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STIL binding to Polo-box 3 of PLK4 regulates centriole duplication.

Arquint C, Gabryjonczyk AM, Imseng S, Böhm R, Sauer E, Hiller S, Nigg EA, Maier T - Elife (2015)

Bottom Line: STIL-CC is the first identified interaction partner of Polo-box 3 (PB3) of PLK4 and also uses a secondary interaction site in the PLK4 L1 region.In vivo analysis of structure-guided STIL mutants reveals distinct binding modes to PLK4-PB3 and L1, as well as interplay of STIL oligomerization with PLK4 binding.We suggest that the STIL-CC/PLK4 interaction mediates PLK4 activation as well as stabilization of centriolar PLK4 and plays a key role in centriole duplication.

View Article: PubMed Central - PubMed

Affiliation: Biozentrum, University of Basel, Basel, Switzerland.

ABSTRACT
Polo-like kinases (PLK) are eukaryotic regulators of cell cycle progression, mitosis and cytokinesis; PLK4 is a master regulator of centriole duplication. Here, we demonstrate that the SCL/TAL1 interrupting locus (STIL) protein interacts via its coiled-coil region (STIL-CC) with PLK4 in vivo. STIL-CC is the first identified interaction partner of Polo-box 3 (PB3) of PLK4 and also uses a secondary interaction site in the PLK4 L1 region. Structure determination of free PLK4-PB3 and its STIL-CC complex via NMR and crystallography reveals a novel mode of Polo-box-peptide interaction mimicking coiled-coil formation. In vivo analysis of structure-guided STIL mutants reveals distinct binding modes to PLK4-PB3 and L1, as well as interplay of STIL oligomerization with PLK4 binding. We suggest that the STIL-CC/PLK4 interaction mediates PLK4 activation as well as stabilization of centriolar PLK4 and plays a key role in centriole duplication.

No MeSH data available.


Related in: MedlinePlus

Secondary structure elements of PLK4-PB3 in solution.Secondary backbone 13C-chemical shifts of PLK4-PB3 (dark blue) and PLK4-PB3 as part of the PLK4-PB3/STIL–CC complex (light blue) and their differences (black), plotted against the amino acid residue number of PLK4-PB3. Asterisks indicate unassigned residues. Secondary structural elements as identified in the PLK4-PB3/STIL-CC crystal structure are shown at the top. STIL–CC binding increases the helicity at residues 954 and 955 in helix α1, as indicated by an increase of secondary 13C chemical shifts.DOI:http://dx.doi.org/10.7554/eLife.07888.013
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fig5s3: Secondary structure elements of PLK4-PB3 in solution.Secondary backbone 13C-chemical shifts of PLK4-PB3 (dark blue) and PLK4-PB3 as part of the PLK4-PB3/STIL–CC complex (light blue) and their differences (black), plotted against the amino acid residue number of PLK4-PB3. Asterisks indicate unassigned residues. Secondary structural elements as identified in the PLK4-PB3/STIL-CC crystal structure are shown at the top. STIL–CC binding increases the helicity at residues 954 and 955 in helix α1, as indicated by an increase of secondary 13C chemical shifts.DOI:http://dx.doi.org/10.7554/eLife.07888.013

Mentions: To further characterize differences in structure and dynamics of PLK4-PB3 seen upon STIL-CC binding in aqueous solution, free and STIL-CC-bound PLK4-PB3 were subjected to 2D [15N,1H]-TROSY experiments to reveal chemical shift perturbations and to 15N-{1H}NOE measurements to characterize backbone dynamics on the ps- to ns-timescale (Figure 5—figure supplements 1, 2). Chemical shift perturbations are observed throughout most of the PB3 backbone and comprise direct STIL-CC interactions and perpetuated structural changes throughout PLK4-PB3. The most significant changes locate however around residues C954 and L955 on helix α1, where a slight kink is formed in apo PLK4-PB3. Notably, the secondary chemical shifts of these residues are substantially increased upon binding STIL-CC, suggesting a stabilization of helical conformation (Figure 5—figure supplement 3). STIL-CC binding leads to an increase in averaged heteronuclear NOEs for β-strands and the α1 helix, suggesting a general reduction of fast backbone motions of PLK4-PB3 upon STIL-CC binding. Overall, the structural data reveal two core differences in PLK4-PB3 induced by STIL-CC binding: first, strand β1 is N-terminally extended by three residues to the range 888–893, resulting in a shortening of the unstructured N-terminal region in the STIL-CC complex. Second, helix α1 slightly changes its orientation and is stabilized by STIL-CC binding (Figure 5C).


STIL binding to Polo-box 3 of PLK4 regulates centriole duplication.

Arquint C, Gabryjonczyk AM, Imseng S, Böhm R, Sauer E, Hiller S, Nigg EA, Maier T - Elife (2015)

Secondary structure elements of PLK4-PB3 in solution.Secondary backbone 13C-chemical shifts of PLK4-PB3 (dark blue) and PLK4-PB3 as part of the PLK4-PB3/STIL–CC complex (light blue) and their differences (black), plotted against the amino acid residue number of PLK4-PB3. Asterisks indicate unassigned residues. Secondary structural elements as identified in the PLK4-PB3/STIL-CC crystal structure are shown at the top. STIL–CC binding increases the helicity at residues 954 and 955 in helix α1, as indicated by an increase of secondary 13C chemical shifts.DOI:http://dx.doi.org/10.7554/eLife.07888.013
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4530586&req=5

fig5s3: Secondary structure elements of PLK4-PB3 in solution.Secondary backbone 13C-chemical shifts of PLK4-PB3 (dark blue) and PLK4-PB3 as part of the PLK4-PB3/STIL–CC complex (light blue) and their differences (black), plotted against the amino acid residue number of PLK4-PB3. Asterisks indicate unassigned residues. Secondary structural elements as identified in the PLK4-PB3/STIL-CC crystal structure are shown at the top. STIL–CC binding increases the helicity at residues 954 and 955 in helix α1, as indicated by an increase of secondary 13C chemical shifts.DOI:http://dx.doi.org/10.7554/eLife.07888.013
Mentions: To further characterize differences in structure and dynamics of PLK4-PB3 seen upon STIL-CC binding in aqueous solution, free and STIL-CC-bound PLK4-PB3 were subjected to 2D [15N,1H]-TROSY experiments to reveal chemical shift perturbations and to 15N-{1H}NOE measurements to characterize backbone dynamics on the ps- to ns-timescale (Figure 5—figure supplements 1, 2). Chemical shift perturbations are observed throughout most of the PB3 backbone and comprise direct STIL-CC interactions and perpetuated structural changes throughout PLK4-PB3. The most significant changes locate however around residues C954 and L955 on helix α1, where a slight kink is formed in apo PLK4-PB3. Notably, the secondary chemical shifts of these residues are substantially increased upon binding STIL-CC, suggesting a stabilization of helical conformation (Figure 5—figure supplement 3). STIL-CC binding leads to an increase in averaged heteronuclear NOEs for β-strands and the α1 helix, suggesting a general reduction of fast backbone motions of PLK4-PB3 upon STIL-CC binding. Overall, the structural data reveal two core differences in PLK4-PB3 induced by STIL-CC binding: first, strand β1 is N-terminally extended by three residues to the range 888–893, resulting in a shortening of the unstructured N-terminal region in the STIL-CC complex. Second, helix α1 slightly changes its orientation and is stabilized by STIL-CC binding (Figure 5C).

Bottom Line: STIL-CC is the first identified interaction partner of Polo-box 3 (PB3) of PLK4 and also uses a secondary interaction site in the PLK4 L1 region.In vivo analysis of structure-guided STIL mutants reveals distinct binding modes to PLK4-PB3 and L1, as well as interplay of STIL oligomerization with PLK4 binding.We suggest that the STIL-CC/PLK4 interaction mediates PLK4 activation as well as stabilization of centriolar PLK4 and plays a key role in centriole duplication.

View Article: PubMed Central - PubMed

Affiliation: Biozentrum, University of Basel, Basel, Switzerland.

ABSTRACT
Polo-like kinases (PLK) are eukaryotic regulators of cell cycle progression, mitosis and cytokinesis; PLK4 is a master regulator of centriole duplication. Here, we demonstrate that the SCL/TAL1 interrupting locus (STIL) protein interacts via its coiled-coil region (STIL-CC) with PLK4 in vivo. STIL-CC is the first identified interaction partner of Polo-box 3 (PB3) of PLK4 and also uses a secondary interaction site in the PLK4 L1 region. Structure determination of free PLK4-PB3 and its STIL-CC complex via NMR and crystallography reveals a novel mode of Polo-box-peptide interaction mimicking coiled-coil formation. In vivo analysis of structure-guided STIL mutants reveals distinct binding modes to PLK4-PB3 and L1, as well as interplay of STIL oligomerization with PLK4 binding. We suggest that the STIL-CC/PLK4 interaction mediates PLK4 activation as well as stabilization of centriolar PLK4 and plays a key role in centriole duplication.

No MeSH data available.


Related in: MedlinePlus