A Screen for X-Linked Mutations Affecting Drosophila Photoreceptor Differentiation Identifies Casein Kinase 1α as an Essential Negative Regulator of Wingless Signaling

Abstract

The Wnt and Hedgehog signaling pathways are essential for normal development and are misregulated in cancer. The casein kinase family of serine/threonine kinases regulates both pathways at multiple levels. However, it has been difficult to determine whether individual members of this family have distinct functions in vivo, due to their overlapping substrate specificities. In Drosophila melanogaster, photoreceptor differentiation is induced by Hedgehog and inhibited by Wingless, providing a sensitive system in which to identify regulators of each pathway. We used a mosaic genetic screen in the Drosophila eye to identify mutations in genes on the X chromosome required for signal transduction. We recovered mutations affecting the transcriptional regulator CREB binding protein, the small GTPase dynamin, the cytoskeletal regulator Actin-related protein 2, and the protein kinase Casein kinase 1a. Consistent with its reported function in the b-Catenin degradation complex, Casein Kinase 1a mutant cells accumulate b-Catenin and ectopically induce Wingless target genes. In contrast to previous studies based on RNA interference, we could not detect any effect of the same Casein Kinase 1a mutation on Hedgehog signaling. We thus propose that Casein kinase 1a is essential to allow b-Catenin degradation and prevent inappropriate Wingless signaling, but its effects on the Hedgehog pathway are redundant with other Casein kinase 1 family members.

Drosophila Vps4 promotes Epidermal growth factor receptor signaling independently of its role in receptor degradation

“…In a mosaic screen of the X chromosome (Legent et al, 2012), we recovered a mutation that prevents photoreceptor differentiation. In eye imaginal discs, clones of 3B1 mutant cells showed a cellautonomous lack of expression of the pan-neuronal marker Elav (Fig.…”

“…Although some studies have demonstrated that it contributes to signal termination through lysosomal EGFR degradation (Bache et al, 2006;Razi and Futter, 2006), other results argue that it enables receptor signaling from intracellular organelles (Miaczynska et al, 2004;Shilo and Schejter, 2011). Vps4 clones in Dronc mutant eye discs resemble clones lacking positive intracellular effectors of EGFR transduction, in which R8 cells fail to recruit additional photoreceptors (Legent et al, 2012). We found that the expression of lacZ reporters for two direct transcriptional targets of the EGFR pathway in R1-R7, argos (aos) (Golembo et al, 1996) and hedgehog (hh) (Rogers et al, 2005), was downregulated in Vps4 mutant clones (Fig.…”

“…We therefore generated double mutant clones for Vps4 and shibire (shi), which encodes Dynamin, a GTPase required to internalize EGFR and other receptors from the plasma membrane (Henriksen et al, 2013;Sousa et al, 2012;Windler and Bilder, 2010). shi 7C7 mutant cells have enhanced EGFR signaling, leading to increased photoreceptor differentiation (Legent et al, 2012) (Fig. 6A).…”

“…Adult eye phenotypes thus provide a rapid screen for new regulators of the EGFR pathway (Legent et al, 2012;Miura et al, 2008;Roignant et al, 2006;Roignant and Treisman, 2010). Here, we report that a mutation in Vps4 disrupts EGFR signal transduction as well as the transduction of other signaling pathways.…”

“…Blocking EGFR internalization by removing Dynamin prevents its degradation, enhancing some downstream signaling events, but other aspects of EGFR signal transduction require an endosomal localization for the receptor (Jones and Rappoport, 2014;Legent et al, 2012;Miura et al, 2008;Teis et al, 2006;Vieira et al, 1996). Internalized EGFR can be either degraded or recycled to the plasma membrane, a choice that depends on the concentration and nature of the ligand, as ligands that remain bound in acidic late endosomes promote more extensive receptor ubiquitylation (Eden et al, 2012;French et al, 1995;Roepstorff et al, 2009;Sigismund et al, 2005).…”