Genetic analysis demonstrates a direct link between rho signaling and nonmuscle myosin function during Drosophila morphogenesis.
A dynamic actomyosin cytoskeleton drives many morphogenetic events. Conventional nonmuscle myosin-II (myosin) is a key chemomechanical motor that drives contraction of the actin cytoskeleton. We have explored the regulation of myosin activity by performing genetic screens to identify gene products that collaborate with myosin during Drosophila morphogenesis. Specifically, we screened for second-site noncomplementors of a mutation in the zipper gene that encodes the nonmuscle myosin-II heavy chain. We determined that a single missense mutation in the zipper(Ebr) allele gives rise to its sensitivity to second-site noncomplementation. We then identify the Rho signal transduction pathway as necessary for proper myosin function. First we show that a lethal P-element insertion interacts genetically with zipper. Subsequently we show that this second-site noncomplementing mutation disrupts the RhoGEF2 locus. Next, we show that two EMS-induced mutations, previously shown to interact genetically with zipper(Ebr), disrupt the RhoA locus. Further, we have identified their molecular lesions and determined that disruption of the carboxyl-terminal CaaX box gives rise to their mutant phenotype. Finally, we show that RhoA mutations themselves can be utilized in genetic screens. Biochemical and cell culture analyses suggest that Rho signal transduction regulates the activity of myosin. Our studies provide direct genetic proof of the biological relevance of regulation of myosin by Rho signal transduction in an intact metazoan.
Halsell
S R
SR
Department of Cell Biology, Duke University Medical Center, Durham, North Carolina 27710, USA.
Chu
B I
BI
Kiehart
D P
DP
D000483:Alleles; D000818:Animals; D002875:Chromosomes / Q000235:genetics; D029721:Drosophila Proteins; D004331:Drosophila melanogaster / Q000196:embryology* / Q000235:genetics* / Q000378:metabolism; D005816:Genetic Complementation Test; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism; D015151:Immunoblotting; D008565:Membrane Proteins / Q000235:genetics; D009024:Morphogenesis / Q000235:genetics; D020125:Mutation, Missense; D018995:Myosin Heavy Chains / Q000235:genetics; D009218:Myosins / Q000378:metabolism*; D020161:Physical Chromosome Mapping; D017434:Protein Structure, Tertiary / Q000235:genetics; D064067:Rho Guanine Nucleotide Exchange Factors; D015398:Signal Transduction / Q000235:genetics*; D020742:rhoA GTP-Binding Protein / Q000235:genetics*
D029721:Drosophila Proteins; D020662:Guanine Nucleotide Exchange Factors; D008565:Membrane Proteins; D064067:Rho Guanine Nucleotide Exchange Factors; C097862:Zip protein, Drosophila; D018995:Myosin Heavy Chains; D009218:Myosins; D020742:rhoA GTP-Binding Protein