spacer gif spacer gif spacer gif spacer gif spacer gif
QUICK SEARCH:   [advanced]


spacer gif
     Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents    

doi: 10.1242/10.1242/dev.00356

This Article
Right arrow Summary Freely available
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Supplemental Data
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Ang, L.-H.
Right arrow Articles by Hing, H.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Ang, L.-H.
Right arrow Articles by Hing, H.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati   Add to Twitter  
What's this?

Dock and Pak regulate olfactory axon pathfinding in Drosophila

Lay-Hong Ang, Jenny Kim, Vitaly Stepensky and Huey Hing*

Department of Cell and Structural Biology, University of Illinois at Urbana-Champaign, 601 South Goodwin Avenue, C626, Urbana, IL 61801, USA



View larger version (145K):

[in a new window]
  		Fig. 1. DM2, DM3 and VA1lm glomeruli are mis-shapen and ectopically localized in dock and Pak mutants. Confocal micrographs of adult ALs stained with the nc82 (red) and anti-GFP (green) antibodies, revealing the neuropil structure. (A) Wild-type ALs showing distinct anatomical subdivisions: the glomeruli. (B) A pair of dockP1/dockP1 ALs and (C) a pair of Pak6/Pak11 ALs showing an amorphous neuropil structure. (D-F) DM2, (G-I) DM3 and (J-L) VA1lm glomeruli were visualized by expressing UAS-nsyb::GFP with Or22a-Gal4, Or47a-Gal4 and Or47b-Gal4, respectively. In the wild type (D,G,J), DM2, DM3 and VA1lm have characteristic shapes and are located in invariant positions in the AL. In dockP1/dockP1 (E,H,K), and Pak6/Pak11 (F,I,L) mutants, DM2 and DM3 glomerular structures are smaller and scattered randomly in the AL. While VA1lm is located approximately in the right position, it is severely misshapen and frequently engulfs the adjacent VA1d glomerulus. Scale bar: 20 µm.

 


View larger version (112K):

[in a new window]
  		Fig. 2. dock and Pak function autonomously in ONs for AL development. (A) ALs from animals with mosaic wild-type antennae: ey-FLP/+; Or47b-Gal4/+; FRT82/FRT82 M(3) arm-lacZ UNG. UNG stands for UAS-nysb::GFP. (B) ALs from animal with mosaic dockP1 antennae: ey-FLP/+; dockP1 FRT40/cycE FRT40; Or47a-Gal4 UNG/+. (C) ALs from animal with mosaic Pak16 antennae: ey-FLP/+; Or47b-Gal4/+; FRT82 Pak16 UNG/FRT82 M(3) arm-lacZ. (D-F) Animals expressing UAS-mCD8::GFP under the control of SG18.1-Gal4. (D) Confocal stereo pair of a 30 hAPF antenna double-stained with anti-mCD8 (green) and 22C10 mAb (red). Cells with a bipolar morphology, characteristic of ONs, express both GFP and Futsch. (E) Optical section of the adult ALs, showing GFP expression in the outer nerve layer (arrowhead) and in many glomeruli. Little GFP staining is observed outside of the AL. (F) Optical section at an equivalent plane as in E of an animal from which both the third segments of the antennae and the maxillary palps were surgically removed 6 days earlier. GFP staining in the outer nerve layer (arrowhead) and glomeruli is lost. (G-J) Expression of UAS-dock and UAS-Pak under the control of SG18.1-Gal4 rescues the mutant AL phenotypes. 3D reconstructions of ALs from dock (G) and Pak (I) mutants show that their ALs are mis-shapen and aglomerular. (H) Expression of the dock cDNA in dockP1 homozygotes using SG18.1-Gal4 rescues the development of most of the glomeruli in the mutant. (J) Similarly, expression of Pak cDNA using SG18.1-Gal4 in Pak6/Pak11 mutants strongly rescues the development of most of the glomeruli. (K) Quantification of the rescue of the dock and Pak AL phenotype by wild-type dock and Pak cDNAs. The frequencies with which four indicator glomeruli (VA1d, DA4, DM6 and VA6) were observed in the different genotypes are presented. Scale bars: in A, 25 µm for A-C,E-J; in D, 25 µm for D.

 


View larger version (66K):

[in a new window]
  		Fig. 3. Olfactory axon trajectories are severely disrupted in dock and Pak mutants. ALs of animals expressing GFP under the control of various Gal4 drivers were either observed with fluorescence microscopy (I,K,L) or stained with anti-GFP (green) and nc82 mAb (red) and examined by confocal microscopy (A,C-E,G,H). (B,F,J) Interpretive drawings of confocal images (A,B1) In the wild type, upon entering the AL, SG18.1-Gal4 expressing axons radiate over its surface in characteristic tracks in search of their target glomeruli. These pathways are bilaterally symmetric and invariant from one lobe to another. (B2 ,C) In the dockP1/dockP1 and (D) Pak4/Pak6 mutants, however, SG18.1-Gal4 axons take circuitous paths in the nerve layer creating a homogenous mat. Some axons project to dorsal brain regions (arrowheads) in dockP1 mutants. (E,F1) In the wild type, Or47a axons take direct paths, with little sidetracking, from the entry point to their target, DM3. (F2,G) However, in dockP1/dockP1 and (H) Pak4/Pak6 mutants, Or47a axons make chaotic and meandering projections, terminating in numerous ectopic glomeruli. (I,J1) In the wild type, Or47b axons terminate immediately on VA1lm upon entering the AL. A distinct fascicle, containing contralateral axons, connects the glomerulus with the commissure. (J2,K) In dockP1/dockP1 and (L) Pak6/Pak11 mutants, the VA1lm glomerulus is severely mis-shapen and contralaterally projecting axons enter and exit the glomerulus over its entire surface. Asterisks in A,E,I indicate the antennal nerve entry points at the anterior lateral region of the AL. Scale bar: 25 µm.

 


View larger version (99K):

[in a new window]
  		Fig. 6. dock and Pak do not function in antennal neuron differentiation. (A-C) Confocal micrographs of 30 hAPF pupal antennae double-stained with anti-Elav (green) and 22C10 mAb (red). ONs express Elav and project their axons out of the antenna normally in the dock (B) and Pak (C) mutants, compared with the wild type (A). (D-F) Adult antennae mounted in Faure's mountant; coeloconic sensilla are highlighted by blue dots. The morphology, number and distribution of sensilla in dock (E) and Pak (F) mutants are similar to those in wild type (D). See Table 2 for a summary of the results. (G-I) Antennae from Or47a-Gal4/UAS-lacZnuclear stained with X-gal showing the distribution and number of Or47a ONs. The number and distribution of Or47a neurons, in a diagonal lateral-to-medial stripe, in dock (H) and Pak (I) mutants are similar to those in wild type (G). Scale bar: 30 µm.

 


View larger version (161K):

[in a new window]
  		Fig. 4. Or47a and Or47b axons show strong pathfinding defects and terminate in ectopic glomeruli in dock mutants. Antennal lobes, in which single Or47a or Or47b axons were labeled with mCD8::GFP using the MARCM technique, were stained with anti-CD8 (green) and nc82 mAb (red). See Materials and Methods for the genotypes. (A,B) In wild type, each Or47a axon can be seen to synapse on the ipsilateral DM3 and then send a collateral to the contralateral DM3. (C,D) In the dock mutant, Or47a axons branch normally. However, both axon branches are severely misrouted and arborize away from their normal target area. (E,F) Wild-type Or47b axons terminate on VA1lm and extend a collateral across the commissure to the corresponding glomerulus on the contralateral lobe. (G,H) In the dock mutant, some Or47b axons can be seen extending to dorsal brain regions (arrowhead), while others stop short of VA1lm. Broken outlines in all panels indicate the approximate positions of the glomerular targets. Scale bar: 25 µm.

 


View larger version (144K):

[in a new window]
  		Fig. 5. Interaction between Dock and Pak is necessary for antennal lobe development. ALs of adult dockP1/dockP1 mutants, expressing wild-type and various mutant UAS-dock cDNA constructs under the control of SG18.1-Gal4, were stained with nc82 mAb to assess AL development. (A) Wild-type dock cDNA: UAS-dockWT; dockP1/dockP1. (B) No cDNA: dockP1/dockP1. (C) Mutation in the first SH3 domain: UAS-dockW48K; dockP1/dockP1. (D) Mutation in the second SH3 domain: UAS-dockW151K; dockP1/dockP1. (E) Mutation in the third SH3 domain: UAS-dockW225K; dockP1/dockP1. (F) Mutation in the SH2 domain: UAS-dockR336Q; dockP1/dockP1. Scale bar: 25 µm.

 

Add to CiteULike CiteULike   Add to Complore Complore   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati   Add to Twitter Twitter    What's this?



© The Company of Biologists Ltd 2003