RECQL4 is a RanGTP dependent MAP

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RECQL4 is a RanGTP dependent MAP RECQL4 is a RanGTP dependent MAP. (A) Endogenous human RECQL4 binds to microtubules (MTs) in a salt-sensitive manner. RECQL4 is a RanGTP dependent MAP. (A) Endogenous human RECQL4 binds to microtubules (MTs) in a salt-sensitive manner. HeLa nuclear extract was incubated in the presence or absence of taxol-stabilized MTs and 0.5 M NaCl and centrifuged. MT pellets were immunoblotted using human RECQL4 and α-tubulin antibodies. (B) Preparation of human RECQL4 antibody. Recombinant human RECQL4 aa 1–831 was expressed in bacteria, purified, and used for antibody production in rabbits. The generated antibody detected endogenous hRECQL4 in HeLa cell lysate. *, nonspecific band. (C) Endogenous Xenopus RECQL4 binds to MTs. Xenopus CSF egg extract was incubated with taxol-stabilized MTs. MTs and MAPs were spun down. MAPs are eluted from MTs with high salt and separated by centrifugation. The eluate and MT pellet were immunoblotted. (D) Recombinant Xenopus RECQL4 expressed in insect cells and purified with TALON beads. Coomassie staining and Western blot (WB) using an anti-His 6-antibody are shown. (E)Xenopus RECQL4 binds to MTs in a RanGTP-regulated manner. Recombinant RECQL4 was incubated with MTs as in Fig 1B, but, where indicated, in the presence of recombinant importin α, importin β, and/or RanQ69L mutant loaded with GTP (RanGTP). After centrifugation, supernatant (s) and pellet (p) were analyzed by Western blot. Hideki Yokoyama et al. LSA 2019;2:e201800120 © 2019 Yokoyama et al.