Par3 cooperates with Sanpodo for the assembly of Notch clusters following asymmetric division of Drosophila sensory organ precursor cells
- Univ Rennes, CNRS, IGDR (Institut de Génétique et Développement de Rennes) - UMR 6290, F- 35000, France
- Equipe Labellisée Ligue Nationale contre le cancer, United Kingdom
Figures
Figure 1 with 1 supplement
Distribution of polarity markers and Notch during sensory organ precursor (SOP) cell division.
(A–D’’) Time-lapse imaging of Crumbs (Crb)-GFP (A, A’, n = 22), atypical protein kinase C (aPKC)-GFP (B, B’, n = 10), Bazooka (Baz)-GFP (C, C’, n = 28), and NiGFP (D, D’, n = 25) during SOP …
Figure 1—figure supplement 1
Distribution of polarity markers and Notch during sensory organ precursor (SOP) and epidermal cell divisions.
(A–D) Time-lapse imaging of neuroglian (Nrg)-YFP (A, n = 11 SOPs; A’, n = 10 epidermal cells); discs-large (Dlg)-GFP (B, n = 9 SOPs; B’, n = 11 epidermal cells); Crumbs (Crb)-GFP (C, n = 11 …
Figure 2
Dynamics of colocalisation of NiGFP and Bazooka (Baz)-mCherry at pIIa-pIIb interface.
(A–A’) Schematic representation and time-lapse imaging of a dividing sensory organ precursor (SOP) expressing NiGFP (green) adjacent to epidermal cells expressing untagged version of Notch (n = 3). …
Figure 3
Notch loss-of-function does not impair the formation of Bazooka (Baz) clusters along the pIIb-pIIb-like interface.
(A–C) SO lineage analysis using the SO marker Cut (green), the socket marker Su(H) (magenta) and the neuronal marker Elav (blue) in control lineage (A, n = 3 nota), upon silencing of Notch (B, n = 2 …
Figure 4 with 2 supplements
Bazooka (Baz) and Delta are required for localisation of Notch in lateral clusters and for Notch activation.
(A–D’’) Time-lapse imaging of NiGFP (green) together with H2B-RFP expressed under the neur minimal driver (magenta) during sensory organ precursor (SOP) cytokinesis in control (A), upon silencing of …
Figure 4—figure supplement 1
NiGFP localisation in bazEH747 mutant clones.
(A–A’) Bazooka (Baz) immunostaining using an anti-Nterm (green, A) or an anti-PDZ antibody (green, A’). bazEH747 mutant clones are identified by the loss of the nuclear marker nls RFP (magenta). …
Figure 4—figure supplement 2
Requirements for Delta and Bazooka (Baz) for Notch activation.
(A–D) SO lineage analysis using the SO marker Cut (blue), the socket marker Su(H) (red), and the neuronal marker Elav (green) in control (A, n = 4 nota), upon silencing of Baz (B, n = 4 nota), Delta …
Figure 5
Neuralized (Neur) localises in and regulates the number of NiGFP/Bazooka (Baz)-positive clusters.
(A) Time-lapse imaging of Neur-GFP (green) together with Baz-Scarlet (red) during sensory organ precursor (SOP) cytokinesis in control (n = 8). Yellow dashed rectangles highlight the high …
Figure 6
Numb regulates the number of NiGFP/Bazooka (Baz)-positive clusters.
(A) Time-lapse imaging of Numb-GFP (green) together with Baz-Scarlet (red) during sensory organ precursor (SOP) cytokinesis in control (n = 5). Yellow-dashed rectangles highlight the high …
Figure 7 with 1 supplement
Sanpodo is required for the assembly of NiGFP/Bazooka (Baz) clusters.
(A) Localisation of Sanpodo (Spdo)-GFP (green) together with Baz-Scarlet (magenta) at t21 during sensory organ precursor (SOP) cytokinesis in control (n = 10). Yellow-dashed rectangles highlight the …
Figure 7—figure supplement 1
Sanpodo is required for the assembly of NiGFP/Bazooka (Baz) clusters.
(A-A’) Localisation of NiGFP (green) together with H2B-IRF670 and growth-associated protein 43 (GAP43) expressed under the neur minimal driver (magenta) at t21 during sensory organ precursor (SOP) …
Videos
Video 1
3D viewing of the time-lapse of Baz-GFP (green) and GAP43-IR (magenta) from t0 to t18, illustrating the position of the Baz-positive lateral clusters along the pIIa-pIIb interface at t18.
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Gene (Drosophila melanogaster) | w1118 | Bloomington Drosophila Stock Center | BDSC: 3,605FLYB: FBal0018186;RRID:BDSC_3605 | |
| Gene (Drosophila melanogaster) | Nrg::YFPP{PTT-GA}NrgG00305 | Kyoto Stock Center Morin et al., 2001 | FLYB: FBgn0264975;RRID:KSC_110658 | |
| Gene (Drosophila melanogaster) | neur-H2B::IR | This study | Two lines generated (IId and IIId) | |
| Gene (Drosophila melanogaster) | Crb::GFP (A) | Huang et al., 2009 | FLYB: FBgn0259685 | Gift from Dr Y Hong |
| Gene (Drosophila melanogaster) | aPKC::GFP | Besson et al., 2015 | FLYB: FBgn0261854 | Kind gift from Dr F Schweisguth |
| Gene (Drosophila melanogaster) | Baz::GFPP{PTT-GC}bazCC01941 | Bloomington Drosophila Stock Center Buszczak et al., 2007 | FLYB:FBgn0000163;RRID:BDSC_51572 | |
| Gene (Drosophila melanogaster) | Baz::mScarlet | kind gift from Dr J Januschke | FLYB:FBgn0000163 | |
| Gene (Drosophila melanogaster) | neur-GAP43::IR | This study | Two lines generated (IId and IIId) | |
| Gene (Drosophila melanogaster) | Dlg::GFP | Bloomington Drosophila Stock Center Buszczak et al., 2007 | FLYB: FBgn0001624;RRID: BDSC_ 50859 | |
| Gene (Drosophila melanogaster) | Ubi-p63E-Baz-mCherry (II) and (III) | Bosveld et al., 2012 | Transgenic lines;FLYB:FBgn0000163 | Kind gift from Dr Y Bellaiche |
| Gene (Drosophila melanogaster) | NiGFP | Bellec et al., 2018 | FLYB: FBgn0004647 | CRISPR line |
| Gene (Drosophila melanogaster) | neur-H2B-RFP | Gomes et al., 2009 | Kind gift from Dr F Schweisguth | |
| Gene (Drosophila melanogaster) | bazEH747, FRT19A/Y | Kind gift of A Wodarz | FLYB:FBgn0000163 | |
| Gene (Drosophila melanogaster) | Ubi-mRFP.nls, w*, hsFLP, FRT19A | Bloomington Drosophila Stock Center | RRID:BDSC_31418 | |
| Gene (Drosophila melanogaster) | UAS-Baz dsRNA (III) | Bloomington Drosophila Stock Center | RRID:BDSC_35002 | |
| Gene (Drosophila melanogaster) | UAS-Notch dsRNA (III) | Bloomington Drosophila Stock Center | FLYB: FBgn0004647; RRID:BDSC_28981 | |
| Gene (Drosophila melanogaster) | UAS-Dl dsRNA (III) | Bloomington Drosophila Stock Center | FLYB: FBgn0000463; RRID:BDSC_28032 | |
| Gene (Drosophila melanogaster) | UAS-Nslmb-vhhGFP4 (II) | Bloomington Drosophila Stock Center | RRID:BDSC_38422 | |
| Gene (Drosophila melanogaster) | UAS-Nslmb-vhhGFP4 (III) | Bloomington Drosophila Stock Center | RRID:BDSC_38421 | |
| Gene (Drosophila melanogaster) | pnr-GAL4 | Bloomington Drosophila Stock Center | FLYB: FBgn0003117; RRID:BDSC_3039 | Calleja et al., 1996 |
| Gene (Drosophila melanogaster) | UAS-Nb dsRNA (III) | Bloomington Drosophila Stock Center | FLYB: FBgn0002973; RRID:BDSC_35045 | |
| Gene (Drosophila melanogaster) | FRT82B, neur1F65 | Yeh et al., 2000 | FLYB: FBgn0002932 | |
| Gene (Drosophila melanogaster) | FRT82B, nls-RFP | Bloomington Drosophila Stock Center | RRID:BDSC_30555 | |
| Gene (Drosophila melanogaster) | hsFLP | Bloomington Drosophila Stock Center | RRID:BDSC_6938 | |
| Gene (Drosophila melanogaster) | FRT82B, spdoG104 | O’Connor-Giles and Skeath, 2003 | FLYB: FBgn0260440 | |
| Gene (Drosophila melanogaster) | Spdoi-GFP (II) | Couturier et al., 2013 | FLYB: FBgn0260440 | BAC Rescue, Kind gift from Dr F Schweisguth |
| Gene (Drosophila melanogaster) | UAS-Spdo dsRNA (II) | Vienna Drosophila Resource Center | FLYB: FBgn0260440; RRID:VDRC_104092 | |
| Gene (Drosophila melanogaster) | UAS-Neur dsRNA | Bloomington Drosophila Stock Center | FLYB: FBgn0002932; RRID:BDSC_26023 | |
| Gene (Drosophila melanogaster) | y, w, PB[y + attP-3B Neur::GFP 22A3] | Perez-Mockus et al., 2017 | FLYB: FBgn0002932 | BAC Rescue, Kind gift from Dr F Schweisguth |
| Antibody | Anti-Elav(rat monoclonal) | Developmental Studies Hybridoma Bank | RRID:Rat-Elav-7E8A10 | IF (1:200) |
| Antibody | Anti-cut(mouse monoclonal) | Developmental Studies Hybridoma Bank | RRID:2B10 | IF (1:500) |
| Antibody | Anti-Su(H)(goat polyclonal) | Santa Cruz | Cat# sc15813 | IF (1:500) |
| Antibody | Anti-Baz N-term(rabbit polyclonal) | Wodarz et al., 1999 | IF (1:1000),Kind gift from Dr A Wodarz | |
| Antibody | anti-Baz-PDZ (guinea pig) | Shahab et al., 2015 | IF (1:1000),Kind gift from Dr A Wodarz | |
| Antibody | Anti-GFP(goat polyclonal) | AbCam | Cat# Ab5450 | IF (1:500) |
| Antibody | Cy2-, Cy3- and Cy5-coupled secondary antibodies(donkey anti-goat) | Jackson Laboratories | 705-225-147, 705-165-147, and 705-175-147, respectively | IF (1:400) |
| Antibody | Cy2-, Cy3- and Cy5-coupled secondary antibodies(goat anti-rabbit) | Jackson Laboratories | 111-225-144, 111-165-144, and 111-175-144, respectively | IF (1:400) |
| Antibody | Cy2-, Cy3- and Cy5-coupled secondary antibodies(donkey anti-mouse min cross-react with rat) | Jackson Laboratories | 715-225-151, 715-165-151, and 715-175-151, respectively | IF (1:400) |
| Antibody | Cy2-, Cy3- and Cy5-coupled secondary antibodies(donkey anti-rat min cross-react with mouse) | Jackson Laboratories | 712-225-153, 712-165-153, and 712-175-153, respectively | IF (1:400) |
| Software, algorithm | ImageJ/FIJI | Open source Java image processing program | https://imagej.nih.gov/ij/ | |
| Software, algorithm | Countdots macro for FIJI | This study | ||
| Software, algorithm | Excel | Microsoft Office 2013 | RRID:Microsoft Excel 2013 | |
| Software, algorithm | Illustrator | Adobe Systems | RRID: Adobe Illustrator CS3 | |
| Software, algorithm | Prism | GraphPad | RRID: GrpahpadPrism |
