Lineage mapping identifies molecular and architectural similarities between the larval and adult Drosophila central nervous system
- Janelia Research Campus, Howard Hughes Medical Institute, United States
Figures
Figure 1 with 1 supplement
Tracing individual NB lineages identifies postembryonic progeny of NBs.
(A) Schematic illustration of the strategies used in this study to trace NB lineages. Since expression of GAL4 lines is usually not maintained throughout the development, GAL4 expression was …
Figure 1—figure supplement 1
Schematic representations of postembryonic lineages in different segments of the nerve cord shown.
T2 and T3 segments virtually have the same lineages with the exception of lineage 11 (asterisk), which is dramatically reduced in cell number in the T3 segment. Among A2-A7 segments, NB6-2 survives …
Figure 2
Sample GAL4 lines that mark NBs and their progeny.
(A-K) Expression patterns of selected GAL4 lines in the nerve cords of late-stage embryos were visualized by driving mCD8-GFP (green). Only the T2-A2 segments are shown. (A-H) GAL4 lines uniquely …
Figure 3
NB2-4, NB2-5, NB3-5 and NB6-2 generate lineage 18, 17, 9 and 19, respectively.
Images on panels B, E, I, and M are from the larval CNS; the rest are from the embryonic CNS. (A) Projection of dorsally located NBs, marked by Dpn (green), in the thoracic segments of a stage-17 …
Figure 4 with 3 supplements
NB1-2, NB4-2, NB6-1 and NB7-2 generate lineages 1, 13, 12 and 11, respectively.
(A-D) Four examples showing identification of postembryonic lineages via reporter immortalization of NB-specific GAL4 lines. (i and ii) Nerve cords dissected from late stage embryos; Dpn (red) marks …
Figure 4—figure supplement 1
Postembryonic progeny of the remaining NBs.
See Figure 4 legends for more information. See Table 1 for efficiency and specificity of these lines on marking postembryonic lineages. (A-C) R70D06AD-R28H10DBD, R19H09 AD -R28H10DBD, and 5172J …
Figure 4—figure supplement 2
(A) NB1-1 generates lineage 1.
Top panels: A lineage clone generated via R28H10 showing embryonic progeny of NB1-1. Bottom panels: a lineage 16 MARCM clone generated by elav-GAL4. Hb9 (red) is expressed in both neuronal …
Figure 4—figure supplement 3
(A-B) NB5-5 disappears in the late stage embryos.
(A) In the stage 13 embryonic nerve cord, unpg-lacZ (green) marks NB5-5 (arrows), which is the most dorsal NB in the nerve cord and isolated from other NBs. (B) In the stage 15 embryonic nerve cord, …
Figure 5 with 1 supplement
Thoracic-specific NB2-3 and NB3-4 generate lineage 15 and lineage 25, respectively.
(A) R10C12 marks uniquely Msh+ NB2-3 (arrows) in each thoracic hemisegment but not in abdominal ganglia. Dpn, blue; Msh, red (B) A lineage clone containing NB2-3 (arrow) extends only efferent axons, …
Figure 5—figure supplement 1
(A) Stage 12 embryonic nerve cord; Engrailed (En; red) marks row 6 and 7 NBs as well as NB1-2.
R10C12 marked NB (arrows) is located immediately posterior and lateral to NB1-2. (B-C) Individual channels for T3 (A) and A1 (B) segments of the nerve cord shown in Figure 4D. These projections …
Figure 6 with 1 supplement
NB5-4 and NB5-7, a novel NB, generate almost identical lineages, lineage 22 and 20, respectively.
(A, F) R19B03AD-R45D04DBD marks two adjacent NB in the embryonic (A-E) and the larval CNS (F). Hemisegments on the right of the midline are shown. (A-B) GFP (green) shows complete GAL4 expression …
Figure 6—figure supplement 1
(A-C) Late stage embryos shown.
(A) Expression pattern of R719B03AD-R45D04DBD whole mount-embryo. Green, mcd8-GFP; red, nls-tdTomato. (B) A NB5-4 lineage clone, which was generated with R719B03AD-R45D04DBD in a thoracic segment, …
Figure 7 with 1 supplement
Hox genes restrict NB2-3 and NB3-4 lineages to the thoracic segments.
(A-G) 77B09-lexAp65 driven GFP marks NB2-3 and NB3-4 lineages in wild type embryos (A, B, D and E) and embryos mutant for antp (C), ubx (F), and double mutant for abd-A and ubx (G). All embryos are …
Figure 7—figure supplement 1
Apoptosis is not responsible for the lack of NB2-3, NB3-4 and NB 5–7 in the abdominal segments.
Nerve cords from H99 mutant embryos at stage 12 (A) and stage 16 (B). (A) Blocking apoptosis does not introduce new Msh+ NBs in the abdominal segments (compare to Figure 4D). No additional NBs are …
Figure 8
NB5-6 generates postembryonic progeny only in the S3 segments.
Expression of lbe-K-Gal4 in the embryonic nerve cord (A-B) and immortalization of its expression in NB5-6 progeny with (C) or without p35 in the larval CNS (B). (A) S3-A1 segments of stage 16 …
Figure 9
Embryonic progeny of NB2-3 are developmentally arrested during larval life, but differentiate during metamorphosis to be leg motor neurons.
(A, E) R77B09AD-R28H10DBD (A) and VT006878 (E) expression in T2 hemisegments from a late stage larva shown. EDU (red) labels all of the postembryonic neurons but not NB2-3 progeny (outlined with …
Figure 10 with 1 supplement
Late-born embryonic neurons and postembryonic neurons show significant similarities.
(A) A transverse view of the larval thoracic nerve cord. R24B02 driven GFP is in green; flip-out clones, which are induced in the embryo via R24B02, are in red. Within the NB6-1 lineage, mature …
Figure 10—figure supplement 1
(A-C) NB5-3 lineage clones, which were induced at different time points (A, earliest; C, latest).
All embryos at stage 17. Clones generated with R86D02. Late born neurons extend contralateral axons locally. (D) Birth order in NB5-3 lineage is shown in a color code. Red cells are born last. (E-F) …
Figure 11
Dicheaete and Dbx marks both embryonic ands postembryonic NB4-2 progeny.
Nerve cord images from the embryo (A-D) and larva (E). (A) R81F02 drives reporter expression only in thoracic segments. R81F02 marks D+ neurons (red) of NB4-2; abdominal segments do not have these D+…
Tables
Table 1
GAL4 lines used to mark NBs and for reporter immortalization.
| Line | NB expression | Immortalization* | Figure |
|---|---|---|---|
| R16A05AD-R28H10DBD | NB1-2 | lin1 (30/30) | Figure 2, Figure 4 |
| R70D06AD-R28H10DBD | NB2-1 | lin 2 (18/ 30), lin10 (7/ 30) | Figure 2, Figure 4—figure supplement 1 |
| R19H09AD-R28H10DBD | NB2-2 | lin10 (16/30), lin7 (4/30) | Figure 4—figure supplement 1 |
| R10C12 | NB2-3 | lin15 (30/30), lin25(7/30) | Figure 5 |
| R65G02 | NB2-4 | lin 18 (21/30), lin20-22 (9/30), lin5 (5/30) | Figure 3 |
| 5172J-Gal4 | NB3-1 | lin4 (12/18) | Figure 4—figure supplement 1 |
| R21E09AD-R16H11DBD | NB3-2, | lin7 (18/24), | Figure 4, Figure 4—figure supplement 1 |
| R21E09AD-R28H10DBD | NB1-2, | lin1 (15/24), | |
| ems-Gal4 | NB2-2, , | lin10(10/30), , | Figure 3, Figure 4—figure supplement 1 |
| R59E09 | NB3-5 | lin9 (30/30) | Figure 2, Figure 3 |
| R77B09AD-R28H10DBD | NB2-3, , | lin15 (30/30), ; | Figure 1, Figure 4 |
| VT0048571 | NB4-1, | lin14 (34/36), | Figure 4—figure supplement 1 |
| R81C12AD-R42F01DBD | NB4-2 | Lin13 (30/30) | Figure 2, Figure 4 |
| R19B03AD-R16H11DBD | NB4-3, , | lin21 (30/30), , , | Figure 2, Figure 4—figure supplement 1 |
| VT0041296 | NB4-4 | Lin24 (20/24); lin18 (5/18) | Figure 4—figure supplement 1 |
| R54B10 | NB5-3, NB5-6 | lin5 (17/30) | Figure 4—figure supplement 1 |
| R19B03AD-R45D04DBD | NB5-4, , | lin20 (30/30), , | Figure 2, Figure 6 |
| R24C10 | NB5-7 | lin20 (23/30), lin18(6/24) | Figure 6 |
| R81F01 | NB6-1 | lin12 (13/30), lin18 (5/30) | |
| R70D06AD-R42F01DBD | NB6-1 | lin12 (12/24), lin13(7/24) | Figure 2, Figure 4 |
| R76D11 | NB6-2 | lin19 (16/30) | Figure 2; Figure 3 |
| R51B04 | NB7-1, | lin3 (22/30), | Figure 4—figure supplement 1 |
| R35B12AD-R28H10DBD | NB7-2 | lin11 (12/30) | Figure 2, Figure 4 |
| R35B12 | NB7-1, , | lin3(10/30), , | |
| R19B03AD-R18F07DBD | NB7-4 | lin23 (26/30) | Figure 2, Figure 4—figure supplement 1 |
| R19B03** | NB2-5, | lin17 (20/24), | Figure 3 |
| R13G03 | MNB | lin0 (10/18) | Figure 4—figure supplement 1 |
| lbe-K-GAL4*** | NB5-6 | lin5-6 (20/20) | Figure 8 |
| R45D04 | NB5-2, NB5-3, NB5-4, NB5-7, NB6-2 | lin6 (30/30), lin5(30/30), lin20 (30/30), lin22(30/30), lin19 (18/30) | |
| eg-GAL4 | NB2-4, , , NB6-4, NB7-3 | lin18 (19/30), | Figure 5, Figure 4—figure supplement 1 |
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* The number of immortalized lineage per hemisegment shown in paranthesis. Corresponding NBs and lineages are color matched.
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** Only dorsal part VNC scored
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***NB5-6 generate postembryonic progeny only in S3 segments Lineages marked less than 15% of the time are not included. “lin” refers to postembryonic lineage.
Table 2
Expression profile of transcription factors in NBs and their corresponding embryonic and postembryonic progeny.
| NBs | lin | Embryonic progeny* | Postembryonic lin** | NB Marker*** |
|---|---|---|---|---|
| MNB | lin 0 | En, FoxD, Vg | En | En, unpg-Lacz |
| NB1-1 | lin16 | Hb9, Lim3, Isl, D, Eve, | Hb9, Lim3 | mirr-Lacz |
| NB1-2 | lin1 | Msh, Nmr1, Hb9, Nkx6 | Nmr1, Msh | mirr-Lacz |
| NB2-1 | lin2 | Toy | mirr-Lacz | |
| NB2-2 | lin10 | Hb9, Lim3, Nkx6, | Hb9, Lim3, Nkx6 | mirr-Lacz, Run |
| NB2-3 | lin15 | Lim3, Nkx6, Isl, | Isl, Lim3, Nkx6 | mirr-Lacz, Msh, Run |
| NB2-4 | lin18 | Unc-4, Eg, Toy, Msh | Unc-4 | mirr-Lacz, Msh |
| NB2-5 | lin17 | Unc-4, Isl | Unc-4, Isl | mirr-Lacz |
| NB3-1 | lin4 | Hb9, Msh | Hb9 | Nkx6, Run |
| NB3-2 | lin7 | Hb9, Toy, Barh, Unc-4 | Unc-4 | Ey, Dbx |
| NB3-3 | lin8 | Toy, Lim3, Ems, Acj6, Eg, Eve | Lim3, Ems, Acj6, Toy, | Ems,Run |
| NB3-4 | lin25 | Toy, Ey, Msh, Eg | Toy, Nkx6 | Msh, Run, Ey, Eg-Gal4 |
| NB3-5 | lin9 | Ems, Msh, Islet, Unc-4 | Ems, Msh, Islet | Ems |
| NB4-1 | lin14 | Msh, Unc-4 | Msh | unpg-Lacz |
| NB4-2 | lin13 | Dbx, D, Vg,Ey, Eve | Dbx, D, Vg | Ey |
| NB4-3 | lin21 | Msh, Ey | Msh, Ey | Ey, Msh |
| NB4-4 | lin24 | Ems, Toy | Ems, Toy | Ems, Ey, |
| NB5-1 | - | gsb-Lacz | ||
| NB5-2 | lin6 | Toy, En, Vg, Hb9 | Toy, En, Vg | gsb-Lacz, Run |
| NB5-3 | lin5 | Vg, Toy, Ey, En | Vg, Toy, | gsb-Lacz, Ey, Run |
| NB5-4 | lin22 | BarH | BarH | gsb-Lacz, Msh |
| NB5-5 | - | unpg-Lacz | ||
| NB5-6 | - | EyA, Toy | EyA (S3 segments) | gsb-Lacz,lbe-Gal4 |
| NB5-7 | lin20 | - | gsb-Lacz, Msh | |
| NB6-1 | lin12 | Unc-4, Nmr1, Dbx | Dbx, Unc-4, Nmr1 | gsb-Lacz, En, Dbx |
| NB6-2 | lin19 | Unc-4, Dbx | Dbx, Unc-4 | gsb-Lacz, En, D |
| NB6-4 | - | Eg, Toy, Msh | gsb-Lacz, Eg-lacz | |
| NB7-1 | lin3 | Unc-4, Dbx, Eve | Dbx, Nkx6, | gsb-Lacz, En |
| NB7-2 | lin11 | Unc-4, Nkx6 | Unc-4, Nkx6, | En, unpg-Lacz, Dbx |
| NB7-3 | - | Hb9, Isl, Eg, Ey | En, eg-GAL4 | |
| NB7-4 | lin23 | Unc-4, Acj6 | Unc-4, Acj6 | En, Msh, D |
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* Transcription factors that are also expressed in the postembryonic neurons are highlighted in bold.
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** We failed to detect transcription factors in red in the corresponding embryonic progeny.
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*** Expression of these markers is maintained from embryonic to postembryonic stages. “lin” refers to postembryonic lineage.
Table 3
Lineages that are assigned to different NBs by this study and Birkholz et al. (2015).
| Lin | Findings on lineage tracing supporting assignments of this study | Findings on molecular markers supporting assignments of this study |
|---|---|---|
| lin7 NB3-2* NB4-2** | Immortalization of R21E09AD-R28H10DBD, which marks lin7, labels NB3-2 in the embryo, but not NB4-2. | NB3-2 and NB of lin7 are Dbx+versus NB4-2 is Dbx-. Embryonic progeny of NB3-2 and lin7 neurons are Unc4+ versus NB4-2 progeny are Unc4-. |
| lin8 NB3-3* NB2-4** | Immortalization of ems-GAL4, which marks lin8, labels NB3-3 in the embryo, but not NB2-4 (Figure 4—figure supplement 1). | NB3-3 and NB of lin8 are Ems+versus NB2-4 is Ems-. Embryonic progeny of NB3-3 and lin8 neurons are Acj6+ versus NB2-4 progeny are Acj6-. |
| lin11 NB7-2* NB6-4** | Immortalization of R35B12AD-R28H10DBD, which specifically marks lin11, labels only NB7-2 in the embryo, but not NB6-4 (Figure 4). NB6-4 dies (TUNEL+) in late stage embryos (Figure 4—figure supplement 3). | NB7-2 and NB of lin11 are marked by unpg-LacZ versus NB6-4 is not. NB6-4 is Msh+ versus NB of lin11 is Msh-. Embryonic progeny of NB7-2 and lin11 neurons are Unc4+ versus NB6-4 progeny are Unc4-. |
| lin13 NB4-2* NB3-3** | Immortalization of R81C12AD-R42F01DBD, which specifically marks lin13, labels only NB4-2 in the embryo, but not NB3-3 (Figure 4). | NB4-2 and NB of lin13 are Ey+ versus NB3-3 is Ey-. Embryonic progeny of NB4-2 and lin13 neurons are Dbx+/D+/Vg+ versus NB3-3 progeny are Dbx-/D-/Vg- (Figure 11). |
| lin15 NB2-3* NB3-2** | Immortalization of R10C12, which specifically marks lin15, labels only NB2-3 in the embryo, but not NB3-2 (Figure 5). | Lineage 15 NB and NB2-3 are Msh+ versus NB3-2 is Msh-. |
| lin18 NB2-4* NB3-4** | Immortalization of R65G02, which specifically marks lin18, labels only NB2-3 in the embryo, but not NB3-4 (Figure 5). Dorsal location of NB2-4 correlates with lineage 18. | Embryonic progeny of NB2-4 and lin18 neurons are Unc4+ versus NB3-4 progeny are Unc4-. |
| lin20 NB5-7* NB5-4** | Immortalization of R24C10, which marks lin20, labels NB5-7 in the embryo, but not NB5-4 (Figure 6). | NB5-7 shares similar molecular markers with NB5-4. |
| lin25 NB3-4* -** | Lineage tracing via eg-GAL4, R77B09AD-R28H10DBD, and R19B03AD-R16H11DBD lines, all of which mark lin25, label NB3-4 in the embryo (Figure 5). | Lineage 25 is first described here and assigned to NB3-4, which has not been previously characterized in detail. |
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*NB assignment by this study
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** NB assignment by Birkholz et al. (2015) “lin” refers to postembryonic lineage
