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Development ePress online publication date 14 Sep 2005
doi: 10.1242/dev.02039
Research article
Cortical localization of the G
protein GPA-16 requires RIC-8 function during C. elegans asymmetric cell division
Katayoun Afshar,
Francis S. Willard,
Kelly Colombo,
David P. Siderovski,
and
Pierre Gönczy*
* Author for correspondence (e-mail: pierre.gonczy{at}isrec.unil.ch)
Understanding of the mechanisms governing spindle positioning during asymmetric division remains incomplete. During unequal division of one-cell stage C. elegans embryos, the G
proteins GOA-1 and GPA-16 act in a partially redundant manner to generate pulling forces along astral microtubules. Previous work focused primarily on GOA-1, whereas the mechanisms by which GPA-16 participates in this process are not well understood. Here, we report that GPA-16 is present predominantly at the cortex of one-cell stage embryos. Using co-immunoprecipitation and surface plasmon resonance binding assays, we find that GPA-16 associates with RIC-8 and GPR-1/2, two proteins known to be required for pulling force generation. Using spindle severing as an assay for pulling forces, we demonstrate that inactivation of the G
protein GPB-1 renders GPA-16 and GOA-1 entirely redundant. This suggests that the two G proteins can activate the same pathway and that their dual presence is normally needed to counter G
. Using nucleotide exchange assays, we establish that whereas GPR-1/2 acts as a guanine nucleotide dissociation inhibitor (GDI) for GPA-16, as it does for GOA-1, RIC-8 does not exhibit guanine nucleotide exchange factor (GEF) activity towards GPA-16, in contrast to its effect on GOA-1. We establish in addition that RIC-8 is required for cortical localization of GPA-16, whereas it is not required for that of GOA-1. Our analysis demonstrates that this requirement toward GPA-16 is distinct from the known function of RIC-8 in enabling interaction between G proteins and GPR-1/2, thus providing novel insight into the mechanisms of asymmetric spindle positioning.
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© The Company of Biologists Ltd 2005
