Relationship between dose, distance and time in Sonic Hedgehog-mediated regulation of anteroposterior polarity in the chick limb

Y Yang, G Drossopoulou, P T Chuang, D Duprez, E Marti, D Bumcrot, Neil Vargesson, Jonathan D Clarke, L Niswander, A McMahon, C Tickle

Research output: Contribution to journalArticle

270 Citations (Scopus)

Abstract

Anteroposterior polarity in the vertebrate limb is thought to be regulated in response to signals derived from a specialized region of distal posterior mesenchyme, the zone of polarizing activity. Sonic Hedgehog (Shh) is expressed in the zone of polarizing activity and appears to mediate the action of the zone of polarizing activity. Here we have manipulated Shh signal in the limb to assess whether it acts as a long-range signal to directly pattern all the digits. Firstly, we demonstrate that alterations in digit development are dependent upon the dose of Shh applied. DiI-labeling experiments indicate that cells giving rise to the extra digits lie within a 300 microm radius of a Shh bead and that the most posterior digits come from cells that lie very close to the bead. A response to Shh involves a 12-16 hour period in which no irreversible changes in digit pattern occur. Increasing the time of exposure to Shh leads to specification of additional digits, firstly digit 2, then 3, then 4. Cell marking experiments demonstrate that cells giving rise to posterior digits are first specified as anterior digits and later adopt a more posterior character. To monitor the direct range of Shh signalling, we developed sensitive assays for localizing Shh by attaching alkaline phosphatase to Shh and introducing cells expressing these forms into the limb bud. These experiments demonstrate that long-range diffusion across the anteroposterior axis of the limb is possible. However, despite a dramatic difference in their diffusibility in the limb mesenchyme, the two forms of alkaline phosphatase-tagged Shh proteins share similar polarizing activity. Moreover, Shh-N (aminoterminal peptide of Shh)-coated beads and Shh-expressing cells also exhibit similar patterning activity despite a significant difference in the diffusibility of Shh from these two sources. Finally, we demonstrate that when Shh-N is attached to an integral membrane protein, cells transfected with this anchored signal also induce mirror-image pattern duplications in a dose-dependent fashion similar to the zone of polarizing activity itself. These data suggest that it is unlikely that Shh itself signals digit formation at a distance. Beads soaked in Shh-N do not induce Shh in anterior limb mesenchyme ruling out direct propagation of a Shh signal. However, Shh induces dose-dependent expression of Bmp genes in anterior mesenchyme at the start of the promotion phase. Taken together, these results argue that the dose-dependent effects of Shh in the regulation of anteroposterior pattern in the limb may be mediated by some other signal(s). BMPs are plausible candidates.
Original languageEnglish
Pages (from-to)4393-4404
Number of pages12
JournalDevelopment
Volume124
Issue number21
Publication statusPublished - 1 Nov 1997

Fingerprint

Hedgehogs
Extremities
Mesoderm
Alkaline Phosphatase
Hedgehog Proteins
Limb Buds
Vertebrates
Membrane Proteins
Gene Expression
Peptides

Keywords

  • Animals
  • Antigens, CD4
  • Body Patterning
  • COS Cells
  • Cell Membrane
  • Chick Embryo
  • Dose-Response Relationship, Drug
  • Gene Expression Regulation, Developmental
  • Hedgehog Proteins
  • Proteins
  • Recombinant Proteins
  • Signal Transduction
  • Trans-Activators
  • Wing

Cite this

Yang, Y., Drossopoulou, G., Chuang, P. T., Duprez, D., Marti, E., Bumcrot, D., ... Tickle, C. (1997). Relationship between dose, distance and time in Sonic Hedgehog-mediated regulation of anteroposterior polarity in the chick limb. Development, 124(21), 4393-4404.

Relationship between dose, distance and time in Sonic Hedgehog-mediated regulation of anteroposterior polarity in the chick limb. / Yang, Y; Drossopoulou, G; Chuang, P T; Duprez, D; Marti, E; Bumcrot, D; Vargesson, Neil; Clarke, Jonathan D; Niswander, L; McMahon, A; Tickle, C.

In: Development, Vol. 124, No. 21, 01.11.1997, p. 4393-4404.

Research output: Contribution to journalArticle

Yang, Y, Drossopoulou, G, Chuang, PT, Duprez, D, Marti, E, Bumcrot, D, Vargesson, N, Clarke, JD, Niswander, L, McMahon, A & Tickle, C 1997, 'Relationship between dose, distance and time in Sonic Hedgehog-mediated regulation of anteroposterior polarity in the chick limb', Development, vol. 124, no. 21, pp. 4393-4404.
Yang Y, Drossopoulou G, Chuang PT, Duprez D, Marti E, Bumcrot D et al. Relationship between dose, distance and time in Sonic Hedgehog-mediated regulation of anteroposterior polarity in the chick limb. Development. 1997 Nov 1;124(21):4393-4404.
Yang, Y ; Drossopoulou, G ; Chuang, P T ; Duprez, D ; Marti, E ; Bumcrot, D ; Vargesson, Neil ; Clarke, Jonathan D ; Niswander, L ; McMahon, A ; Tickle, C. / Relationship between dose, distance and time in Sonic Hedgehog-mediated regulation of anteroposterior polarity in the chick limb. In: Development. 1997 ; Vol. 124, No. 21. pp. 4393-4404.
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T1 - Relationship between dose, distance and time in Sonic Hedgehog-mediated regulation of anteroposterior polarity in the chick limb

AU - Yang, Y

AU - Drossopoulou, G

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AU - Duprez, D

AU - Marti, E

AU - Bumcrot, D

AU - Vargesson, Neil

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AB - Anteroposterior polarity in the vertebrate limb is thought to be regulated in response to signals derived from a specialized region of distal posterior mesenchyme, the zone of polarizing activity. Sonic Hedgehog (Shh) is expressed in the zone of polarizing activity and appears to mediate the action of the zone of polarizing activity. Here we have manipulated Shh signal in the limb to assess whether it acts as a long-range signal to directly pattern all the digits. Firstly, we demonstrate that alterations in digit development are dependent upon the dose of Shh applied. DiI-labeling experiments indicate that cells giving rise to the extra digits lie within a 300 microm radius of a Shh bead and that the most posterior digits come from cells that lie very close to the bead. A response to Shh involves a 12-16 hour period in which no irreversible changes in digit pattern occur. Increasing the time of exposure to Shh leads to specification of additional digits, firstly digit 2, then 3, then 4. Cell marking experiments demonstrate that cells giving rise to posterior digits are first specified as anterior digits and later adopt a more posterior character. To monitor the direct range of Shh signalling, we developed sensitive assays for localizing Shh by attaching alkaline phosphatase to Shh and introducing cells expressing these forms into the limb bud. These experiments demonstrate that long-range diffusion across the anteroposterior axis of the limb is possible. However, despite a dramatic difference in their diffusibility in the limb mesenchyme, the two forms of alkaline phosphatase-tagged Shh proteins share similar polarizing activity. Moreover, Shh-N (aminoterminal peptide of Shh)-coated beads and Shh-expressing cells also exhibit similar patterning activity despite a significant difference in the diffusibility of Shh from these two sources. Finally, we demonstrate that when Shh-N is attached to an integral membrane protein, cells transfected with this anchored signal also induce mirror-image pattern duplications in a dose-dependent fashion similar to the zone of polarizing activity itself. These data suggest that it is unlikely that Shh itself signals digit formation at a distance. Beads soaked in Shh-N do not induce Shh in anterior limb mesenchyme ruling out direct propagation of a Shh signal. However, Shh induces dose-dependent expression of Bmp genes in anterior mesenchyme at the start of the promotion phase. Taken together, these results argue that the dose-dependent effects of Shh in the regulation of anteroposterior pattern in the limb may be mediated by some other signal(s). BMPs are plausible candidates.

KW - Animals

KW - Antigens, CD4

KW - Body Patterning

KW - COS Cells

KW - Cell Membrane

KW - Chick Embryo

KW - Dose-Response Relationship, Drug

KW - Gene Expression Regulation, Developmental

KW - Hedgehog Proteins

KW - Proteins

KW - Recombinant Proteins

KW - Signal Transduction

KW - Trans-Activators

KW - Wing

M3 - Article

C2 - 9334287

VL - 124

SP - 4393

EP - 4404

JO - Development

JF - Development

SN - 0950-1991

IS - 21

ER -