KAP, the Accessory Subunit of Kinesin-2, Binds the Predicted Coiled-Coil Stalk of the Motor Subunits

Abstract

Kinesin-2 is an anterograde motor involved in intraflagellar transport and certain other intracellular transport processes. It consists of two different motor subunits and an accessory protein KAP (kinesin accessory protein). The motor subunits were shown to bind each other through the coiled-coil stalk domains, while KAP was proposed to bind the tail domains of the motor subunits. Although several genetic studies established that KAP plays an important role in kinesin-2 functions, its exact role remains unclear. Here, we report the results of a systematic analysis of the KAP binding sites by using recombinant Drosophila kinesin-2 subunits as well as the endogenous proteins. These show that at least one of the coiled-coil stalks is sufficient to bind the N-terminal region of DmKAP. The soluble complex involving the recombinant kinesin-2 fragments is reconstituted in vitro at high salt concentrations, suggesting that the interaction is primarily nonionic. Furthermore, independent distant homology modeling indicated that DmKAP may bind along the coiled-coil stalks through a combination of predominantly hydrophobic interactions and hydrogen bonds. These observations led us to propose that KAP would stabilize the motor subunit heterodimer and help assemble a greater kinesin-2 complex in vivo.

Kinesin-2 motors: Kinetics and biophysics

“…KIF3AB-KAP transports multimeric protein complexes (designated IFT particles) into the cilium, and its transport role for ciliogenesis is considered the reason that KIF3AB-KAP is essential for development. It also appears to be the basis of similarity between KIF3AB-KAP and other kinesin-2 heterotrimeric orthologs.In contrast, there is not strong experimental evidence that mammalian KIF3AC binds KAP (5,52,55,65). Another observation to rule out an association of KAP with KIF3AC is the lack of sequence similarity of KIF3C at the putative KAP binding region of KIF3AB.…”

Kinesin-2 heterodimerization alters entry into a processive run along the microtubule but not stepping within the run

“…KIF3AB-KAP transports multimeric protein complexes (designated intraflagellar transport (IFT) particles) into the cilium and has also been linked to ciliadependent signal transduction pathways including the Hedgehog signaling pathway (16,17). Moreover, KIF3A, KIF3B, and KAP are all essential genes (18)(19)(20)(21)(22)(23), and the transport role of KIF3AB-KAP in ciliogenesis is considered the reason that KIF3AB-KAP is essential for development and the basis of similarity with other heterotrimeric kinesin-2 motor proteins.In contrast, there is not strong evidence that mammalian KIF3AC binds KAP, and KIF3C lacks the sequence similarity to KIF3AB at the putative C-terminal KAP binding region (2,11,14,24). In spite of similarities in sequence and structure, KIF3AC appears to function specifically in neurons whereas heterotrimeric KIF3AB-KAP is more ubiquitously expressed and transports ciliary IFT particles, melanosomes, and organelles as well as cell signaling and cell adhesion molecules (21,23,(25)(26)(27)(28)(29)(30)(31)(32).…”

“…In contrast, there is not strong evidence that mammalian KIF3AC binds KAP, and KIF3C lacks the sequence similarity to KIF3AB at the putative C-terminal KAP binding region (2,11,14,24). In spite of similarities in sequence and structure, KIF3AC appears to function specifically in neurons whereas heterotrimeric KIF3AB-KAP is more ubiquitously expressed and transports ciliary IFT particles, melanosomes, and organelles as well as cell signaling and cell adhesion molecules (21,23,(25)(26)(27)(28)(29)(30)(31)(32).…”

“…KIF3AB and its orthologs form a heterotrimeric complex by association with KAP, a distinctive http://www.jbc.org/cgi/doi/10.1074/jbc.RA118.002767 The latest version is at JBC Papers in Press. Published on July 10, 2018 as Manuscript RA118.002767 by guest on April 27, 2019 http://www.jbc.org/ Downloaded from 2 adaptor protein for cargo linkage because it is largely composed of armadillo repeats (10)(11)(12)(13)(14)(15). It is the armadillo repeats that provide specificity of the interaction between KIF3AB and KAP and between KIF3AB-KAP and its cargo.…”

“…adaptor protein for cargo linkage because it is largely composed of armadillo repeats (10)(11)(12)(13)(14)(15). It is the armadillo repeats that provide specificity of the interaction between KIF3AB and KAP and between KIF3AB-KAP and its cargo.…”