(A) Schematic of the adult morphologies of R7 and R8 photoreceptors and lamina neurons L1–L5. The visual system comprises topographically matched modules (i.e., ommatidia, cartridges and columns). Adapted from .
(B) Axons of R7, R8, and L1–L5 are shown together within a single column as determined from serial EM reconstruction. The color of different neurons is the same as in (A). Dotted lines represent layer boundaries. Arrowheads indicate the plane of section shown in electron micrographs in (D) and (E). Courtesy of S. Takemura, I. Meinertzhagen, and L. Scheffer (JRC/HHMI).
(C) Examples of three general classes of medulla neurons that are synaptic targets for R7, R8, and L1–L5 neurons (two examples are shown for each class). Within each class, there are many cell types that display similar morphologies and branch in different layers. Adapted from .
(D and E) Cross-sections through medulla columns reconstructed by serial EM within the M2 and M3 layers (see arrows in B). Axons are colored as in (A) and (B). Each column contains processes from over 100 different neuronal cell types (A. Nern, personal communication). Scale bar, 2 µm. Courtesy of S. Takemura, I. Meinertzhagen, and L. Scheffer (JRC/HHMI).
(F and G) Isolation of R8 neurons at 40 hr APF using fluorescence-activated cell sorting (FACS). Only R8 neurons express both retinal-specific TdTom and R8-specific GFP. Senseless is an R8-specific transcription factor. Scale bars, 10 µm. In (F), arrows indicate double-labeled cells in developing tissue and the asterisks indicate single GFP-labeled cells of different cell types (i.e., contaminants).
(H and I) Isolation of L3 neurons at 40 hr APF using FACS. Only L3 neurons express lamina-specific TdTom and L3-specific GFP. Scale bars, 10 µm. In (H), arrows indicate double-labeled cells in developing tissue and the asterisks indicate single GFP-labeled cells of different cell types (i.e., contaminants). See Experimental Procedures for purification protocols for other cells and additional details.

(A) Correlograms showing the correlation score matrix across all libraries of all seven cell types (R, Pearson correlation coefficient).
(B) Principal component analysis plot of the RNA-seq data for the indicated cell types. Each red dot represents an RNA-seq sample.
(C) RPKM values (left) and antibody staining (right) for transcription factors in the lamina (L1–L5) and retina (R7, R8) at 40 hr APF. Cell-type-specific markers are shown in green and antibodies for cell-type-specific transcription factors are shown in red (L1–L5 and R7) or blue (R8). Arrows in L2 panels indicate glial cells also stained with antibody to Bab2. A general retinal marker is shown in blue and red in the R7 and R8 panels, respectively. Scale bars, 10 µm.
See also .

(A) Heat map showing expression of all genes encoding CSMs expressed in at least one cell with an RPKM > 5 (n = 444). See also .
(B) Heat map representing expression of genes encoding immunoglobulin (Ig) superfamily of cell surface proteins. Each gene in this list is expressed in at least one cell type with an RPKM greater than five and five times greater in one cell type than at least one of the six other cell types. Genes shown in color are members of gene sub-families. Note all Side family members (with the exception of Side) are shown as CG numbers. See also and .
(C) Synaptic connections of L1 and L2 in medulla neuropil. They are largely different. Arrows indicate directionality of connection from pre-synaptic neuron to post-synaptic neuron. For example, L1 is pre-synaptic to C3. And L1 is both pre- and post-synaptic to L5. C2 in red means that C2 is a shared synaptic partner with both L1 and L2. L5 in green means that L5 is also a shared synaptic partner for both L1 and L2.
(D) Numbers of genes exhibiting differences of less than two times (shared), two to five times, and greater than five times in expression between L1 and L2 with RPKM greater than five in at least one cell type, with an adjusted p value < 0.05. Enriched means the level of a gene in one cell type is higher than the other cell type. Numbers of genes in each category are shown. See also .
(E) Lists of genes encoding Ig superfamily cell surface proteins that are enriched in L1 and L2 by greater than five times. RPKM values in L1 and L2 are also listed. CG42313 and CG14372 are Side protein family members. Asterisk indicates that the interacting partner of the protein is not known yet. Interacting partners for all other proteins in this table have been identified (, , and ).
See also and .

(A) Schematic of a MiMIC-based protein trap. MiMIC protein traps contain GFP in frame flanked by splice acceptor and donor sites. They are generated by cassette exchange using ϕC31 recombinase to catalyze recombination with the Minos insertion between the AttP and AttB sites. The green inverted arrows after recombination represent recombined recombination sites (i.e., attR sites). Genes modified in this way encode chimeric proteins containing GFP.
(B and B′) Arrangement of lamina neuron cell bodies at 40 hr and 72 hr APF. L2 and L3 are intermingled at the top of lamina cell clusters. L4 and L5 make up the bottom two rows with L5 beneath L4.
(C–E′) Dpr17, Dpr2, and Dpr13 expression in lamina neurons visualized using MiMIC protein traps. See for lamina neuron markers. Scale bars, 10 µm.
(F and F′) Summary of Dpr expression using protein trap lines (10 of 21 dpr genes). RPKM values from the RNA-seq results indicating level of gene expression are included in (F). Dpr2, Dpr13, and Dpr17 are orange colored in bold to indicate changes in staining with the preceding panels. *Indicates Dpr10 expression level in L5 is variable at 72 hr APF.
See also .

(A) Schematic of three classes of medulla neurons. A transmedullary neuron (Tm, in yellow), an amacrine-like distal medulla neuron (Dm, in red), and a medulla intrinsic neuron (Mi, in magenta) are shown. Each class of medulla neurons can be further divided into specific cell types based on different patterns of layer-specific branching. Adapted from .
(B–C′) DIP protein traps are expressed in scattered cells in the medulla cortex (arrows) and in layer-specific patterns in the medulla neuropil. DIP-β and DIP-η are shown as examples (green). Photoreceptor axons are visualized by staining for the cell surface protein Chp (red).
(D–J) All six DIP genes for which protein trap lines are available were expressed in neurons exhibiting unique layer-specific patterns of processes within the outer medulla neuropil at 40 hr APF. (D) Schematic of R8 and R7 axon morphology and layer distribution in the outer medulla. (E–J) Protein expression of six DIPs (green) in the outer medulla. The six DIPs are expressed in one to three layers; each layer is defined by a unique combination of DIPs.
(K–R) The DIP expression pattern at 72 hr is shown. (K) Schematic of R7 and R8 axons at 72 hr. The medulla expands and R7 and R8 layers change between 40 and 72 hr. (L–Q) The layered expression of DIPs is largely the same as at 40 hr. Expression in an additional layer, however, appears in DIP-θ. Dm3 axons are labeled with td-tomato (magenta). They run parallel to layers and mark the M2 and M3 border. (R) Summary of expression of DIPs in the medulla and the projection of R8, R7, and L1–L5 terminals at 72 hr APF. Scale bars, 10 µm.
See also .

(A–E) Co-localization of DIPs in synaptic partners of L1–L5. Left panels: co-localization of indicated DIP (green) and cell-type-specific marker (red) in the adult. Middle panels: schematic of morphology of lamina neurons (green) and a subset of their synaptic partners (red) within the medulla neuropil. Right panels: summary of Dpr expression in L1–L5 and DIPs in synaptic partners. Layer patterns for DIPs in the medulla are the same as at 72 hr. Synaptic partner assignments from and S. Takemura, I. Meinertzhagen, and L. Scheffer, personal communication.
(F) Summary of the Dpr/DIP interactome ( and ).
See also .