Limits...
Kinetochore-dependent microtubule rescue ensures their efficient and sustained interactions in early mitosis.

Gandhi SR, Gierliński M, Mino A, Tanaka K, Kitamura E, Clayton L, Tanaka TU - Dev. Cell (2011)

Bottom Line: Meanwhile, microtubule rescue distal to the kinetochore is also promoted by Stu2, which is transported by a kinesin-8 motor Kip3 along the microtubule from the kinetochore.Microtubule extension following rescue facilitates interaction with other widely scattered kinetochores, diminishing long delays in collecting the complete set of kinetochores by microtubules.Thus, kinetochore-dependent microtubule rescue ensures efficient and sustained kinetochore-microtubule interactions in early mitosis.

View Article: PubMed Central - PubMed

Affiliation: Wellcome Trust Centre for Gene Regulation & Expression, College of Life Sciences, University of Dundee, Dundee DD1 5EH, UK.

Show MeSH

Related in: MedlinePlus

Microtubule Rescue at the Kinetochore Is Facilitated by Stu2 and Prevents a Loss of Kinetochore-Microtubule Attachment(A) Stu2 facilitates MT rescue at the KT and prevents loss of KT-MT attachment. PSTU2-stu2ΔTOG1 (T9323; stu2 hypomorphic mutant [hypo]) and PSTU2-STU2+(T9345; control) cells (i.e., with an extra stu2 or STU2+ gene at an auxotroph marker locus) with PMET3-CDC20 PGAL-CEN3-lacOs TetR-GFP GFP-TUB1 STU2+ were treated, and their images were acquired as in Figure 1C. (i) Schematic diagrams show wild-type STU2 and a mutant stu2 with the TOG1 domain deleted. 7/281: 7–281 amino acid residues are deleted. (ii) Graph showing the proportion of the events (shown in the diagram) after the plus end of a shrinking MT catches up with CEN3 in the control (n = 20) and stu2 hypomorphic mutant (n = 38). Representative examples of (iii) CEN3 detachment from the MT end (preceded by MT pausing) and (iv) end-on attachment/pulling (preceded by MT pausing), found in the stu2 hypomorphic mutant. Zero time is set arbitrarily for the first panel. Scale bar, 1 μm. The graphs show MT length (orange) and CEN3–spindle-pole distance (blue).(B) The double mutant of stu2 hypomorph and the ndc80 loop region shows frequent CEN3 detachment from the MT end. (i) Diagram showing the deletion in ndc80Δ490–510 (Maure et al., 2011). (ii) NDC80+ PSTU2-STU2+ (T9460), ndc80Δ490–510 PSTU2-STU2+ (T9456), NDC80+ PSTU2-stu2ΔTOG1 (T9470), and ndc80Δ490–510 PSTU2-stu2ΔTOG1 (T9455) cells with PMET3-CDC20 PGAL-CEN3-tetOs TetR-GFP Venus-TUB1 STU2+ were treated, and their images were acquired as in Figure 1C, except that the temperature for cell culture was shifted from 25°C to 35°C, 30 min prior to image acquisition. The restrictive temperature for ndc80Δ490–510 is 35°C (Maure et al., 2011). Graph shows the proportion of the events (categorized as in Aii, diagram) after the plus end of a shrinking MT catches up with CEN3 (n = 31, 33, 23, and 39 from top to bottom). Ndc80 control was Ndc80 wild-type. Stu2 hypo and control were as defined in (A).See also Figure S1.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3277888&req=5

fig2: Microtubule Rescue at the Kinetochore Is Facilitated by Stu2 and Prevents a Loss of Kinetochore-Microtubule Attachment(A) Stu2 facilitates MT rescue at the KT and prevents loss of KT-MT attachment. PSTU2-stu2ΔTOG1 (T9323; stu2 hypomorphic mutant [hypo]) and PSTU2-STU2+(T9345; control) cells (i.e., with an extra stu2 or STU2+ gene at an auxotroph marker locus) with PMET3-CDC20 PGAL-CEN3-lacOs TetR-GFP GFP-TUB1 STU2+ were treated, and their images were acquired as in Figure 1C. (i) Schematic diagrams show wild-type STU2 and a mutant stu2 with the TOG1 domain deleted. 7/281: 7–281 amino acid residues are deleted. (ii) Graph showing the proportion of the events (shown in the diagram) after the plus end of a shrinking MT catches up with CEN3 in the control (n = 20) and stu2 hypomorphic mutant (n = 38). Representative examples of (iii) CEN3 detachment from the MT end (preceded by MT pausing) and (iv) end-on attachment/pulling (preceded by MT pausing), found in the stu2 hypomorphic mutant. Zero time is set arbitrarily for the first panel. Scale bar, 1 μm. The graphs show MT length (orange) and CEN3–spindle-pole distance (blue).(B) The double mutant of stu2 hypomorph and the ndc80 loop region shows frequent CEN3 detachment from the MT end. (i) Diagram showing the deletion in ndc80Δ490–510 (Maure et al., 2011). (ii) NDC80+ PSTU2-STU2+ (T9460), ndc80Δ490–510 PSTU2-STU2+ (T9456), NDC80+ PSTU2-stu2ΔTOG1 (T9470), and ndc80Δ490–510 PSTU2-stu2ΔTOG1 (T9455) cells with PMET3-CDC20 PGAL-CEN3-tetOs TetR-GFP Venus-TUB1 STU2+ were treated, and their images were acquired as in Figure 1C, except that the temperature for cell culture was shifted from 25°C to 35°C, 30 min prior to image acquisition. The restrictive temperature for ndc80Δ490–510 is 35°C (Maure et al., 2011). Graph shows the proportion of the events (categorized as in Aii, diagram) after the plus end of a shrinking MT catches up with CEN3 (n = 31, 33, 23, and 39 from top to bottom). Ndc80 control was Ndc80 wild-type. Stu2 hypo and control were as defined in (A).See also Figure S1.

Mentions: Establishment of end-on attachment (which is verified by subsequent end-on pulling) is probably crucial for subsequent sister KT biorientation (Maure et al., 2011; Tanaka, 2010). However, if end-on attachment is the essential next step for KT-MT interaction, what are the benefits of KT-dependent MT rescue, which extends the duration of lateral attachment and seems to delay establishment of end-on attachment? A clue to this question was obtained regarding MT rescue at the KT, when we recently studied mutants of the Ndc80 loop region (Maure et al., 2011), a distinct motif looping out from the coiled-coil shaft of the Ndc80 complex (see Figure 2Bi). These mutants were defective in forming end-on attachment; indeed, when the MT plus end caught up with a laterally associated KT, end-on attachment was rarely established. Given this, we expected that KTs would detach from MT ends in these mutants. However, this was rare, and instead MT rescue at the KT happened in most cases (Maure et al., 2011). This raised the possibility that MT rescue at a KT may happen, following failure to achieve end-on attachment.


Kinetochore-dependent microtubule rescue ensures their efficient and sustained interactions in early mitosis.

Gandhi SR, Gierliński M, Mino A, Tanaka K, Kitamura E, Clayton L, Tanaka TU - Dev. Cell (2011)

Microtubule Rescue at the Kinetochore Is Facilitated by Stu2 and Prevents a Loss of Kinetochore-Microtubule Attachment(A) Stu2 facilitates MT rescue at the KT and prevents loss of KT-MT attachment. PSTU2-stu2ΔTOG1 (T9323; stu2 hypomorphic mutant [hypo]) and PSTU2-STU2+(T9345; control) cells (i.e., with an extra stu2 or STU2+ gene at an auxotroph marker locus) with PMET3-CDC20 PGAL-CEN3-lacOs TetR-GFP GFP-TUB1 STU2+ were treated, and their images were acquired as in Figure 1C. (i) Schematic diagrams show wild-type STU2 and a mutant stu2 with the TOG1 domain deleted. 7/281: 7–281 amino acid residues are deleted. (ii) Graph showing the proportion of the events (shown in the diagram) after the plus end of a shrinking MT catches up with CEN3 in the control (n = 20) and stu2 hypomorphic mutant (n = 38). Representative examples of (iii) CEN3 detachment from the MT end (preceded by MT pausing) and (iv) end-on attachment/pulling (preceded by MT pausing), found in the stu2 hypomorphic mutant. Zero time is set arbitrarily for the first panel. Scale bar, 1 μm. The graphs show MT length (orange) and CEN3–spindle-pole distance (blue).(B) The double mutant of stu2 hypomorph and the ndc80 loop region shows frequent CEN3 detachment from the MT end. (i) Diagram showing the deletion in ndc80Δ490–510 (Maure et al., 2011). (ii) NDC80+ PSTU2-STU2+ (T9460), ndc80Δ490–510 PSTU2-STU2+ (T9456), NDC80+ PSTU2-stu2ΔTOG1 (T9470), and ndc80Δ490–510 PSTU2-stu2ΔTOG1 (T9455) cells with PMET3-CDC20 PGAL-CEN3-tetOs TetR-GFP Venus-TUB1 STU2+ were treated, and their images were acquired as in Figure 1C, except that the temperature for cell culture was shifted from 25°C to 35°C, 30 min prior to image acquisition. The restrictive temperature for ndc80Δ490–510 is 35°C (Maure et al., 2011). Graph shows the proportion of the events (categorized as in Aii, diagram) after the plus end of a shrinking MT catches up with CEN3 (n = 31, 33, 23, and 39 from top to bottom). Ndc80 control was Ndc80 wild-type. Stu2 hypo and control were as defined in (A).See also Figure S1.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3277888&req=5

fig2: Microtubule Rescue at the Kinetochore Is Facilitated by Stu2 and Prevents a Loss of Kinetochore-Microtubule Attachment(A) Stu2 facilitates MT rescue at the KT and prevents loss of KT-MT attachment. PSTU2-stu2ΔTOG1 (T9323; stu2 hypomorphic mutant [hypo]) and PSTU2-STU2+(T9345; control) cells (i.e., with an extra stu2 or STU2+ gene at an auxotroph marker locus) with PMET3-CDC20 PGAL-CEN3-lacOs TetR-GFP GFP-TUB1 STU2+ were treated, and their images were acquired as in Figure 1C. (i) Schematic diagrams show wild-type STU2 and a mutant stu2 with the TOG1 domain deleted. 7/281: 7–281 amino acid residues are deleted. (ii) Graph showing the proportion of the events (shown in the diagram) after the plus end of a shrinking MT catches up with CEN3 in the control (n = 20) and stu2 hypomorphic mutant (n = 38). Representative examples of (iii) CEN3 detachment from the MT end (preceded by MT pausing) and (iv) end-on attachment/pulling (preceded by MT pausing), found in the stu2 hypomorphic mutant. Zero time is set arbitrarily for the first panel. Scale bar, 1 μm. The graphs show MT length (orange) and CEN3–spindle-pole distance (blue).(B) The double mutant of stu2 hypomorph and the ndc80 loop region shows frequent CEN3 detachment from the MT end. (i) Diagram showing the deletion in ndc80Δ490–510 (Maure et al., 2011). (ii) NDC80+ PSTU2-STU2+ (T9460), ndc80Δ490–510 PSTU2-STU2+ (T9456), NDC80+ PSTU2-stu2ΔTOG1 (T9470), and ndc80Δ490–510 PSTU2-stu2ΔTOG1 (T9455) cells with PMET3-CDC20 PGAL-CEN3-tetOs TetR-GFP Venus-TUB1 STU2+ were treated, and their images were acquired as in Figure 1C, except that the temperature for cell culture was shifted from 25°C to 35°C, 30 min prior to image acquisition. The restrictive temperature for ndc80Δ490–510 is 35°C (Maure et al., 2011). Graph shows the proportion of the events (categorized as in Aii, diagram) after the plus end of a shrinking MT catches up with CEN3 (n = 31, 33, 23, and 39 from top to bottom). Ndc80 control was Ndc80 wild-type. Stu2 hypo and control were as defined in (A).See also Figure S1.
Mentions: Establishment of end-on attachment (which is verified by subsequent end-on pulling) is probably crucial for subsequent sister KT biorientation (Maure et al., 2011; Tanaka, 2010). However, if end-on attachment is the essential next step for KT-MT interaction, what are the benefits of KT-dependent MT rescue, which extends the duration of lateral attachment and seems to delay establishment of end-on attachment? A clue to this question was obtained regarding MT rescue at the KT, when we recently studied mutants of the Ndc80 loop region (Maure et al., 2011), a distinct motif looping out from the coiled-coil shaft of the Ndc80 complex (see Figure 2Bi). These mutants were defective in forming end-on attachment; indeed, when the MT plus end caught up with a laterally associated KT, end-on attachment was rarely established. Given this, we expected that KTs would detach from MT ends in these mutants. However, this was rare, and instead MT rescue at the KT happened in most cases (Maure et al., 2011). This raised the possibility that MT rescue at a KT may happen, following failure to achieve end-on attachment.

Bottom Line: Meanwhile, microtubule rescue distal to the kinetochore is also promoted by Stu2, which is transported by a kinesin-8 motor Kip3 along the microtubule from the kinetochore.Microtubule extension following rescue facilitates interaction with other widely scattered kinetochores, diminishing long delays in collecting the complete set of kinetochores by microtubules.Thus, kinetochore-dependent microtubule rescue ensures efficient and sustained kinetochore-microtubule interactions in early mitosis.

View Article: PubMed Central - PubMed

Affiliation: Wellcome Trust Centre for Gene Regulation & Expression, College of Life Sciences, University of Dundee, Dundee DD1 5EH, UK.

Show MeSH
Related in: MedlinePlus