scholarly journals Genetic Evidence for a Protein Kinase A/Cubitus Interruptus Complex That Facilitates Processing of Cubitus Interruptus in Drosophila

Genetics ◽  
2001 ◽  
Vol 158 (3) ◽  
pp. 1157-1166
Author(s):  
John A Kiger ◽  
Cristin O'Shea
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Target Gene ◽  
Target Genes ◽  
Signal Transduction Pathway ◽  
Transduction Pathway ◽  
Cubitus Interruptus ◽  
Mutant Forms ◽  
The Relationship ◽  
Kinase A

Abstract Hedgehog (Hh) activates a signal transduction pathway regulating Cubitus interruptus (Ci). In the absence of Hh, full-length Ci (Ci-155) is bound in a complex that includes Costal2 (Cos2) and Fused (Fu). Ci-155 is phosphorylated by protein kinase A (PKA), inducing proteolysis to Ci-75, a transcriptional repressor. Hh signaling blocks proteolysis and produces an activated Ci-155 transcriptional activator. The relationship between PKA and the Ci/Cos2/Fu complex is unclear. Here we examine Hh target gene expression caused by mutant forms of PKA regulatory (PKAr) and catalytic (PKAc) subunits and by the PKAc inhibitor PKI(1-31). The mutant PKAr*, defective in binding cAMP, is shown to activate Hh target genes solely through its ability to bind and inhibit endogenous PKAc. Surprisingly, PKAcA75, a catalytically impaired mutant, also activates Hh target genes. To account for this observation, we propose that PKAc phosphorylation targeting Ci-155 for proteolysis is regulated within a complex that includes PKAc and Ci-155 and excludes PKI(1-31). This complex may permit processive phosphorylation of Ci-155 molecules, facilitating their processing to Ci-75.

Proteolysis of cubitus interruptus in Drosophila requires phosphorylation by protein kinase A

Development ◽  
1999 ◽  
Vol 126 (19) ◽  
pp. 4331-4339 ◽  
Author(s):  
M.A. Price ◽  
D. Kalderon
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Hedgehog Signaling ◽  
Target Genes ◽  
Ectopic Expression ◽  
Full Length ◽  
Mutant Form ◽  
Wild Type ◽  
Cubitus Interruptus ◽  
Kinase A

The Hedgehog signal transduction pathway is involved in diverse patterning events in many organisms. In Drosophila, Hedgehog signaling regulates transcription of target genes by modifying the activity of the DNA-binding protein Cubitus interruptus (Ci). Hedgehog signaling inhibits proteolytic cleavage of full-length Ci (Ci-155) to Ci-75, a form that represses some target genes, and also converts the full-length form to a potent transcriptional activator. Reduction of protein kinase A (PKA) activity also leads to accumulation of full-length Ci and to ectopic expression of Hedgehog target genes, prompting the hypothesis that PKA might normally promote cleavage to Ci-75 by directly phosphorylating Ci-155. Here we show that a mutant form of Ci lacking five potential PKA phosphorylation sites (Ci5m) is not detectably cleaved to Ci-75 in Drosophila embryos. Moreover, changes in PKA activity dramatically altered levels of full-length wild-type Ci in embryos and imaginal discs, but did not significantly alter full-length Ci5m levels. We corroborate these results by showing that Ci5m is more active than wild-type Ci at inducing ectopic transcription of the Hh target gene wingless in embryos and that inhibition of PKA enhances induction of wingless by wild-type Ci but not by Ci5m. We therefore propose that PKA phosphorylation of Ci is required for the proteolysis of Ci-155 to Ci-75 in vivo. We also show that the activity of Ci5m remains Hedgehog responsive if expressed at low levels, providing further evidence that the full-length form of Ci undergoes a Hedgehog-dependent activation step.

scholarly journals An absolute requirement for Cubitus interruptus in Hedgehog signaling

Development ◽  
2001 ◽  
Vol 128 (5) ◽  
pp. 733-742 ◽  
Author(s):  
N. Methot ◽  
K. Basler
Keyword(s):  
Gene Expression ◽  
Signal Transduction ◽  
Hedgehog Signaling ◽  
Target Gene ◽  
Target Genes ◽  
Signal Transduction Pathway ◽  
Transduction Pathway ◽  
Cubitus Interruptus ◽  
Target Gene Expression ◽  
Hh Signaling

Hedgehog (Hh) proteins play diverse organizing roles in animal development by regulating gene expression in responding cells. Several components of the Hh signal transduction pathway have been identified, yet their precise role in mediating the various outputs of the pathway is still poorly understood. The Gli homolog Cubitus interruptus (Ci) is involved in controlling the transcription of Drosophila Hh target genes and thus represents the most downstream component known in this pathway. We address the question of whether the Hh pathway is distally branched or, in other words, whether the regulation of Ci activity is the sole output of Hh signaling. Putative Ci-independent branches of Hh signaling are explored by analyzing the behavior of cells that lack Ci but have undergone maximal activation of the Hh transduction pathway due to the removal of Patched (Ptc). The analysis of target gene expression and morphogenetic read-outs of Hh in embryonic, larval and adult stages indicates that Ci is absolutely required for all examined aspects of Hh outputs. We interpret this as evidence against the existence of Ci-independent branches in the Hh signal transduction pathway and propose that most cases of apparent Ci/Gli-independent Hh output can be attributed to the derepression of target gene expression in the absence of Ci/Gli repressor function.

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