Korean J Physiol Pharmacol.  2008 Aug;12(4):199-204. 10.4196/kjpp.2008.12.4.199.

Sorting Nexin 17 Interacts Directly with Kinesin Superfamily KIF1B beta Protein

• Seog DH
• Han J

• KMID: 1838359
• DOI: http://doi.org/10.4196/kjpp.2008.12.4.199

Abstract

KIF1B beta is a member of the Kinesin superfamily proteins (KIFs), which are microtubule-dependent molecular motors that are involved in various intracellular organellar transport processes. KIF1B beta is not restricted to neuronal systems, however, is widely expressed in other tissues, even though the function of KIF1B beta is still unclear. To elucidate the KIF1B beta-binding proteins in non-neuronal cells, we used the yeast two-hybrid system, and found a specific interaction of KIF1B beta and the sorting nexin (SNX) 17. The C-terminal region of SNX17 is required for the binding with KIF1B beta. SNX17 protein bound to the specific region of KIF1B beta (813-916. aa), but not to other kinesin family members. In addition, this specific interaction was also observed in the Glutathione S-transferase pull-down assay. An antibody to SNX17 specifically co-immunoprecipitated KIF1B beta associated with SNX17 from mouse brain extracts. These results suggest that SNX17 might be involved in the KIF1B beta-mediated transport as a KIF1B beta adaptor protein.

Keyword

Kinesin; Molecular motors; Microtubule; Adaptor proteins; SNX

MeSH Terms

Animals
Brain
Glutathione Transferase
Humans
Kinesin
Mice
Microtubules
Neurons
Proteins
Sorting Nexins
Two-Hybrid System Techniques
Glutathione Transferase
Kinesin
Proteins
Sorting Nexins

Figure

• 

  Fig. 1. Identification of the proteins interacting with KIF1Bβ by yeast two-hybrid screening. (A) Schematic illustrations of KIF1A and KIF1B isoforms. Motor domain and KIF1Bβ specific region are indicated gray and hatched box, respectively. Amino acid homology in KIF1A and KIF1B isoforms are represented as a percentage. The positions of the amino acids are indicated. (B) Schematic diagram of domain structure of SNX17. The open box corresponds to PX domain and the filled box to the B41 (FERM) domain. Clone 1, 2 and 5 were found in yeast two-hybrid screen. Clone 1, 2 and 5 were overlapped at the C-terminal region of SNX17. aa, the amino acid residue number. (C) Schematic representation of the SNX17 truncation clones. Several truncated forms of SNX17 were tested in the yeast two-hybrid assay for interaction with KIF1B β. +, interaction with KIF1B β; -, no interaction with KIF1B β.

• 

  Fig. 2. Interaction between KIFs and SNX17. The C-terminal regions of each KIF protein were fused to the pLexA DNA binding domain. SNX17 specifically interacted with KIF1B β, but not with KIF1A, KIF1B α, KIF3A, KIF5B, or KIF17 (+ + +, interaction with SNX17; -, no interaction with SNX17).

• 

  Fig. 3. Association of KIF1B β with SNX17 in the GST pull-down assay and co-immunoprecipitation. (A) Proteins in the mouse brain lysate were allowed to bind to GST alone, GST-SNX17-PX, GST-SNX17-FERM and GST-SNX17-C fusion proteins. The elution fractions were resolved by SDS-PAGE, and Western blotting was performed using an antibody to KIF1Bβ. (B) Mouse brain lysates were immunoprecipitated with an anti-SNX17 antibody or preimmune serum, and the precipitates were immunoblotted with anti-KIF antibodies. Input: 10% of the mouse brain lysates used for each co-immunoprecipitation assay.

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