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The Caenorhabditis elegans protein SAS-5 forms large oligomeric assemblies critical for centriole formation.

Rogala KB, Dynes NJ, Hatzopoulos GN, Yan J, Pong SK, Robinson CV, Deane CM, Gönczy P, Vakonakis I - Elife (2015)

Bottom Line: Here, we combine biophysical and structural analysis to uncover the architecture of SAS-5 and examine its functional implications in vivo.Our work reveals that two distinct self-associating domains are necessary to form higher-order oligomers of SAS-5: a trimeric coiled coil and a novel globular dimeric Implico domain.Disruption of either domain leads to centriole duplication failure in worm embryos, indicating that large SAS-5 assemblies are necessary for function in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, University of Oxford, Oxford, United Kingdom.

ABSTRACT
Centrioles are microtubule-based organelles crucial for cell division, sensing and motility. In Caenorhabditis elegans, the onset of centriole formation requires notably the proteins SAS-5 and SAS-6, which have functional equivalents across eukaryotic evolution. Whereas the molecular architecture of SAS-6 and its role in initiating centriole formation are well understood, the mechanisms by which SAS-5 and its relatives function is unclear. Here, we combine biophysical and structural analysis to uncover the architecture of SAS-5 and examine its functional implications in vivo. Our work reveals that two distinct self-associating domains are necessary to form higher-order oligomers of SAS-5: a trimeric coiled coil and a novel globular dimeric Implico domain. Disruption of either domain leads to centriole duplication failure in worm embryos, indicating that large SAS-5 assemblies are necessary for function in vivo.

No MeSH data available.


The SAS-5 architecture allows formation of higher oligomers.Schematic representation of SAS-5 oligomerization, showing the coiled-coil and Implico domains, without the disordered N- and C-termini of this protein. Tight dimers of SAS-5, mediated by the Implico domain, could come together through the coiled coil to form, first, a tetramer and ultimately a hexamer in a concentration-dependent manner. This oligomerisation model does not take into account protein aggregation mediated by the predicted β-strand at the SAS-5 C-terminus.DOI:http://dx.doi.org/10.7554/eLife.07410.026
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fig7: The SAS-5 architecture allows formation of higher oligomers.Schematic representation of SAS-5 oligomerization, showing the coiled-coil and Implico domains, without the disordered N- and C-termini of this protein. Tight dimers of SAS-5, mediated by the Implico domain, could come together through the coiled coil to form, first, a tetramer and ultimately a hexamer in a concentration-dependent manner. This oligomerisation model does not take into account protein aggregation mediated by the predicted β-strand at the SAS-5 C-terminus.DOI:http://dx.doi.org/10.7554/eLife.07410.026

Mentions: SAS-5 concentrates at centrioles, as illustrated by well-defined GFP foci of this protein (Figure 6F,I), and we propose that SAS-5 dimers associate further to form higher-order assemblies there. Our SEC-MALS results show that SAS-5 forms tetramers already at 1 μM concentration and a mixture of tetramers and hexamers at 10 μM (Figure 4B). Although the SAS-5 architecture, with two independent oligomerization interfaces, could theoretically sustain the formation of open-ended protein chains or networks, we believe it is more likely that higher-order SAS-5 assemblies cap at a hexameric size. Such hexamers, formed through the sequential association of three SAS-5 dimers (Figure 7), should be both kinetically and entropically favored compared to open-ended SAS-5 chains. Hexamer breakdown would require simultaneous disruption of two SAS-5 coiled coils, a rarer event compared to the disruption of a single coiled coil in an open-ended SAS-5 oligomer. Further, in the sequential assembly model of Figure 7, once the first coiled-coil sequence of a SAS-5 dimer has associated into a folded coiled-coil structure, the degrees of freedom of the second SAS-5 coiled-coil sequence would be reduced, and thus its association into the second folded coiled-coil structure would become entropically favored.10.7554/eLife.07410.026Figure 7.The SAS-5 architecture allows formation of higher oligomers.


The Caenorhabditis elegans protein SAS-5 forms large oligomeric assemblies critical for centriole formation.

Rogala KB, Dynes NJ, Hatzopoulos GN, Yan J, Pong SK, Robinson CV, Deane CM, Gönczy P, Vakonakis I - Elife (2015)

The SAS-5 architecture allows formation of higher oligomers.Schematic representation of SAS-5 oligomerization, showing the coiled-coil and Implico domains, without the disordered N- and C-termini of this protein. Tight dimers of SAS-5, mediated by the Implico domain, could come together through the coiled coil to form, first, a tetramer and ultimately a hexamer in a concentration-dependent manner. This oligomerisation model does not take into account protein aggregation mediated by the predicted β-strand at the SAS-5 C-terminus.DOI:http://dx.doi.org/10.7554/eLife.07410.026
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4471805&req=5

fig7: The SAS-5 architecture allows formation of higher oligomers.Schematic representation of SAS-5 oligomerization, showing the coiled-coil and Implico domains, without the disordered N- and C-termini of this protein. Tight dimers of SAS-5, mediated by the Implico domain, could come together through the coiled coil to form, first, a tetramer and ultimately a hexamer in a concentration-dependent manner. This oligomerisation model does not take into account protein aggregation mediated by the predicted β-strand at the SAS-5 C-terminus.DOI:http://dx.doi.org/10.7554/eLife.07410.026
Mentions: SAS-5 concentrates at centrioles, as illustrated by well-defined GFP foci of this protein (Figure 6F,I), and we propose that SAS-5 dimers associate further to form higher-order assemblies there. Our SEC-MALS results show that SAS-5 forms tetramers already at 1 μM concentration and a mixture of tetramers and hexamers at 10 μM (Figure 4B). Although the SAS-5 architecture, with two independent oligomerization interfaces, could theoretically sustain the formation of open-ended protein chains or networks, we believe it is more likely that higher-order SAS-5 assemblies cap at a hexameric size. Such hexamers, formed through the sequential association of three SAS-5 dimers (Figure 7), should be both kinetically and entropically favored compared to open-ended SAS-5 chains. Hexamer breakdown would require simultaneous disruption of two SAS-5 coiled coils, a rarer event compared to the disruption of a single coiled coil in an open-ended SAS-5 oligomer. Further, in the sequential assembly model of Figure 7, once the first coiled-coil sequence of a SAS-5 dimer has associated into a folded coiled-coil structure, the degrees of freedom of the second SAS-5 coiled-coil sequence would be reduced, and thus its association into the second folded coiled-coil structure would become entropically favored.10.7554/eLife.07410.026Figure 7.The SAS-5 architecture allows formation of higher oligomers.

Bottom Line: Here, we combine biophysical and structural analysis to uncover the architecture of SAS-5 and examine its functional implications in vivo.Our work reveals that two distinct self-associating domains are necessary to form higher-order oligomers of SAS-5: a trimeric coiled coil and a novel globular dimeric Implico domain.Disruption of either domain leads to centriole duplication failure in worm embryos, indicating that large SAS-5 assemblies are necessary for function in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, University of Oxford, Oxford, United Kingdom.

ABSTRACT
Centrioles are microtubule-based organelles crucial for cell division, sensing and motility. In Caenorhabditis elegans, the onset of centriole formation requires notably the proteins SAS-5 and SAS-6, which have functional equivalents across eukaryotic evolution. Whereas the molecular architecture of SAS-6 and its role in initiating centriole formation are well understood, the mechanisms by which SAS-5 and its relatives function is unclear. Here, we combine biophysical and structural analysis to uncover the architecture of SAS-5 and examine its functional implications in vivo. Our work reveals that two distinct self-associating domains are necessary to form higher-order oligomers of SAS-5: a trimeric coiled coil and a novel globular dimeric Implico domain. Disruption of either domain leads to centriole duplication failure in worm embryos, indicating that large SAS-5 assemblies are necessary for function in vivo.

No MeSH data available.