Cholesterol modification restricts the spread of Shh gradient in the limb bud

Li, Yina; Zhang, Huimin; Litingtung, Ying; Chiang, Chin

doi:10.1073/pnas.0600124103

Cited by 162 publications

(135 citation statements)

References 47 publications

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“…In vertebrates, ShhN showed reduced signaling capabilities and diffusion in comparison with the multimeric wild-type form (6, 38 -40), but the opposite situation was found in the limb bud (24) and in the Drosophila wing imaginal disc (41,42). Thus, based on those reports, it was unclear whether multimerization restricts or promotes morphogen spreading, also because observed differences had been linked to morphogen lipidation rather than multimerization or differential HS binding of monomeric and multimeric morphogens (24,42,43). In our work, we demonstrate variable ionic interactions between HS and ShhN or ShhNp and demonstrate that ionic interactions depend on HS sulfation.…”

Section: Discussionmentioning

confidence: 99%

Dual Roles of the Cardin-Weintraub Motif in Multimeric Sonic Hedgehog

Farshi¹,

Ohlig²,

Pickhinke³

et al. 2011

Journal of Biological Chemistry

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The fly morphogen Hedgehog (Hh) and its mammalian orthologs, Sonic, Indian, and Desert hedgehog, are secreted signaling molecules that mediate tissue patterning during embryogenesis and function in tissue homeostasis and regeneration in the adult. The function of all Hh family members is regulated at the levels of morphogen multimerization on the surface of producing cells, multimer release, multimer diffusion to target cells, and signal reception. These mechanisms are all known to depend on interactions of positively charged Hh amino acids (the Cardin-Weintraub (CW) motif) with negatively charged heparan sulfate (HS) glycosaminoglycan chains. However, a precise mechanistic understanding of these interactions is still lacking. The proteins of the Hh family are powerful morphogens that control growth and patterning of developing embryos. Current models for Hh activity suggest that the morphogen disperses from a localized source and forms a gradient that patterns fields of responsive cells expressing the Hh receptor Patched (Ptc).Genetic evidence suggests that this process critically depends on heparan sulfate proteoglycan (HSPG) 2 expression. Upon secretion to the cell surface, Drosophila Hh forms nanoscale oligomers on the cell surface that co-localize with HSPGs (1). HS binds to the Cardin-Weintraub (CW) motif found on all known Hhs and regulates their function in flies (2, 3) and mice (4). In Drosophila (5) as well as in mammalian cell culture (6, 7), Hhs are always released from producing cells in multimeric form, as demonstrated by gel filtration analysis of the soluble morphogen. Release of the multimeric morphogen (the processed Hh N-terminal signaling domain, designated HhNp) from the cell surface depends on the expression of Dispatched (8) and A disintegrin and metalloprotease (ADAM) family members that mediate ectodomain shedding from transfected Bosc23 cells (9). HS is involved in the formation of the HhNp extracellular gradient, which, in the fly, depends on the expression of the Drosophila Exostosin (Ext) family of proteins, encoded by the genes tout velu (ttv), brother of tout velu, and sister of tout velu and the glycosylphosphatidylinositollinked HSPGs Dally and Dally-like, corresponding to vertebrate glypicans (2, 3, 10). HS expression and Dally-like/glypican expression are also essential for signal reception and modulation on Ptc-expressing receiving cells (10 -14) and participate in HhNp-Ihog interaction (15). However, the essential role of direct morphogen-HSPG interactions in embryonic patterning was recently challenged (16). In that report, transgenic mice made deficient in two ShhNp CW amino acid residues implicated in HS binding (17) lacked an Shh-related phenotype, suggesting that direct morphogen-HS interactions were not essential for normal development. However, in that report as well as in others (16 -18), CW-dependent HS interactions were characterized using a recombinant, non-physiological monomeric morphogen termed ShhN, whereas embryogenesis depends entirely on the activity of morp...

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Section: Discussionmentioning

confidence: 99%

Dual Roles of the Cardin-Weintraub Motif in Multimeric Sonic Hedgehog

Farshi¹,

Ohlig²,

Pickhinke³

et al. 2011

Journal of Biological Chemistry

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“…This modified peptide forms a posterior to anterior gradient in the developing limb bud (Fig. 3C) (Zeng et al 2001;Li et al 2006). Genetic alteration such that the active SHH peptide is no longer cholesterated increases its spread, which results in formation of additional anterior digits ( preaxial polydactyly) (Li et al 2006).…”

Section: Early Specification and Progressive Expansion/differentiatiomentioning

confidence: 99%

“…3C) (Zeng et al 2001;Li et al 2006). Genetic alteration such that the active SHH peptide is no longer cholesterated increases its spread, which results in formation of additional anterior digits ( preaxial polydactyly) (Li et al 2006). In contrast, the palmitoyl modification increases long-range signaling by promoting the formation of multimeric SHH ligand complexes (for more details, see Chen et al 2004).…”

Section: Early Specification and Progressive Expansion/differentiatiomentioning

confidence: 99%

Vertebrate Limb Development: Moving from Classical Morphogen Gradients to an Integrated 4-Dimensional Patterning System

Bénazet¹,

Zeller²

2009

Cold Spring Harbor Perspectives in Biology

135

117

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A wealth of classical embryological manipulation experiments taking mainly advantage of the chicken limb buds identified the apical ectodermal ridge (AER) and the zone of polarizing activity (ZPA) as the respective ectodermal and mesenchymal key signaling centers coordinating proximodistal (PD) and anteroposterior (AP) limb axis development. These experiments inspired Wolpert's French flag model, which is a classic among morphogen gradient models. Subsequent molecular and genetic analysis in the mouse identified retinoic acid as proximal signal, and fibroblast growth factors (FGFs) and sonic hedgehog (SHH) as the essential instructive signals produced by AER and ZPA, respectively. Recent studies provide good evidence that progenitors are specified early with respect to their PD and AP fates and that morpho-regulatory signaling is also required for subsequent proliferative expansion of the specified progenitor pools. The determination of particular fates seems to occur rather late and depends on additional signals such as bone morphogenetic proteins (BMPs), which indicates that cells integrate signaling inputs over time and space. The coordinate regulation of PD and AP axis patterning is controlled by an epithelial-mesenchymal feedback signaling system, in which transcriptional regulation of the BMP antagonist Gremlin1 integrates inputs from the BMP, SHH, and FGF pathways. Vertebrate limb-bud development is controlled by a 4-dimensional (4D) patterning system integrating positive and negative regulatory feedback loops, rather than thresholds set by morphogen gradients.

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“…There are several mechanisms that assist in shaping morphogen gradients. Post-translational modification, such as the addition of cholesterol to Sonic hedgehog, can regulate the range of spreading of this factor through tissue in several systems (Li et al, 2006). For FGF8, endocytosis regulates the amount of extracellular protein available for signaling, and the distance the protein can travel from the source of production (Scholpp and Brand, 2004).…”

Section: Introductionmentioning

confidence: 99%

Xenopus Lefty requires proprotein cleavage but not N‐linked glycosylation to inhibit nodal signaling

Westmoreland

Takahashi

Wright

2007

Developmental Dynamics

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The Nodal and Nodal-related morphogens are utilized for the specification of distinct cellular identity throughout development by activating discrete target genes in a concentration-dependant manner. Lefty is a principal extracellular antagonist involved in the spatiotemporal regulation of the Nodal morphogen gradient during mesendoderm induction. The Xenopus Lefty proprotein contains a single N-linked glycosylation motif in the mature domain and two potential cleavage sites that would be expected to produce long (Xlefty L ) and short (Xlefty S ) isoforms. Here we demonstrate that both isoforms were secreted from Xenopus oocytes, but that Xlefty L is the only isoform detected when embryonic tissue was analyzed. In mesoderm induction assays, Xlefty L is the functional blocker of Xnr signaling. When secreted from oocytes, vertebrate Lefty molecules were N-linked glycosylated. However, glycan addition was not required to inhibit Xnr signaling and did not influence its movement through the extracellular space. These findings demonstrate that Lefty molecules undergo post-translational modifications and that some of these modifications are required for the Nodal inhibitory function.

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Cholesterol modification restricts the spread of Shh gradient in the limb bud

Cited by 162 publications

References 47 publications

Dual Roles of the Cardin-Weintraub Motif in Multimeric Sonic Hedgehog

Dual Roles of the Cardin-Weintraub Motif in Multimeric Sonic Hedgehog

Vertebrate Limb Development: Moving from Classical Morphogen Gradients to an Integrated 4-Dimensional Patterning System

Xenopus Lefty requires proprotein cleavage but not N‐linked glycosylation to inhibit nodal signaling

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