Abstract
Heterotrimeric G proteins, composed of α, β, and γ subunits, are activated by exchange of GDP for GTP on the Gα subunit. Canonically, Gα is stimulated by the guanine-nucleotide exchange factor (GEF) activity of ligand-bound G protein-coupled receptors. However, Gα subunits may also be activated in a noncanonical manner by members of the Ric-8 family, cytoplasmic proteins that also act as GEFs for Gα subunits. We used a signaling pathway active during Drosophila gastrulation as a model system to study Ric-8/Gα interactions. A component of this pathway, the Drosophila Gα12/13 subunit, Concertina (Cta), is necessary to trigger actomyosin contractility during gastrulation events. Ric-8 mutants exhibit similar gastrulation defects to Cta mutants. Here we use a novel tissue culture system to study a signaling pathway that controls cytoskeletal rearrangements necessary for cellular morphogenesis. We show that Ric-8 regulates this pathway through physical interaction with Cta and preferentially interacts with inactive Cta and directs its localization within the cell. We also use this system to conduct a structure-function analysis of Ric-8 and identify key residues required for both Cta interaction and cellular contractility.
Publication types
-
Research Support, N.I.H., Extramural
-
Research Support, Non-U.S. Gov't
MeSH terms
-
Amino Acid Sequence
-
Animals
-
Binding Sites / genetics
-
Cell Line
-
Drosophila Proteins / chemistry
-
Drosophila Proteins / genetics
-
Drosophila Proteins / metabolism*
-
Drosophila melanogaster / cytology
-
Drosophila melanogaster / genetics
-
Drosophila melanogaster / metabolism
-
GTP-Binding Protein alpha Subunits, G12-G13 / chemistry
-
GTP-Binding Protein alpha Subunits, G12-G13 / genetics
-
GTP-Binding Protein alpha Subunits, G12-G13 / metabolism*
-
Gastrulation / genetics
-
Gastrulation / physiology*
-
Green Fluorescent Proteins / genetics
-
Green Fluorescent Proteins / metabolism
-
Guanine Nucleotide Exchange Factors / chemistry
-
Guanine Nucleotide Exchange Factors / genetics
-
Guanine Nucleotide Exchange Factors / metabolism*
-
Immunoblotting
-
Microscopy, Fluorescence
-
Models, Molecular
-
Molecular Sequence Data
-
Mutation
-
Protein Binding / genetics
-
Protein Structure, Tertiary
-
RNA Interference
-
Sequence Homology, Amino Acid
-
Signal Transduction / genetics
-
Signal Transduction / physiology*
Substances
-
Drosophila Proteins
-
FOG protein, Drosophila
-
Guanine Nucleotide Exchange Factors
-
ric8a protein, Drosophila
-
Green Fluorescent Proteins
-
GTP-Binding Protein alpha Subunits, G12-G13
-
cta protein, Drosophila