Mammalian Embryo: Hox Genes

Abstract

Hox genes are evolutionarily conserved transcription factors that play important roles in establishing the basic body plan of animals. Mammals have 39 Hox genes clustered into four chromosomal complexes. This gene family regulates the regional character and patterning of diverse structures along the anterior–posterior (A/P) axis of the embryo. Nested patterns of Hox gene expression generate a Hox combinatorial protein code that orchestrates the morphogenesis of structures in the nervous system, axial skeleton, limbs, intestine and many other tissues. In light of their key role in regulating morphogenesis across animal species, modulation of Hox expression or function over the course of evolution is believed to have been important in generating diversity.

Key Concepts

  • Axial patterning is the process that generates different regional characteristics during the development of a tissue, such as the nervous system or skeleton.
  • Hox genes encode a family of transcription factors that regulate the identity of structures along the anterior–posterior (A/P) axis of embryos.
  • Co‐linearity is the correlation between the order of Hox genes along a chromosome and their expression along the axis of an embryo.
  • The collection of Hox proteins expressed in a region provides a combinatorial code for specifying diversity.
  • Posterior prevalence is a model for explaining why some Hox proteins dominate over others when they are co‐expressed.
  • Selector genes control the identity of a tissue.
  • Homeotic transformation is the conversion of one structure into another due to loss or gain of selector gene activity.
  • Segmentation subdivides a developing tissue, such as the hindbrain or skeleton, into repeating units that ultimately generate different structures along an axis.
  • Subfunctionalisation is the partitioning of function and regulation between duplicated genes compared with the ancestral gene.
  • Changes in Hox expression or function may be important for generating differences in structures during evolution of vertebrates.
  • Cooption refers to the redeployment or coupling of a common molecular pathway to multiple patterning processes.

Keywords: homeobox; axial patterning; gene regulation; transcription factors; embryogenesis

Figure 1. Schematic representation of the mammalian Hox gene clusters. The four mammalian clusters, named A to D, arose by two rounds of whole genome duplications from a single Hox cluster that is hypothesised to have closely resembled the Hox cluster of the invertebrate chordate amphioxus. The similarity in structure and function of the mammalian and Drosophila complexes supports the view that a Hox cluster composed of Anterior, Group3, Central and Posterior classes of Hox genes existed before the divergence of the protostome and deuterstome lineages. The mammalian Hox genes are arranged in the same transcriptional orientation, so the cluster is said to have a 3′–5′ polarity. 3′ Hox genes are expressed first, followed by the expression of progressively more 5′ genes, a phenomenon referred to as temporal co‐linearity. Mammalian Hox gene expression extends from the posterior of the embryo to an anterior boundary characteristic for each Hox gene. Spatial co‐linearity describes the correlation between the axial level of this anterior boundary and Hox gene order within the cluster; the further 5′ a gene is located within the cluster, the more posterior is its anterior expression limit.
Figure 2. Regulatory subfunctionalisation. In this example, the ancestral gene is regulated by three discrete cis‐elements (red, yellow and blue). After duplication, the gene copies will be functionally redundant. The most likely outcome is the loss of one of the gene copies to deleterious mutations. However, both gene copies may be retained to perform all of the functions of the ancestral gene if complementary, loss‐of‐subfunction mutations (white circles) occur in the modular enhancers.
Figure 3. Activation of the Hox genes in the presomitic mesoderm and throughout the trunk of the developing mouse embryo is accompanied by dynamic changes in histone methylation patterns and their chromatin domains. (a) During gastrulation, ingressing cells past through the primitive streak near the node (white arrowhead) and acquire their unique A/P character. Cells within the presomitic mesoderm (light gray; PSM) migrate anteriorly from the node and form somites (dark gray squares). The activation of each gene within the complex is preceded by changes in their acetylation and methylation patterns. Before their activation, they are covered in repressive marks (such as H3K27me3) which are replaced by activation marks (such as H3K4me3). (b) At later time points, additional cells within the complexes express Hox genes and the change in their chromatin marks reflect their collinear activation profiles. (c) These uniform changes in the acetylation/methylation patterns correlate with their local associations into either a repressed domain or an active domain. The distribution of genes between the inactive and the active domains changes depending on the A/P level and corresponds to their transcriptional status.
Figure 4. Hox gene expression is controlled by microRNAs and long non‐coding RNAs. (a) Several microRNAs (miR; pink arrowheads) and long non‐coding RNAs (lnRNAs; purple boxes) have been identified within, or flanking, the Hox complexes. (b) lncRNAs can function as both gene activators or repressors by selectively recruiting Polycomb or Trithorax protein complexes to specific sites within the genome. In this example, the lncRNA, Hotair, specifically guides the Polycomb Repressive Complex 2 and LSD1 to the posterior HoxD genes resulting in the accumulation of H3K27me3 chromatin marks (red pins), silencing their transcription. (c) microRNAs code for shorter RNA transcripts which are processed into shorter, RNA hairpins (open hairpin displayed in pink). These hairpins associated with the multiprotein miRNA‐induced silencing complex (miRISC) in the cytoplasm where they guide the complex to their target mRNAs and block its translation or facilitate its degradation.
Figure 5. Expression patterns of Hox genes in the developing CNS of the mouse embryo. At E9.5, the patterns of Hox gene expression become restricted and localised to specific rhombomere (r) segments during the transient segmentation of the hindbrain. In general, they follow a two‐segmental pattern of expression where Hox2 expression commences in r3, Hox3 expression starts in r5 and Hox4 expression starts in r7. For each gene, their strongest domain of expression corresponds to their anterior domain and becomes progressively weaker in the posterior domains. In the developing motor neurons of the E12.5 spinal cord, Hox gene expression follows a collinear pattern of A/P distribution but becomes increasingly refined through negative cross‐regulation by more posteriorly expressed paralogues via the posterior prevalence rule. These discrete domains tend to coincide either with specific motor neuron columns (PMC, LMC, PGC and HMC) or within smaller motor neuron pools contained within these columns. For illustration purposes, the positioning of the motor neuron columns has been relaxed. Normally, they occupy a medial position within the basal plate of the neural tube.
Figure 6. Dorsoventral (D/V) expression domains within the CNS are unique and characteristic for each Hox complex. (a) Transverse schematic illustrating the neuroanatomy of the neural tube. Motor neurons project from the basal plate while sensory neurons will form synapses in the alar plates that transmit signals from the periphery to the medulla and cerebellum. (b) Hox genes display complex‐specific patterns of expression within the DV axis of the neural tube. The products of the HoxA and HoxC complexes regulate the patterning of the motor nerves and are predominantly expressed in the ventral portion of the neural tube where these nerves originate. In contrast, the products of the HoxB complex participate in the patterning of the sensory nerves and their expression is concentrated to the dorsal half of the neural tube from which sensory nerves form synpases. (c) The complex‐specific D/V patterns are established early during spinal cord maturation. In the case of Hoxb9, its initial, weak expression at E9.0 becomes concentrated in the dorsal half of the spinal cord at later stages.
Figure 7. Posterior genes from both the HoxA and HoxD complexes pattern the appendicular skeleton. (a) HoxD expression in the developing limb bud is regulated by two separate enhancer regions which are loosely organised within gene deserts flanking the complex. These enhancer regions correspond to topological associated domains (TADs; red pyramids) which represent chromatin interaction frequencies. In the E9.5 limb bud, the arm/forearm enhancers are more active, based on their association with Hoxd promoters, in driving early domains of HoxD expression. (b) In the E11.5 limb bud, there is a switch whereby the digit enhancers, within the opposing TAD, make more frequently contacts with the Hoxd promoters. This switch coincides with the expansion of HoxD expression in the progenitors of the digits. (c) The concentration of the HoxD transcripts in the developing digits is highly variable between the HoxD genes, with lower levels of Hoxd10 expression being observed. Hoxd13 displays the highest concentration of mRNA in the developing hand plate while Hoxd11 and Hoxd12 produce intermediate levels of mRNA. Hoxd13 is the only HoxD gene to be expressed in the presumptive thumb region.
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Further Reading

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Zakany J and Duboule D (2007) The role of Hox genes during vertebrate limb development. Current Opinion in Genetics & Development 17: 359–366.

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Nolte, Christof, Alexander, Tara B, and Krumlauf, Robb(Apr 2015) Mammalian Embryo: Hox Genes. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0000740.pub3]