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Single molecule imaging reveals differences in microtubule track selection between Kinesin motors.

Cai D, McEwen DP, Martens JR, Meyhofer E, Verhey KJ - PLoS Biol. (2009)

Bottom Line: In contrast, individual Kinesin-2 (KIF17) and Kinesin-3 (KIF1A) motors do not select subsets of microtubules.Surprisingly, KIF17 and KIF1A motors that overtake the plus ends of growing microtubules do not fall off but rather track with the growing tip.These results indicate that kinesin families can be distinguished by their ability to recognize microtubule heterogeneity.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Developmental Biology, University of Michigan, Ann Arbor, Michigan, USA.

ABSTRACT
Cells generate diverse microtubule populations by polymerization of a common alpha/beta-tubulin building block. How microtubule associated proteins translate microtubule heterogeneity into specific cellular functions is not clear. We evaluated the ability of kinesin motors involved in vesicle transport to read microtubule heterogeneity by using single molecule imaging in live cells. We show that individual Kinesin-1 motors move preferentially on a subset of microtubules in COS cells, identified as the stable microtubules marked by post-translational modifications. In contrast, individual Kinesin-2 (KIF17) and Kinesin-3 (KIF1A) motors do not select subsets of microtubules. Surprisingly, KIF17 and KIF1A motors that overtake the plus ends of growing microtubules do not fall off but rather track with the growing tip. Selection of microtubule tracks restricts Kinesin-1 transport of VSVG vesicles to stable microtubules in COS cells whereas KIF17 transport of Kv1.5 vesicles is not restricted to specific microtubules in HL-1 myocytes. These results indicate that kinesin families can be distinguished by their ability to recognize microtubule heterogeneity. Furthermore, this property enables kinesin motors to segregate membrane trafficking events between stable and dynamic microtubule populations.

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Related in: MedlinePlus

Preferential motility of Kinesin-1 occurs along stable microtubules marked by acetylation and detyrosination of α-tubulin.Single KHC(1-560)-3xmCit motors in live COS cells were imaged by TIRF microscopy. The cells were fixed and stained for retrospective immunofluorescence. SD Maps of the KHC(1-560)-3xmCit motility events were created from the time series (far left images) and compared to the fixed images of (A) total tubulin (middle panel) and acetylated tubulin (right panel), (B) total tubulin (middle panel) and detyrosinated tubulin (right panel), or (C) total tubulin (middle panel) and polyglutamylated tubulins (right panel). Scale bar, 4 µm. The far right panels indicate schematic representations of the overlap between the Kinesin-1 motility events in the SD Map (green lines) and the modified microtubules (red lines).
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pbio-1000216-g003: Preferential motility of Kinesin-1 occurs along stable microtubules marked by acetylation and detyrosination of α-tubulin.Single KHC(1-560)-3xmCit motors in live COS cells were imaged by TIRF microscopy. The cells were fixed and stained for retrospective immunofluorescence. SD Maps of the KHC(1-560)-3xmCit motility events were created from the time series (far left images) and compared to the fixed images of (A) total tubulin (middle panel) and acetylated tubulin (right panel), (B) total tubulin (middle panel) and detyrosinated tubulin (right panel), or (C) total tubulin (middle panel) and polyglutamylated tubulins (right panel). Scale bar, 4 µm. The far right panels indicate schematic representations of the overlap between the Kinesin-1 motility events in the SD Map (green lines) and the modified microtubules (red lines).

Mentions: We then tested whether Kinesin-1 motors move preferentially on stable microtubule tracks. To do this, we performed retrospective immunofluorescence staining using antibodies that recognize the PTMs that mark stable microtubules. Cells expressing KHC(1-560)-3xmCit were imaged in the TIRF microscope, fixed, stained with antibodies to acetylated α-tubulin and total tubulin, and the previously imaged cells were again viewed on the TIRF microscope. The pattern of KHC(1-560)-3xmCit motility events in the resulting SD Map was similar to the pattern of acetylated microtubules (Figure 3A, representative of 11 cells in six experiments). Kinesin-1 motility events colocalized with 90.3%±5.5% of microtubules marked by acetylated tubulin (Table 1). This suggests that Kinesin-1 moves preferentially along stable microtubules marked by acetylation of α-tubulin.


Single molecule imaging reveals differences in microtubule track selection between Kinesin motors.

Cai D, McEwen DP, Martens JR, Meyhofer E, Verhey KJ - PLoS Biol. (2009)

Preferential motility of Kinesin-1 occurs along stable microtubules marked by acetylation and detyrosination of α-tubulin.Single KHC(1-560)-3xmCit motors in live COS cells were imaged by TIRF microscopy. The cells were fixed and stained for retrospective immunofluorescence. SD Maps of the KHC(1-560)-3xmCit motility events were created from the time series (far left images) and compared to the fixed images of (A) total tubulin (middle panel) and acetylated tubulin (right panel), (B) total tubulin (middle panel) and detyrosinated tubulin (right panel), or (C) total tubulin (middle panel) and polyglutamylated tubulins (right panel). Scale bar, 4 µm. The far right panels indicate schematic representations of the overlap between the Kinesin-1 motility events in the SD Map (green lines) and the modified microtubules (red lines).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2749942&req=5

pbio-1000216-g003: Preferential motility of Kinesin-1 occurs along stable microtubules marked by acetylation and detyrosination of α-tubulin.Single KHC(1-560)-3xmCit motors in live COS cells were imaged by TIRF microscopy. The cells were fixed and stained for retrospective immunofluorescence. SD Maps of the KHC(1-560)-3xmCit motility events were created from the time series (far left images) and compared to the fixed images of (A) total tubulin (middle panel) and acetylated tubulin (right panel), (B) total tubulin (middle panel) and detyrosinated tubulin (right panel), or (C) total tubulin (middle panel) and polyglutamylated tubulins (right panel). Scale bar, 4 µm. The far right panels indicate schematic representations of the overlap between the Kinesin-1 motility events in the SD Map (green lines) and the modified microtubules (red lines).
Mentions: We then tested whether Kinesin-1 motors move preferentially on stable microtubule tracks. To do this, we performed retrospective immunofluorescence staining using antibodies that recognize the PTMs that mark stable microtubules. Cells expressing KHC(1-560)-3xmCit were imaged in the TIRF microscope, fixed, stained with antibodies to acetylated α-tubulin and total tubulin, and the previously imaged cells were again viewed on the TIRF microscope. The pattern of KHC(1-560)-3xmCit motility events in the resulting SD Map was similar to the pattern of acetylated microtubules (Figure 3A, representative of 11 cells in six experiments). Kinesin-1 motility events colocalized with 90.3%±5.5% of microtubules marked by acetylated tubulin (Table 1). This suggests that Kinesin-1 moves preferentially along stable microtubules marked by acetylation of α-tubulin.

Bottom Line: In contrast, individual Kinesin-2 (KIF17) and Kinesin-3 (KIF1A) motors do not select subsets of microtubules.Surprisingly, KIF17 and KIF1A motors that overtake the plus ends of growing microtubules do not fall off but rather track with the growing tip.These results indicate that kinesin families can be distinguished by their ability to recognize microtubule heterogeneity.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Developmental Biology, University of Michigan, Ann Arbor, Michigan, USA.

ABSTRACT
Cells generate diverse microtubule populations by polymerization of a common alpha/beta-tubulin building block. How microtubule associated proteins translate microtubule heterogeneity into specific cellular functions is not clear. We evaluated the ability of kinesin motors involved in vesicle transport to read microtubule heterogeneity by using single molecule imaging in live cells. We show that individual Kinesin-1 motors move preferentially on a subset of microtubules in COS cells, identified as the stable microtubules marked by post-translational modifications. In contrast, individual Kinesin-2 (KIF17) and Kinesin-3 (KIF1A) motors do not select subsets of microtubules. Surprisingly, KIF17 and KIF1A motors that overtake the plus ends of growing microtubules do not fall off but rather track with the growing tip. Selection of microtubule tracks restricts Kinesin-1 transport of VSVG vesicles to stable microtubules in COS cells whereas KIF17 transport of Kv1.5 vesicles is not restricted to specific microtubules in HL-1 myocytes. These results indicate that kinesin families can be distinguished by their ability to recognize microtubule heterogeneity. Furthermore, this property enables kinesin motors to segregate membrane trafficking events between stable and dynamic microtubule populations.

Show MeSH
Related in: MedlinePlus