1. Neuroscience
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A multicellular rosette-mediated collective dendrite extension

  1. Li Fan
  2. Ismar Kovacevic
  3. Maxwell G Heiman
  4. Zhirong Bao  Is a corresponding author
  1. Sloan Kettering Institute, United States
  2. Boston Children’s Hospital, United States
  3. Harvard Medical School, United States
Research Article
Cite this article as: eLife 2019;8:e38065 doi: 10.7554/eLife.38065
4 figures, 1 video, 1 table and 1 additional file

Figures

Figure 1 with 1 supplement
Multicellular rosette precedes collective dendrite extension.

(A, B) Amphid neurons form a multicellular rosette. (A) Image of embryo prior to amphid dendrite extension stained with phalloidin. A multicellular rosette is shaded in pink. In this and all subsequent figures, embryos are oriented with anterior to the left. (B) cnd-1p::PH::GFP labels amphid neurons in the rosette. Lineage-derived cell identities are shown. (C, D) PAR-6 and DYF-7 are localized to the rosette vertex. cnd-1p::PH::mCherry labels neurons. (E) Amphid dendrite tip migrates anteriorly. PAR-6 accumulates in the dendrite tip. cnd-1p::PH::mCherry labels neurons. Vertical dashed lines show initial position of amphid dendrite tip and posterior extent of amphid cell bodies. Arrows indicate dendrite tip. Closed arrowhead indicates amphid axon commissure. Open arrowhead indicates sensory depression. (F) Measured displacement of amphid dendrite tips from five embryos during dendrite extension. Scale bars in A-E, 10 µm.

https://doi.org/10.7554/eLife.38065.002
Figure 1—source data 1

Displacement of dendrite tip from five embryos.

https://doi.org/10.7554/eLife.38065.004
Figure 1—figure supplement 1
Retrograde extension in wild-type embryos.

After the initial anterior extension is complete and the dendrite tips reach the sensory depression (top panel), cell bodies start to move posteriorly as the embryo elongates, further extending the dendrites through retrograde extension. Arrows indicate amphid dendrite tips. Closed arrowheads indicate amphid commissure. Open arrowheads indicate the sensory depression. Dashed line marks the posterior end of the amphid neurons at the time when anterior dendrite extension is complete. Scale bar, 10 µm.

https://doi.org/10.7554/eLife.38065.003
The amphid rosette vertex is attached to the epidermis.

(A) PAR-6-labeled amphid tip (arrows) is localized at the hyp5 leading edge. The apical junction of epidermal cells was labeled by DLG-1::GFP with xnIs17 transgene. The inset is shown magnified on the right. (B) A kymograph generated along the dashed line in the inset in A, showing the correlated anterior movement of the PAR-6 signal and the leading edge of the epidermal cell. (C) Example of an elt-1(RNAi) embryo with weak RNAi effect which is similar to the wild type (left) and an elt-1(RNAi) embryo with strong RNAi effect and minimal lin-26::GFP expression (right). In the latter group dendrites failed to extend (arrow), even though the amphid rosette (circle in the upper panel) and the amphid axon commissure both formed normally (arrowheads). (D) A representative elt-1(RNAi) embryo with moderate lin-26::GFP expression shows a partial anterior migration of the epidermis (upper panel) followed by posterior retraction. Dashed lines mark the leading edge of the epidermis before and after the retraction. The amphid dendrite tips follow the leading edge of the epidermis. Arrows mark the position of the dendrite tip. Scale bars in A-D, 10 µm.

https://doi.org/10.7554/eLife.38065.008
Figure 3 with 1 supplement
Multiple adhesion molecules function redundantly in amphid dendrite anterior extension.

(A–D) Localization of DYF-7, HMR-1, SAX-7 and DLG-1 (dlg-1(cp301)) in amphid neurons and epidermal cells. Middle panels, superficial focal plane showing localization in epidermal cells. Lower panels, deeper focal plane showing localization in amphid neurons. Arrows indicate amphid tips and arrowheads indicate epidermal cells. (E) Dynamics of dendrite anterior extension in wt, dyf-7(m537) and dyf-7;sax-7;hmr-1(RNAi) triple loss of function. (F) Frequency of anterior extension defects in single, double and triple loss of function embryos. Number of amphids scored is indicated. P-values were calculated with Fisher’s exact test (two-tailed). Threshold for significance was adjusted by Bonferroni correction to 0.01 for five comparisons. Scale bars in A-E, 10 µm.

https://doi.org/10.7554/eLife.38065.009
Figure 3—source data 1

P values in multiple comparasions.

https://doi.org/10.7554/eLife.38065.005
Figure 3—figure supplement 1
dyf-7 phenotype and genetic redundancy.

(A) Retrograde extension defects in dyf-7(m537) embryos.The dendrite tips detach from the sensory depression during embryo elongation, after successful anterior extension. See Figure 1—figure supplement 1 for the wild-type control. Arrows indicate amphid dendrite tips. Closed arrowheads indicate amphid commissure. Open arrowheads indicate the sensory depression. Dashed line marks the posterior end of the amphid neurons at the time when anterior dendrite extension is complete. Scale bar, 10 µm. (B) Frequency of anterior extension defects in single, double and triple loss of function embryos with dyf-7(ns119), sax-7(eq1), hmr-1(RNAi) or dlg-1(RNAi). Number of amphids scored is indicated. P-values were calculated with Fisher’s exact test (two-tailed). Threshold for significance was adjusted by Bonferroni correction to 0.01 for five comparisons.

https://doi.org/10.7554/eLife.38065.010
Figure 4 with 1 supplement
PAR-6 regulates localization of DYF-7.

(A, B) DYF-7 shows more dispersed localization in par-6(M/Z). (C) Quantification of the size of DYF-7::GFP in WT and par-6(M/Z) embryos before the dendrite tips migrate anteriorly. Data was shown as mean ±SD. P value was calculated with student t-test. (D) Schematic model of molecular localization between the amphid dendrite tips and epidermal cell. Scale bars in A-B, 10 µm.

https://doi.org/10.7554/eLife.38065.011
Figure 4—source data 1

Size of DYF-7::GFP in wt and par-6(M/Z) embryos.

https://doi.org/10.7554/eLife.38065.006
Figure 4—figure supplement 1
PAR-6 is required for amphid dendrite extension.

(A) The wild type. (B) Partial extension of amphid dendrites in par-6(M/Z) embryos that arrested at the 1.5-fold stage, indicating abnormal epidermal morphogenesis. (C) Partial extension of amphid dendrites in a par-6(M/Z) embryo that developed beyond the 1.5-fold stage, indicating more or less normal epidermal morphogenesis during dendrite extension. Arrows indicate amphid dendrite tips. Closed arrowheads indicate amphid commissure. Open arrowheads indicate the sensory depression. Scale bar, 10 µm.

https://doi.org/10.7554/eLife.38065.012

Videos

Video 1
Amphid dendrite anterior extension.

Time-lapse imaging of a wild-type embryo expressing cnd-1p::PH::mCherry to label sensory neurons and PAR-6::GFP to label dendrite tips. Dashed lines mark the initial position of the amphid neuron cell bodies at 0 min. The dendrite tip is tracked with an arrow. Scale bar, 10 µm.

https://doi.org/10.7554/eLife.38065.007

Tables

Key resources table
Reagent type
(species) or
resource
DesignationSource or
reference
IdentifiersAdditional
information
Genetic reagent
(E. coli)
OP50Caenorhabditis
Genetics Center
OP50
Genetic reagent
(C. elegans)
dex-1(ns42) IIIPMID: 19344940
Genetic reagent
(C. elegans)
dyf-7 (ns119) XPMID: 19344940
Genetic reagent
(C. elegans)
dyf-7 (m537) XPMID: 19344940
Genetic reagent
(C. elegans)
sax-7(ky146) IVCaenorhabditis
Genetics Center
CX2993
Genetic reagent
(C. elegans)
sax-7(eq1) IVCaenorhabditis
Genetics Center
LH81
Genetic reagent
(C. elegans)
par-6(zu170) ICaenorhabditis
Genetics Center
FT36
Genetic reagent
(C. elegans)
par-6(tm1425)/hIn1
[unc-54(h1040)] I
Caenorhabditis
Genetics Center
JJ1743
Genetic reagent
(C. elegans)
unc-101(m1) ICaenorhabditis
Genetics Center
DR1
Genetic reagent
(C. elegans)
zbIs3[cnd-1p::PH::GFP]PMID: 28441532
Genetic reagent
(C. elegans)
ujIs113[pie-1p::mCherry::H2B;
nhr-2p::mCherry::HIS-24
-let-858UTR; unc-119(+)] II
Caenorhabditis
Genetics Center
JIM113
Genetic reagent
(C. elegans)
itIs1024[par-6::PAR-6::GFP]Dr. Kenneth
Kemphues
KK1024
Genetic reagent
(C. elegans)
zyIs36[cnd-1p::PH::mCherry;
myo-2p::mCherry]X
PMID: 28441532
Genetic reagent
(C. elegans)
mcIs40 [lin-26p::ABDvab-
10::mCherry; myo-2p::GFP]
Caenorhabditis
Genetics Center
ML916
Genetic reagent
(C. elegans)
xnIs17[dlg-1::GFP; rol-6(su1006)]Caenorhabditis
Genetics Center
FT63
Genetic reagent
(C. elegans)
axIs1928[mCherry::PAR-6]Caenorhabditis
Genetics Center
JH2648
Genetic reagent
(C. elegans)
dlg-1(cp301[dlg-1::
mNG-C1^3xFlag])X
Caenorhabditis
Genetics Center
LP598
Genetic reagent
(C. elegans)
ntIs1[gcy-5::GFP]Caenorhabditis
Genetics Center
OH3192
Genetic reagent
(C. elegans)
oyIs44[odr-1::RFP + lin-15(+)]VCaenorhabditis
Genetics Center
PY2417
Genetic reagent
(C. elegans)
zuIs43[pie-1::GFP::PAR-6::ZF1;
unc-119(+)]
Caenorhabditis
Genetics Center
JJ1743
Genetic reagent
(C. elegans)
ddIs290[sax-7::TY1::EGFP::
3xFLAG(92C12); unc-119(+)]
Caenorhabditis
Genetics Center
TH502
Genetic reagent
(C. elegans)
xnIs96 [hmr-1p::HMR-1::GFP::
unc-54 3'UTR; unc-119(+)]
Caenorhabditis
Genetics Center
FT250
Genetic reagent
(C. elegans)
kyIs4[ceh-23-unc-76-gfp::lin-15]XCaenorhabditis
Genetics Center
CX2565
Genetic reagent
(C. elegans)
hmnEx149[dyf-7p::DYF-7
(ZP-sfGFP)-mCherry;
rol-6(su1006)]
DOI: 10.1101/393850
Genetic reagent
(E. coli)
elt-1(RNAi) expressed
in HT115 (DE3)
Source BioScienceC. elegans RNAi
collection (Vadel)
RNAi Bacteria,
Clone ID: 10019-B-8
Genetic reagent
(E. coli)
hmr-1(RNAi) expressed
in HT115 (DE3)
Source BioScienceC. elegans RNAi
collection (Ahringer)
RNAi Bacteria,
Clone ID: I-5F23
Genetic reagent
(E. coli)
dlg-1(RNAi) expressed
in HT115 (DE3)
Source BioScienceC. elegans RNAi
collection (Ahringer)
RNAi Bacteria,
Clone ID: X-8A08
Software, algorithmFijihttps://fiji.sc/

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