Cell migration in Drosophila optic lobe neurons is controlled by eyeless/Pax6

Abstract

In the developing Drosophila optic lobe, eyeless, apterous and distal-less, three genes that encode transcription factors with important functions during development, are expressed in broad subsets of medulla neurons. Medulla cortex cells follow two patterns of cell movements to acquire their final position: first, neurons are arranged in columns below each neuroblast. Then, during pupation, they migrate laterally, intermingling with each other to reach their retinotopic position in the adult optic lobe. eyeless, which encodes a Pax6 transcription factor, is expressed early in progenitors and controls aspects of this cell migration. Its loss in medulla neurons leads to overgrowth and a failure of lateral migration during pupation. These defects in cell migration among medulla cortex cells can be rescued by removing DE-Cadherin. Thus, eyeless links neurogenesis and neuronal migration.

Fig. 1.

Medulla cortex organogenesis. (A,C) Anterior (A) and middle (C) sections of late L3 larval brains. Neuroepithelial cells were visualized with DE-Cadherin (blue) and neuroblasts with Deadpan (red). Solid white line in A separates optic lobe from central brain (CB). IPC, inner proliferation center; OPC, outer proliferation center. (B,D,E) Expression patterns of Eyeless (green), Distal-less (red) and ap-lacZ (blue) in cell populations in anterior (B) and middle (D,E) sections of late L3 larvae. Arrowheads in E indicate Ey expression in postmitotic cells and arrows indicate Ey-positive neuroblasts. (F,G) Expression pattern of Ey (F) and ey-Gal4 driving UAS-H2B-YFP (G) in OPC-derived neuroblasts (red). Neuroblasts were labeled with Miranda in F and Deadpan in G. Neuroepithelial cells were visualized with DE-Cadherin (blue). Arrows indicate neuroblasts. (H) Neurogenesis in the OPC. Lamina precursor cells (LPC) differentiate into lamina neurons (gray) in the most lateral part of the neuroepithelium (NE) (yellow). Medulla neurons (MeC) derive from eyeless neuroblasts (Nb) on the medial side of the NE. Postmitotic Ey-positive cells (green), ap-positive cells (blue), Dll-positive cells (red) and other cell types (white) are shown. A, anterior; D, dorsal; M, medial; L, lateral. (I-L) Expression patterns of Ey (green), Dll (red) and ap-lacZ (blue) in cell populations in anterior sections at P0 (I), P20 (J,K) and adult (L).

Fig. 2.

G-TRACE analysis of OPC-derived neurons. (A-L) G-TRACE analysis with ey-Gal4 (A-D), ap-Gal4 (E-H) and dll-Gal4 (I-L) reporter lines. Images show OPC-derived cells at late L3 (A,B,E,F,I,J) and P40 (C,D,G,H,K,L). Neuroepithelial cells in A,B,E,F,I,J were labeled with DE-Cadherin (blue). Arrowheads in C,D indicate persisting ey expression in pupal cells (red and green) and arrows indicate cells with previous ey expression that no longer express the reporter (green).

Fig. 3.

Cell migration pattern of eyeless and apterous cells. (A-F,H-J) Migration patterns of ap-(A-C), ey-(D-F) and ey-misexpressing ey^DN (H-J) MARCM clones (green) generated 72 hours AEL and analyzed in late L3 (A,D,H), in P44 (B,E,I) and in P68 (C,F,J). Brackets in B,C,E,F indicate layers of medulla neuropils. Dotted circles in I,J highlight the accumulation of neurons in large clumps. Neurons were visualized with Elav (red) and the neuropil with nc82 (blue). (G,K) Proliferation analysis in ey-expressing (G) and ey-misexpressing ey^DN (K) MARCM clones (green) generated 72 hours AEL and analyzed at P5. Mitosis was visualized with anti-phospho Histone 3 (red) and the neuropil with DN-Cad (blue).

Fig. 4.

Cell migration pattern of clonally related cells. (A-C) Migration patterns of tub-MARCM clones (green) generated 72 hours AEL and analyzed in late L3 (A), P10 (B) and P44 (C). Dll-positive cells visualized with anti-Dll (red) and ap-positive cells with ap-lacZ (blue). White arrows in A-C mark ap-positive cells, green arrows mark GFP-positive cells negative for ap and arrowheads label Dll-positive cells. (D) Schematic representation of a larval `column' in the OPC. During pupation, these OPC-derived cells lose their columnar organization and become dispersed throughout the medulla cortex with non-lineage-related cells (gray cells).

Fig. 5.

Impaired optic lobe cell migration in ey^DN MARCM clones rescued by DE-Cadherin. (A-D) Migration patterns of ey-positive cells in wild-type MARCM clones (A), in clones misexpressing ey^DN (B), in clones lacking DE-Cadherin (C), and in clones misexpressing ey^DN and lacking DE-Cadherin (D) (green) generated 72 hours AEL and analyzed in adult. Neuropil (red) was visualized with DN-Cadherin and photoreceptors with 24B10 (blue). Dotted circles in B highlight the accumulation of neurons in large clumps. (E) Number of cells per clone in ey-positive MARCM clones (mean±s.e.m.: 4.08±0.3), in clones mis-expressing ey^DN (19.43±1.56), and in clones lacking DE-Cadherin and mis-expressing ey^DN (6.04±0.31). (F) Number of large clumps per optic lobe in ey-positive MARCM clones (0), in clones mis-expressing ey^DN (5.44±0.28), and in clones lacking DE-Cadherin and mis-expressing ey^DN (3.11±0.35). ^**P<0.001.

Fig. 6.

Increased expression of DE-Cadherin in ey^DN MARCM clones. (A-D) DE-Cadherin (red) staining in ey-MARCM wild-type clones (A,B) and in ey-MARCM clones misexpressing ey^DN (C,D) generated 72 hours AEL and analyzed in L3.