The cytoskeletal regulator Genghis khan is required for columnar target specificity in theDrosophilavisual system

Abstract

SUMMARYA defining characteristic of neuronal cell type is the growth of axons and dendrites into specific layers and columns of the brain. Although differences in cell surface receptors and adhesion molecules are known to cause differences in synaptic specificity, differences in downstream signaling mechanisms that determine cell type-appropriate targeting patterns are unknown. Using a forward genetic screen in Drosophila, we identify the GTPase effector Genghis khan (Gek) as playing a crucial role in the ability of a subset of photoreceptor (R cell) axons to innervate appropriate target columns. In particular, single-cell mosaic analyses demonstrate that R cell growth cones lacking Gek function grow to the appropriate ganglion, but frequently fail to innervate the correct target column. Further studies reveal that R cell axons lacking the activity of the small GTPase Cdc42 display similar defects, providing evidence that these proteins regulate a common set of processes. Gek is expressed in all R cells, and a detailed structure-function analysis reveals a set of regulatory domains with activities that restrict Gek function to the growth cone. Although Gek does not normally regulate layer-specific targeting, ectopic expression of Gek is sufficient to alter the targeting choices made by another R cell type, the targeting of which is normally Gek independent. Thus, specific regulation of cytoskeletal responses to targeting cues is necessary for cell type-appropriate synaptic specificity.

An apical MRCK-driven morphogenetic pathway controls epithelial polarity

“…2a ). Strikingly, differentiating photoreceptors displayed active Myosin caps at the apical cortex, which were not formed in cells mutant for gek /MRCK 25 ( Fig. 2b ).…”

“…Flies were maintained at 25°C on standard food. To generate whole mutant eyes, the EGUF/GMRhid system was used 45 in combination with the gek NP5192 and gek omb1080 alleles 25 . Canton S flies were used as control.…”

“…Whole mount retinas were prepared as described in 48 . The following antibodies were used: rabbit anti-p-MLC Ser19, 1/40 (Cell Signaling Technology 3671); rabbit anti-PKCζ 1/200 (Sigma Aldrich SAB4502380), mouse anti-Arm 1/200 (N27-A1, Developmental Studies Hybridoma Bank), rabbit anti-Gek 1/25 (gift from TR Clandinin 25 , rabbit anti-P-Moesin 1/500 49 , phalloidin-Texas Red (Sigma Aldrich, 300nM) with the appropriate combination of secondary antibodies conjugated to Dy405, Alexa488, Cy3 or Cy5 as appropriate at 1/200 each (Jackson ImmunoResearch). Retinas were mounted in VectaShield and imaging was performed using a Leica SP5 confocal.…”

Gogo Receptor Contributes to Retinotopic Map Formation and Prevents R1-6 Photoreceptor Axon Bundling

“…Several studies revealed that afferent-afferent interactions among R axon growth cones of the same fascicle mediate their specific lateral directions during cartridge selection [6], [9], [17], [27], [31], [34], [35], [36]. However, Gogo is not mediating afferent-afferent interactions between R1–R6 growth cones as loss of Gogo in single R cells does not affect target cartridge selection.…”

The cell biology of synaptic specificity during development

“…The actin cytoskeleton is also linked to presynaptic assembly, as disruption of F-actin in C. elegans can affect presynaptic assembly in vivo [77 • ] (Figure 3b). In the Drosophila visual system, the cytoskeletal regulatory protein Ghengis Khan (GEK) is required for columnar targeting specificity [78 • ]. The actin cytoskeleton underlies other neurodevelopmental programs besides synaptic specificity, including morphogenesis, cell migration and guidance.…”