QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

This Article Summary Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Latimer, A. J. Articles by Appel, B. Search for Related Content PubMed PubMed Citation Articles by Latimer, A. J. Articles by Appel, B.

Delta-Notch signaling induces hypochord development in zebrafish

Department of Biological Sciences, Vanderbilt University, Nashville, TN 37232, USA

View larger version (129K): [in a new window]   Fig. 1. Hypochord precursors are closely associated with notochord and muscle precursors near the edge of the shield. (A) View of dorsal margin region of 6 hpf (shield stage) embryo. Caged fluorescein was photoactivated in a cluster of three to five cells near the margin (broken black line) and centered about 15 cell diameters from the dorsal midline (broken white line). (B) Side view of trunk region of living embryo at 24 hpf showing labeled hypochord (hc) and notochord (nc) cells. Floor-plate (fp) cells were not labeled. (C) Transverse section of 24 hpf embryo in which the photoactivated fluorescein signal was converted to a blue precipitate. Hypochord, muscle (mu) cells near the periphery of the myotome and periderm (asterisk) were labeled. The periderm precursors were enveloping layer cells that were above the hypochord and muscle precursors at the time of photoactivation. (D) Transverse section of 30 hpf embryo labeled with F59 antibody to reveal slow muscle cells (red). Hypochord (green) and three slow muscle cells (yellow, arrows) were labeled by photoactivated fluorescein. (E) Dorsal view, anterior towards the top, of 9.5 hpf living embryo. Cells labeled by photoactivated fluorescein (green) are next to notochord cells (between arrows). Scale bar: 20 µm for all panels.

View larger version (105K): [in a new window]   Fig. 2. Presumptive hypochord precursors express notch5 and her4. (A,B) Dorsal views, anterior towards the top, of 8.5 hpf (A) and 9.5 hpf (B) embryos probed for notch5 expression. Cells near the dorsal midline express notch5 (arrows). (C,D) Dorsal views, anterior towards the top, of 8.5 hpf (C) and 9.5 hpf (D) embryos probed for her4 expression. Cells near the dorsal midline express her4 (arrows). (E) Sagittal section, anterior towards the left, of 9.5 hpf embryo probed for notch5 expression. Deep mesendodermal cells express notch5 (arrows). (F) Transverse section of 9.5 hfp embryo probed for her4 (blue, arrows) and myod (red) expression. Adaxial cells, marked by myod expression, do not express her4. (G) Transverse section of 10 hpf embryo probed for notch5 (blue, arrows) and myod (red) expression. myod-positive adaxial cells do not express notch5. (H) Dorsal view, anterior towards the top, of 9.5 hpf embryo probed for her4 (blue) and ntl (red) expression. Broken lines indicate approximate positions of transverse sections shown in I,J. (I,J) Transverse sections of 9.5 hpf embryo probed for her4 (blue) and ntl (red) expression. (I) Posterior transverse section with several double-labeled cells (arrows). (J) Anterior transverse section in which one her4-positive cell expresses ntl (arrow) whereas two others do not. (K) Transverse section of 9.5 hpf embryo probed for notch5 (blue, arrows) and ntl (red) expression. The notch5-positive cell on the right expresses ntl whereas the one on the left does not. (L) Transverse section of 9.5 hpf embryo probed for notch5 (blue) and her4 expression. Cells bordering the midline express both genes (arrows). (M) Transverse section of 11.5 hpf embryo probed for her4 expression. A single hypochord cell (arrow), ventral to notochord (nc), and neural keel (nk) cells expresses her4. (N) Transverse section of YPC stage embryo probed for notch1a expression. Many dorsal mesoderm cells express notch1a. Scale bar: 100 µm for A-D,H; 50 µm for E; 25 µm for I-N.

View larger version (94K): [in a new window]   Fig. 3. Paraxial mesoderm cells next to midline precursors express dlc and dld. (A,B) Dorsal views, anterior towards the top, of 8.5 hpf (A) and 9.5 hpf (B) embryos probed for dld expression. Cells at the dorsal midline (arrows) do not express dld. (C,D) Dorsal views, anterior towards the top, of 8.5 hpf (C) and 9.5 hpf (D) embryos probed for dlc expression. Similar to dld, dorsal midline cells (arrows) do not express dlc. (E,F) Transverse sections of 9.5 hpf embryos probed for dld (E) and dlc (F) expression. Paraxial mesoderm expresses dld and dlc, whereas midline (flanked by arrows) and ectoderm (asterisks) do not. (G) Dorsal view, anterior towards the top, of 9.5 hpf embryo probed for dlc (purple) and ntl (red) expression. (H) Transverse section of 9.5 hpf embryo probed for dlc (blue) and ntl (red) expression. In both G and H, dlc-expressing paraxial mesoderm borders ntl-positive midline precursors (arrows). (I) Transverse section of 9.5 hpf embryo showing her4-positive cells (red, arrows) adjacent to dlc-expressing cells (blue, arrows). Scale bar: 100 µm for A-D,G; 20 µm for E,F,H,I.

View larger version (128K): [in a new window]   Fig. 4. Delta functions are required for hypochord development. (A-F) Side views, anterior towards the left, of the trunk region of 24-26 hpf embryos probed for col2a1 expression to mark floor-plate (fp) and hypochord (hc) cells. (A) Wild-type embryo showing the normal pattern of floor-plate and hypochord cells. (B) Homozygous mutant aeiAR33 embryo with a deficit of hypochord cells. Floor plate appeared normal. (C) Wild-type embryo injected with dlc antisense morpholino oligonucleotides (MO). Hypochord cell number was reduced, whereas floor plate appeared normal. (D) Homozygous mutant aeiAR33 embryo injected with dlc MO. Most hypochord cells were absent, whereas floor plate appeared normal. (E-G) Dorsal views, anterior towards the top, of 9.5 hpf embryos from aeiAR33/+ intercross injected with dlc MO. notch5 expression was normal (E), whereas her4 expression was reduced (F) or absent (G) (compare with Fig. 2D). Embryos in F,G were double-labeled to reveal pax2.1 expression in pronephros (asterisks), which was normal. (H-K) Dorsal views, anterior towards the top, of 10.5 hpf embryos probed for isl1 expression to reveal prospective Rohon-Beard (RB) neurons and primary motoneurons (pmn). (H) Wild-type, uninjected embryo. (I) aeiAR33 mutant embryo showing small increase in the number of prospective primary motoneurons and RBs. (J) Embryo from aeiAR33/+ intercross injected with dlc MO. The neural phenotype is similar to that of the uninjected aeiAR33 mutant embryo shown in I. (K) Embryo from aeiAR33/+ intercross injected with dla MO. The number of prospective primary motoneurons and RB cells was greater than the embryos shown in I,J. Scale bar: 20 µm in A-D; 80 µm in E-K.

View larger version (71K): [in a new window]   Fig. 5. Constitutive Notch activity promotes her4 and inhibits ntl expression. (A) Transverse section showing lateral region of 9.5 hpf embryo in which NICDMT expression was induced during gastrulation. Some cells that expressed NICDMT (brown) also expressed her4 (blue, arrows). (B,C) Dorsal views, anterior towards the top, of 9.5 hpf embryos labeled for ntl (blue) and NICDMT (brown) expression. These embryos were injected with mRNA encoding NICDMT in a single cell at the eight-cell stage. (B) Embryos in which NICDMT-positive cells were restricted to ventrolateral regions (asterisks). Midline cells expressed ntl normally (arrows). (C) Embryos in which NICDMT-expressing cells occupied the dorsal midline region. These embryos had fewer midline cells that expressed ntl (arrows). (D) Transverse section through dorsal midline region of 9.5 hpf embryo treated as embryos described in B,C. Two NICDMT-positive cells (brown, arrows) occupied the notochord precursor domain but did not express ntl (blue), whereas neighboring cells expressed ntl normally. Scale bar: 20 µm in A,D; 120 µm in B,C.

View larger version (13K): [in a new window]   Fig. 6. Model for Delta-Notch-mediated induction of hypochord fate. At early gastrula stage, ntl-expressing midline precursors occupy the dorsal margin. During gastrulation, Delta signaling, primarily from paraxial mesoderm expression of DeltaD and DeltaC, induce ntl-positive midline precursors to express her4. These cells downregulate ntl expression and, after completion of gastrulation, migrate beneath notochord to form hypochord.