4 S) as shown. γ-TuSC and γ-TuRC were sedimented at 6–10 S and above 22 S, respectively

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