|GENETICS HOME||GENETICS TABLE OF CONTENTS||OBL HOME||OBL REFERENCES|
|Homeobox genes code for a great variety of transcription factors in humans. The majority are involved in the differentiation of tissues in embryonic and fetal development, and as such they can be thought of as developmental “master switches”. One interesting group, the Hox cluster genes, determine the identity of cells along the longitudinal and limb axes of the body. These genes are not unique to humans: humans (and all coelomate animals) have merely modified common ancestral blueprints to form their bodies.|
|In the human body, human cheek cells not only possess all the genes they require to make cheek cell proteins, they also possess all the genes required to make the repertoire of proteins found in nerve cells. Liver cells have all the genes that cells of the ovary require. Bone cells have the same genes that are found in cardiac muscle cells. Since mitotic division faithfully copies the chromosomes of the parent cells and equally divides them between two daughter cells, all nucleated cells in the body (with some minor exceptions in lymphocytes) possess the same genes for the same proteins. Transcription factors in general determine which genes will be activated in each cell, thus inducing nerve cells to make a different set of proteins than are found in liver cells. Not only is the large family of homeodomain genes functional in this regard (as are other groups of transcription factors as well), many homeodomain genes function as “master switches” which, when activated in a cell during embryonic development, begin its differentiation to a specific fate. Some mutations of homeodomain genes can cause an anatomical structure to develop in an inappropriate region of the body. Although such mutations are best studied in flies, they can occur in other organisms as well (Kettle, 1999).|
|There are almost 2000 homeodomain transcription factors known from animals, plants, and fungi. Plants and Drosophila possess at least 70 homeodomain genes in their genomes while mammals possess at least 175. Many of genes and sequences are conserved between animals that are distantly related (Bharathan, 1997; Balavoine, 1995; Stock, 1996), such as C. elegans and humans, and even the regulators of Hox genes may be evolutionarily conserved (such as genes of the polycomb and homothorax group) (Kappen, 2000; Yu, 1998; Nunes, 2001). Yeast proteins which are homologous to the trithorax Hox family regulators in flies and nematodes function in the methylation of histone proteins (Nagy, 2002). Homeodomain proteins play vital roles in the embryonic development of both vertebrates and invertebrates and may have roles in simpler animals as well.|
The homeodomain proteins are transcription factors of the helix-turn-helix type that have a conserved DNA binding region made of about 60 conserved amino acids (the homeodomain). They can be clustered into two large groups, which evolved from an ancient duplication which predated the split between animals, plants, and fungi. The smaller family can be referred to as Group I (which share two characteristics in their sequences) and the larger family Group II. Both of these groups underwent additional duplications early in the evolution of eukaryotes. Group II includes clusters of Hox genes arranged in tandem which direct the development of the anterior-posterior axis in higher animals, which are referred to as Hox clusters. All of the following human genes in Group I are more similar to plant and fungal Group I genes than to any of the Group II genes (Bharathan, 1997).
Of the several hundred homeobox genes in animals, none possess a leucine zipper motif. In plants, such homeobox genes do exist, perhaps originating from the insertion of a leucine zipper exon into a homeobox gene. This ancestral gene underwent multiple duplications in the genome of the small mustard plant Arabidopsis thaliana to produce genes whose effects include the regulation of flowering, pigmentation, and leaf morphology (Schena, 1994; Schena, 1992).
1) GROUP I HOX GENES
a) SIX genes
--The SIX genes in mammals (mice and humans) have roles in the development of the eye and, since they are homologues of the sine oculis gene in Drosophila, are in involved in the development of invertebrate eyes. As a result, the SIX genes (and other genes including eyeless/PAX—another Hox gene) seem to be part of an ancient molecular pathway for eye development, suggesting that both protostomes and deuterostomes inherited similar mechanisms from an earlier bilateran ancestor with some form of eye.
Developing frog eyes are pictured below on either side of the developing brain (OMIM).
--six1 is involved in eye and limb development and is overexpressed in some tumors.
--six2 is expressed in the developing sclera, skeletal muscle, pancreas, and ovary.
--six3 is expressed in the developing retina.
--six4 is expressed in the developing brain, spinal cord, and sense organs in mice.
--six5 is expressed in the embryonic brain, meninges, adrenal glands, cornea, ciliary body, lens, and the photoreceptors of the retina.
--six6 is expressed in the embryonic eye, olfactory system, hypothalamus, and pituitary glands (OMIM).
b) TALE family
--These proteins all share a common a Three Amino acid Loop Extension and, in humans, include Meis1, Meis2, Pbx1, Pbx2, and Pbx3. Interestingly, both Pbx and Meis genes are involved in the limb development of both vertebrates and Drosophila. Although vertebrate and invertebrate limbs are not directly homologous, the last common ancestor of coelomates may have been a worm with some specialization of its lateral body wall (typical in many worms today) and the genetic mechanisms which defined this anatomical region may have been retained in descendants which evolved different types of limbs there (OMIM). Hox/Pbx and Brn proteins bind to the promoter of Pax3 genes to promote their transcription (Pruitt, 2004).
--Meis1 and Meis 2 are needed for the proximal/distal axis of the vertebrate limb. Meis1 is mutated in some mice with leukemia.
--Pbx is regulated by Meis 1. Mutations in Pbx 1 and 2 cause craniofacial and forearm abnormalities. Pbx 2 and 3 are expressed in most fetal tissues. One human Pbx genes is a pseudogene. One leukemia involved a fusion of the Pbx with the nearby E2A gene.
--PBX/knotted1/Homeobox1 is expressed in every tissue. knotted-1 is a family of plant HOX genes involved in plant development (OMIM).
c) TGIF (TG interacting factor)
--The TGIF protein interacts with MHC proteins. Mutations in TGIF can result in holoprosencephaly, a single central incisor, microcephaly, retardation, cleft palate, and the absence of the corpus callosum.
2) GROUP II HOX GENES
a) PAX Family
Paired box homeodomain genes share a large (128 amino acid) DNA binding domain and are most related to the Otx and Arx homeodomain families. The vertebrate Pax genes are homologs of paired box genes in Drosophila and all of them are essential for vertebrate embryonic tissue development. The Pax genes a form a family with 4 subgroups, descended from a common ancestral gene. A Pax gene is known in sponges and up to four Pax genes are known to exist in species of cnidarians. At least 2 of these are homologs of different Pax genes found in vertebrates, indicating that the diversification of Pax gene family members predates the evolution of bilaterans (Miller, 2000; Ogasawara, 2000).
Nine Pax genes have been identified from mammals and eight from Drosophila (Miller, 2000). Most Pax genes function in the development of the nervous system. Mice with mutations in Pax2 and Pax5 suffer a complete loss of the posterior midbrain and cerebellum (OMIM). The following image is of the cerebellar region of an embryonic pig brain.
Both vertebrates and insects have complex eyes, although these eyes are structurally very different. Simpler eyes exist in animals as diverse as tunicates, scallops, flatworms, and jellyfish. Are all animal eyes homologous? In one sense, they cannot be considered homologous since most of the structure of the vertebrate eye is specific to vertebrates and unlike the eyes found in insects and jellyfish. Nevertheless, it is possible that very different types of eyes (such as those found in vertebrates and insects) have descended from different sets of modifications of the same ancestral eyes. Although seemingly distinct, all animal eyes involve the expression of the gene Pax6. Null mutations of Pax6 genes can cause abnormalities and even the absence of eyes in animals as diverse as mice and fruit flies. (Ohno, S., from Muller, 1998). In Drosophila, ectopic expression of this gene can cause eyes to form on inappropriate body parts (including even limbs and genetalia). The effects of PAX6 mutations in humans include the almost complete absence of the iris, cataract, and other visual problems. PAX6 is known to determine the expression of zeta-crystallins in the eye (OMIM).
Pax gene family members initiate the development of eyes in coelomates and are involved in cnidarian eye development as well (Kozmik, 2005).Cnidarians are the simplest animals to possess eyes and many possess Pax genes. One of these cnidarian genes, Pax-B, is related to the vertebrate genes Pax-4/6 and Pax-2/5/8 (Sun, 1997). Pax6 activates the expression of vertebrate crystallin genes and the homologous jellyfish PaxB gene activates jellyfish crystallin genes (Kozmik, 2003). Eyes are found in some species of Hydra which possess a single Pax gene. The simplest cnidarians (anthozoans) can possess at least four genes. One intron has been preserved in coral, invertebrate, and vertebrate Pax genes (Miller, 2000).
Nemertine worms, which may represent a lineage whose origin was close to the protostome-deuterostome divergence, express a Pax-6 homolog in their CNS and eye regions (Loosli, 1996). Tunicates express Pax genes in their eyes, as do vertebrates and fruit flies. Ectopic expression of Pax genes can cause the formation of additional eyes (Glardon, 1997). The expression pattern of Pax6 in the embryonic development of the nervous system has been highly conserved among osteichthyans (bony fish and tetrapods). Some aspects of its expression are also observed in cartilaginous fish and jawless fish, indicating that it was functional in ancestral vertebrates (Derobert, 2002).
The following image is of the developing brain and eye of an embryonic pig.
Pax 3, 7, and 6 expression is involved in the subdivision of neural tube and Pax2, 5, and 8 in midbrain-hindbrain boundary (Ogasawara, 2000). In mice Pax 7 is expressed in rhombomeres 1, 3, and 5 while Pax 3 is expressed in rhombomeres 2 and 4 (Pruitt, 2004).
One of the defining features of chordates is the development of pharyngeal arches, such as those of the pig embryo below.
Pax 1 and 9 are expressed in pharyngeal arches of vertebrate embryos. A single gene Pax1/9 homolog is expressed in the pharyngeal arches of urochordates, cephalochordates, and hemichordates. Homologs of this gene are known in more primitive animals which lack pharyngeal arches, such as sea urchins, flies and nematodes (Ogasawara, 2000). Two genome duplication events seem to have occurred early in vertebrate evolution, as evidenced by the multiple copies of genes in vertebrates whose homologs exist as single copy genes in primitive chordates and other invertebrates. The duplication of the Pax 1/9 gene occurred before the evolution of lampreys and the expression pattern of Pax9 in lampreys is intermediate between that of chordates and gnathostomes. (Ogasawara, 2000)
--PAX1 is important in the development of the vertebral column, sternum, and scapula.
--Mutations in PAX2 result in kidney and eye problems.
--Mutations in PAX3 cause Waardenburg syndrome whose symptoms include deafness and the absence of melanocytes in the skin, hair and eyes.
--PAX4 is involved in the differentiation of the endocrine tissue of the pancreas.
--PAX5 is involved in hematopoeisis.
--Although PAX6 is also involved in the differentiation of the endocrine tissue of the pancreas, it is best known for its role in the development of the eye. PAX6 is the homolog of the Drosophila gene eyeless and is a master gene controlling eye development in vertebrates, insects, cephalopods, ascidians, and nemertians. The reduced eye size observed in the fly on the right is caused by a mutation in a Pax gene (eyeless).
--PAX7 is expressed in satellite cells derived from myoblasts.
--PAX8 is necessary in the thyroid for the development of the thyroxine-developing follicular cells. A fusion gene of PAX8/PPARG1 was involved in a carcinoma.
--In mice, mutations in PAX9 cause symptoms which include cleft palate, abnormal face and limb development, the absence of all teeth, and the absence of a thymus, parathyroid glands, and ultimobranchial bodies (OMIM).
b) POU Family
--The genes of the POU family are important in cell differentiation. POU domain pdm exist in arthropods where they are expressed in wings and epipodytes. Deuterostome share Oct gene members of this family (Averof, 1997a).
--POU1F1 is only expressed in the developing pituitary and hormone secreting cells. Mutations in this gene cause deficiencies in the secretions of pituitary hormones.
--POU2AF1 binds Oct proteins. Although B cells can develop without this gene, there are fewer B cells than normal in mutants.
--The POU Class 3 genes (4 members known in humans) evolved from a common ancestral gene around the time of the origin of the vertebrates. POU3F2 and POU3F3 are specific to the central nervous system. POU3F4 is expressed in the developing brain, neural tube, and otic vesicles; mutations can result in deafness.
--POU5F1 is expressed in oocytes before and after fertilization.
--Oct3 is expressed in mouse oocytes before and after fertilization. In mutant mice, inner mass cells are not pluripotent. Oct3 seems to be a master regulator of pluripotency needed for stem cell self-renewal (OMIM).
c) paired-like Homeobox Family (paralogs of the gene paired in Drosophila)
This family of genes is known in both protostomes and deuterostomes). In vertebrates, gene members are involved in craniofacial and eye development in vertebrates and include the PITX Subfamily. The tunicate Pitx (pituitary homeobox) homolog is expressed in the neural complex (part of the embryonic pharynx) comparable to where the pituitary forms in vertebrates. The Pitx family of homeodomain proteins is related to the Aristaless-related, goosecoid, and Otx families (Christiaen, 2002). Pitx is required for the development of dopaminergic neurons in the substantia nigra of the midbrain (Nunes, 2003).
--CART1—Mice with mutations in CART1 die soon after birth with acrania and meroanencephaly.
--Rx is expressed during the development of the retina.
--Pmx is expressed in the developing cochlea and mutations can result in deafness.
--VSX1 is expressed in the eye.
--PITX1 is expressed in the pituitary and hindlimb. If PITX1 is expressed in a developing chicken’s wing, it develops as a hind limb. The following image is of the developing hindlimb of a chick (OMIM).
--PITX2 is induced by the important Sonic Hedgehog protein. It is expressed in the left side of the heart and gut in the embryo and may have a role in determining left-right asymmetry. Mutations cause abnormalities of the jaws, teeth, pituitary, and heart and can cause Rieger Syndrome.
--PITX3 is expressed in the developing lens. Mutations in mice result in small eyes without lenses; mutations in humans result in cataracts and other ocular problems.
c) LIM Homeobox Family
LIM Homeobox proteins have one homeobox domain and 2 LIM domains. There are more than 40 members of the LIM family known in vertebrates and invertebrates but not all of them are HOX genes. Humans possess at least four LIM-only proteins (LMOs) which lack the homeodomain.
apterous is a LIM-homeodomain gene in Drosophila. It functions in the development of the wing and nervous system and contributes to viability and fertility. Not only do vertebrates (such as humans and mice) possess this gene , the vertebrate proteins can perform the same function in flies as the fly proteins, given that human LHX2 genes can restore normal phenotypes to apterous mutants in Drosophila. In both mice and fruit flies, apterous is expressed in the nerve chords, eyes, brain, limbs, and olfactory structures. (Rincon-Limas, 1999)
Other LIM proteins have a variety of roles. In vertebrates, LIM genes are expressed in nerve cords, eyes, the brain, olfactory regions, and limbs. Xlim-1 is expressed at embryonic organizer regions and is involved in establishing the body axis and induces important embryonic genes goosecoid, Xotx2, and chordin. Lim1 mutations in mice cause the loss of head structures anterior to the third rhombomere (Yamamoto, 2003). Lmx1b is involved in the development of midbrain dopaminergic neurons (Nunes, 2003). Many developing cholinergic neurons require the LIM homeobox Lhx8 (Zhao, 2003). In C. elegans, lin-11 involved in the formation of vulva and mec-3 functions in the formation of mechanosensory neurons. In Drosophila, islet and arrowhead involved in development of neurons and imaginal disks respectively. In vertebrates, Lhx1 functions in the differentiation of mesoderm, Lhx3 in that of the pituitary, Isl1 for motor neurons, and Lmx1 the dorso-ventral limb axis (Rincon-Limas, 1999).
--LMX1A is expressed in the CNS, especially in dorsal cells.
--LMX1B causes the nail-patella syndrome which, among other abnormalities, can result in the absence of nails and patella bones.
--LHX1 is expressed in the brain, thymus, tonsils, lymphoid cells, and motor axons of the limbs.
--LHX2 is the homolog of apterous in Drosophila. It is expressed in the developing and adult CNS and is also expressed in B and T cells. Mutations have been involved in leukemia.
--LHX3 mutations cause deficiencies of pituitary hormones and motor neuron abnormalities.
--LHX4 is expressed in the pituitary and, for a brief period, LHX3 and 4 are expressed in all motor neurons which travel ventrally. Mutations cause shortness of stature, pituitary and cerebellar abnormalities, and a small sella turcica.
--LHX5 is expressed in the developing forebrain; the brain of an embryonic chick is pictured below.
--LHX8 is expressed in the secondary palate; mutations can cause cleft palate.
--Mutations in LHX9 in mice result in the absence of gonads.
--ISL1, when mutated, can cause the agenesis of the pancreas and diabetes.
d) Bar-like genes (homologs of Bar in Drosophila)
--Barx1 and 2 are expressed in the developing head, brain, jaws, neck, and stomach. Some mutations cause Rieger syndrome and eye abnormalities (OMIM). The Bar mutation in Drosophila is pictured below.
e) One-Cut genes (homologs of cut in Drosophila)
--One-cut 1 is expressed in the liver, testis, and skin. Mutations in mice cause endocrine problems of the pancreas.
--One-cut 2 is expressed in the liver, skin, testes, and bladder.
--Cut-like 1 is downregulated in a number of tumors.
--Goosecoid is the homolog of the gene bicoid in Drosophila which is one of the first genes which helps to determine pattern in the embryo. Goosecoid is expressed during gastrulation. Mutations in mice cause abnormalities in a variety of tissues and mutant mice die at birth.
--Goosecoid-like is expressed in both embryonic and adult brain. Mutations can cause DiGeorge syndrome, velocardiofacial syndrome, learning disorders,and psychiatric illness.
j) ALX (homolog of arista-less in Drosophila)
--ALX3 is expressed in the developing head, pharyngeal arches, tail, limbs, and genitalia. The following image is of the pharyngeal arches of a chick embryo.
--Mutations in ALX4 cause the formation of parietal foramina.
k) OTP (homolog of orthopedia in Drosophila)
--OTP is involved in the migration of neurons; mutations can result neural abnormalities including problems of the hypothalamus.
l) DRK11 is expressed in sensory neurons; mutant mice have reduced perception of pain.
m) LBX (homolog of forkhead in Drosophila) is expressed in muscle precursor cells of the limb.
n) Prox1is expressed in the CNS and eye and mutations cause the development of a smaller liver and the abnormal development of lymphatic vessels.
--HESX1is active during gastrulation and has a role in the development of the head. Mutations cause abnormalities of the corpus callosum, hippocampus, and septum pellucidium (the latter can even be absent).
p) Pem is expressed in the placenta, gonads, and epidermis of mice and it is involved in the regulation of spermatogenesis.
q) Mutations in ZFHX1B cause Hirschsprung disease with its mental retardation, epilepsy, and delayed motor development.
r) HHEX is expressed during hematopoeisis and in the liver.
--In humans and mice, this gene is expressed in the fetal and adult retina. Mutations cause micropthalmia, cataracts, blindness, and iris abnormalities (even the absence of a pupil).
--This gene is a homolog of Antennapedia in Drosophila. It is expressed in the developing thymus, blood cell precursors, and activated lymphocytes.
--This gene is the homolog of ceh in C. elegans. It is retina specific and its mutation leads to eye abnormalities.
The sponge homeobox gene EmH-3 is homologous to Hox 11 in higher animals. This gene in mammals and Drosophila is a member of the Tlx group of the NKL/93DE Hox cluster; sponges also possess 2 genes (ox3 and Prox1) which are homologous to two other members of this cluster (sx/sh and Nk3/bap). Human cells in culture activated introduced sponge genes in a way similar to the mammalian homolog, suggesting that the gene’s function in determining cell growth and differentiation was established in early animals (Coutinho, 2003). When mutated, HOX11 can cause leukemia (and thus was the first HOX gene to be linked to human cancer) and the absence of a spleen in mice (OMIM). Cells of the spleen are depicted below.
aa) SHOX--Short Stature HOX
This gene is expressed in bone-making cells, in limbs, and in the first two pharyngeal arches. Although it is on the X chromosome, it is in an area called the pseudoautosomal region that behaves like the autosomes rather than in a sex linked fashion (that is, males have a copy on the Y chromosome and females express both copies, even though one of the chromosomes has condensed to form a Barr body).
Individuals who have an extra copy are taller (although women with this condition are estrogen deficient). Mutations in this gene cause short stature and abnormalities of the palate, teeth, and ears. These same problems are seen in women who have Turner’s syndrome, caused by the presence of only one X chromosome. It may be that many of the characteristics of missing one X chromosome are the result of only one copy of SHOX rather than two (OMIM).
Orothodenticle genes (otd in flies and Otx 1-2 in vertebrates) contain a bicoid-like homeodomain and form the anterior brain. Otx2 mutants lack anterior neurectoderm. Although there are obvious differences between the brains of vertebrates and invertebrates, comparative analysis of gene expression indicates that certain shared patterning was established before the last common ancestor of coelomates. (Boyl, 2001). Hox cluster genes (discussed in the next section) are expressed in along the developing longitudinal axis of the posterior brain in both protostomes and deuterostomes. Otd/Otx genes are expressed in the anterior brain in both groups. Both protostomes and deuterostomes possess a rostral brain which is separated into regions. Both groups use homologs of ems/Emx and otd/Otx in the formation of separate brain regions. Pax and Six genes are also expressed in the developing brains and eyes in both groups. (Kammermeier, 2001; Tallafuβ, 2002). Otx/Otd homeodomain genes are known in eumetazoan animals and is an important regulatory element of the bilateran brain. Fly mutants can be rescued with expression of mammalian proteins and many of the neural mutant phenotypes of mammalian mutants can be rescued with the expression of fly proteins. The single ancestral gene in ancestral chordates was duplicated by the evolution of the jawed fish at which point new roles in the development of the brain were established (Acampora, 2005).Otx is expressed in the photoreceptor structures of lancelets and in the developing eyes of jawless fish. Gnathostomes possess multiple Otx genes which guide the differentiation of eye structures, such as bipolar cells and photoreceptors (Ploughinec, 2005).
Hox proteins can bind DNA as a heterodimer with another homeodomain protein, extradenticle (Exd). This binding affects the specificity of the Hox protein and its ability to promote transcription. The homothorax homeodomain can interact with Exd to determine its nuclear localization (Sprecher, 2004).
The homeobox gene bozozok and the TGF-β gene squint are expressed in the dorsal yolk region and induce the BMP antagonists which are required for the formation of neural tissue (Sirotkin, 2000; Fekany-Lee, 2000)
Fish embryos express the homeobox protein nieuwold/dharma in the yolk prior to gastrulation. Mutations here can disrupt or delete structures such as the notochord and prechordal plate and result in the loss of neural cell fates at the midbrain/hindbrain boundary (Koos, 1999).
Two species of Drosophila, D. mauritania and D. simulans, have involved changes in an orphan Hox gene, Odysseus (Ods) as a “speciation gene” which is responsible for the sterility of males produced by matings between D. mauritania and D. simulans (Ting, 1998).
Anthropoid primates have experienced a higher level of evolutionary change in the gene TGIFLX, a homeobox gene expressed only in the testis (Wang, 2004)
Anthropoid primates have experienced a higher level of evolutionary change in the gene TGIFLX, a homeobox gene expressed only in the testis (Wang, 2004)