Sonic Hedgehog 2
Sofie Kovalcheck
This signaling molecule is key in regulating embryonic morphogenesis in all animals. SHH controls organogenesis and the organization of the central nervous system, limbs, digits and many other parts of the body. Sonic hedgehog is a morphogen that patterns the developing embryo using a concentration gradient characterized by the French flag model.[6] This model has a non-uniform distribution of SHH molecules which governs different cell fates according to concentration. Mutations in this gene can cause holoprosencephaly, a failure of splitting in the cerebral hemispheres,[7] as demonstrated in an experiment using SHH knock-out mice in which the forebrain midline failed to develop and instead only a single fused telencephalic vesicle resulted.[8]
Sonic hedgehog still plays a role in differentiation, proliferation, and maintenance of adult tissues. Abnormal activation of SHH signaling in adult tissues has been implicated in various types of cancers including breast, skin, brain, liver, gallbladder and many more.[9]
The hedgehog gene (hh) was first identified in the fruit fly Drosophila melanogaster in the classic Heidelberg screens of Christiane Nsslein-Volhard and Eric Wieschaus, as published in 1980.[10] These screens, which led to the researchers winning a Nobel Prize in 1995 along with developmental geneticist Edward B. Lewis, identified genes that control the segmentation pattern of the Drosophila embryos. The hh loss of function mutant phenotype causes the embryos to be covered with denticles, i.e. small pointy projections resembling the spikes of a hedgehog. Investigations aimed at finding a hedgehog equivalent in vertebrates by Philip Ingham, Andrew P. McMahon and Clifford Tabin revealed three homologous genes.[11][12][13][14]
Two of these genes, desert hedgehog and Indian hedgehog, were named for species of hedgehogs, while sonic hedgehog was named after the video game character Sonic the Hedgehog.[15][16] The gene was named by Robert Riddle, a postdoctoral fellow at the Tabin Lab, after his wife Betsy Wilder came home with a magazine containing an advert for the game Sonic the Hedgehog.[17][18][19] In the zebrafish, two of the three vertebrate hh genes are duplicated: SHH a[20] and SHH b[21] (formerly described as tiggywinkle hedgehog, named for Mrs. Tiggy-Winkle, a character from Beatrix Potter's books for children) and ihha and ihhb[22] (formerly described as echidna hedgehog, named for the spiny anteater and not for the character Knuckles the Echidna in the Sonic franchise).
Sonic hedgehog has also been shown to act as an axonal guidance cue. It has been demonstrated that SHH attracts commissural axons at the ventral midline of the developing spinal cord.[34] Specifically, SHH attracts retinal ganglion cell (RGC) axons at low concentrations and repels them at higher concentrations.[35] The absence (non-expression) of SHH has been shown to control the growth of nascent hind limbs in cetaceans[36] (whales and dolphins).
The SHH gene is a member of the hedgehog gene family with five variations of DNA sequence alterations or splice variants.[37] SHH is located on chromosome seven and initiates the production of Sonic Hedgehog protein.[37] This protein sends short- and long-range signals to embryonic tissues to regulate development.[38] If the SHH gene is mutated or absent, the protein Sonic Hedgehog cannot do its job properly. Sonic hedgehog contributes to cell growth, cell specification and formation, structuring and organization of the body plan.[39] This protein functions as a vital morphogenic signaling molecule and plays an important role in the formation of many different structures in developing embryos.[39] The SHH gene affects several major organ systems, such as the nervous system, cardiovascular system, respiratory system and musculoskeletal system.[37][38] Mutations in the SHH gene can cause malformation of components of these systems, which can result in major problems in the developing embryo. The brain and eyes, for example, can be significantly impacted by mutations in this gene and cause disorders such as Microphthalmia and Holoprosencephaly.[39] Microphthalmia is a condition that affects the eyes, which results in small, underdeveloped tissues in one or both eyes.[39] This can lead to issues ranging from a coloboma to a single small eye to the absence of eyes altogether.[38] Holoprosencephaly is a condition most commonly caused by a mutation of the SHH gene that causes improper separation or turn of the left and right brain[40] and facial dysmorphia.[38][39] Many systems and structures rely heavily on proper expression of the SHH gene and subsequent sonic hedgehog protein, earning it the distinction of being an essential gene to development.
The neural tube itself is the initial groundwork of the vertebrate CNS, and the floor plate is a specialized structure, located at the ventral midpoint of the neural tube. Evidence supporting the notochord as the signaling center comes from studies in which a second notochord is implanted near a neural tube in vivo, leading to the formation of an ectopic floor plate within the neural tube.[44]
Sonic hedgehog is the secreted protein that mediates signaling activities of the notochord and floor plate.[45] Studies involving ectopic expression of SHH in vitro[46] and in vivo[47] result in floor plate induction and differentiation of motor neuron and ventral interneurons. On the other hand, mice mutants for SHH lack ventral spinal cord characteristics.[48] In vitro blocking of SHH signaling using antibodies against it shows similar phenotypes.[47] SHH exerts its effects in a concentration-dependent manner,[49] so that a high concentration of SHH results in a local inhibition of cellular proliferation.[50] This inhibition causes the floor plate to become thin compared to the lateral regions of the neural tube. Lower concentration of SHH results in cellular proliferation and induction of various ventral neural cell types.[47] Once the floor plate is established, cells residing in this region will subsequently express SHH themselves,[50] generating a concentration gradient within the neural tube.
Although there is no direct evidence of a SHH gradient, there is indirect evidence via the visualization of Patched (Ptc) gene expression, which encodes for the ligand binding domain of the SHH receptor[51] throughout the ventral neural tube.[52] In vitro studies show that incremental two- and threefold changes in SHH concentration give rise to motor neuron and different interneuronal subtypes as found in the ventral spinal cord.[53] These incremental changes in vitro correspond to the distance of domains from the signaling tissue (notochord and floor plate) which subsequently differentiates into different neuronal subtypes as it occurs in vitro.[54] Graded SHH signaling is suggested to be mediated through the Gli family of proteins, which are vertebrate homologues of the Drosophila zinc-finger-containing transcription factor Cubitus interruptus (Ci). Ci is a crucial mediator of hedgehog (Hh) signaling in Drosophila.[55] In vertebrates, three different Gli proteins are present, viz. Gli1, Gli2 and Gli3, which are expressed in the neural tube.[56] Mice mutants for Gli1 show normal spinal cord development, suggesting that it is dispensable for mediating SHH activity.[57] However, Gli2 mutant mice show abnormalities in the ventral spinal cord, with severe defects in the floor plate and ventral-most interneurons (V3).[58] Gli3 antagonizes SHH function in a dose-dependent manner, promoting dorsal neuronal subtypes. SHH mutant phenotypes can be rescued in a SHH/Gli3 double mutant.[59] Gli proteins have a C-terminal activation domain and an N-terminal repressive domain.[56][60]
SHH is suggested to promote the activation function of Gli2 and inhibit repressive activity of Gli3. SHH also seems to promote the activation function of Gli3, but this activity is not strong enough.[59] The graded concentration of SHH gives rise to graded activity of Gli 2 and Gli3, which promote ventral and dorsal neuronal subtypes in the ventral spinal cord. Evidence from Gli3 and SHH/Gli3 mutants show that SHH primarily regulates the spatial restriction of progenitor domains rather than being inductive, as SHH/Gli3 mutants show intermixing of cell types.[59][61]
SHH also induces other proteins with which it interacts, and these interactions can influence the sensitivity of a cell towards SHH. Hedgehog-interacting protein (HHIP) is induced by SHH, which in turn attenuates its signaling activity.[62] Vitronectin is another protein that is induced by SHH; it acts as an obligate co-factor for SHH signaling in the neural tube.[63]
It is important to note that SHH is not the only signaling molecule exerting an effect on the developing neural tube. Many other molecules, pathways and mechanisms are active (e.g., RA, FGF, BMP), and complex interactions between SHH and other molecules are possible. BMPs are suggested to play a critical role in determining the sensitivity of neural cell to SHH signaling. Evidence supporting this comes from studies using BMP inhibitors that ventralize the fate of the neural plate cell for a given SHH concentration.[65] On the other hand, mutation in BMP antagonists (e.g., noggin) produces severe defects in the ventral-most characteristics of the spinal cord, followed by ectopic expression of BMP in the ventral neural tube.[66] Interactions of SHH with Fgf and RA have not yet been studied in molecular detail.
The concentration- and time-dependent, cell-fate-determining activity of SHH in the ventral neural tube makes it a prime example of a morphogen. In vertebrates, SHH signaling in the ventral portion of the neural tube is most notably responsible for the induction of floor plate cells and motor neurons.[67] SHH emanates from the notochord and ventral floor plate of the developing neural tube to create a concentration gradient that spans the dorso-ventral axis and is antagonized by an inverse Wnt gradient, which specifies the dorsal spinal cord.[68][69] Higher concentrations of the SHH ligand are found in the most ventral aspects of the neural tube and notochord, while lower concentrations are found in the more dorsal regions of the neural tube.[68] The SHH concentration gradient has been visualized in the neural tube of mice engineered to express a SHH::GFP fusion protein to show this graded distribution of SHH during the time of ventral neural tube patterning.[70]
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