Mammalian Kinesin-3 Motors Are Dimeric In Vivo and Move by Processive Motility upon Release of Autoinhibition
Figure 4
The NC Is Required for Dimerization of KIF1A
(A) Sequence comparison (top, ClustalW) of the NC region across species. Red, identical residues; orange, similar residues. Schematic (bottom) of NC constructs used in this study and their relation to KIF1A/Unc104 constructs used in previous studies.
(B) Crosslinking. Lysates from COS cells expressing Myc-tagged NC truncations were treated with or without DMP, separated by SDS-PAGE, and immunoblotted with anti-Myc antibodies. Arrow, reduced mobility of KIF1A(1–393) upon crosslinking.
(C and D) Photobleaching analysis. Lysates of COS cells expressing 3xmCit-tagged NC constructs were imaged by TIRF microscopy. (C) Stepwise photobleaching of representative spots. (D) Distribution of the number of photobleaching steps. (1–393), n = 171 spots, average = 3.91 ± 0.10 steps; (1–381), n = 181 spots, average = 2.40 ± 0.07 steps; (1–369), n = 171 spots, average = 2.49 ± 0.07 steps.