X. The Homeobox
Through a variety of experiments, developmental biologists are beginning to learn how the one-dimensional information encoded in the nucleotide sequence of a zygote’s DNA is translated into the three-dimensional form of an animal. There are enough unanswered questions, however, to entertain many future generations of scientists.
Neil Campbell, Biology (4th ed., 1996)
Neil Campbell, Biology (4th ed., 1996)
As the 21st century approached, biologists—despite all their remarkable findings—still knew very little about the specific role of genes in embryonic development. During the 1970s and 80s, teams of researchers working on different problems were beginning to understand that regulatory enzymes and genes were responsible for various aspects of the initial growth phases, including their coordination and timing.
This culminated in 1983 with the concurrent discovery (by three separate labs on two continents) of a cluster, or complex, of regulatory genes responsible for the orchestration of early developmental stages—the ontogeny—of organisms exhibiting segmented body parts.
Inducing mutations in these types of creatures by manipulating DNA has, since Morgan’s time, been an invaluable research tool—the key to identifying things such as genes’ specific locations on chromosomes and various of their attributes (for instance, their roles in gene networks, or whether they were dominant or recessive). The discoveries of the 1980s revealed that various deformities seen in fruit flies—such as extra sets of wings or legs gruesomely sprouting from heads—were the result of mutations to groups of genes controlling the fly’s overall body plan. After years of study, it was found that there were eight of these (what are now known as) homeotic genes in two clusters on one of Drosophila’s chromosomes. Significantly, the relative position of the genes in these complexes were found to correspond to the relative head-to-tail order of those body parts affected. Consequently, it was anticipated that these clusters held clues to the general logic behind the fly’s body patterning.
When DNA from the two gene clusters was isolated and analyzed, it was found that proteins coded for by each of the eight homeotic genes had short sections amounting to 180 nucleotides sharing virtually identical sequences. Researchers dubbed them homeobox genes. The abbreviated form, Hox, is still used in reference to regulatory genes involved with segmentation and related structures but the term “homeobox” now covers a wide range of genes central to an organism’s development and growth.
Homeobox genes are part of what is commonly referred to as the genetic tool kit, which metaphorically consists of a number of “tools.” Many of these are transcription factors (as mentioned earlier, DNA binding proteins that switch genes on or off), plus signaling proteins, hormones, and various cellular receptors. Others include cell-type proteins (those that determine a cell’s identity) and coloration proteins. Homeobox genes control ontogeny by sending signals between each other in the form of chemical messengers—gene products—whose transcription they induce.
Hox genes, again, are identity genes—those responsible for an embryo’s head-to-tail development. Some delineate the right and left sides of the body; others establish top and bottom while still others map out general body regions. Hox genes are responsible for switching on circuit cascades of master genes such as the homeotic genes that are in charge of building structures associated with an embryo’s different segments. Homeobox genes, of course, play multiple roles and virtually never work in isolation.
Here’s an outline of how this process plays out in the case of that superstar of lab animals, Drosophila melanogaster:
Like all arthropods, fruit flies are modular in construction, their bodies arranged in three sections—head, thorax, and abdomen—which are further subdivided. (For example: the first thoracic segment bears a pair of legs; the second, another set of legs plus a pair of wings; and a third carrying the last two legs and a pair of modified wings—balancing organs called halteres.) Starting with an undifferentiated egg, segments appear and these become ever more anatomically distinct as development proceeds in a series of steps determined by gene activation. Regional identity information is established by the types and, importantly, concentrations of regulatory proteins that control gene expression. This begins before eggs are even fertilized.
Egg-polarity genes active in nursery cells surrounding and nourishing unfertilized eggs in a female fruit fly’s ovary begin the process of defining an embryo’s body axes. When transcribed, these genes’ mRNA enters the egg cell, remaining near the point of origin. After being fertilized and laid, the mRNA is translated and its DNA-binding protein product begins to slowly diffuse though the egg away from its point of origin, forming a concentration gradient. In the instance of the polarity gene responsible for establishing anterior-posterior regions in the fertilized egg, its highest concentration—near the point where it was introduced—delineates the head end while areas of decreasing concentration sequentially activate segmentation genes that map out the basic segmentation plan along the anterior-posterior axis. The dorsal-ventral axis is formed in similar fashion.
After the major body axes are laid out, three suites of segmentation genes go to work. Gap genes determine, in succession, the basic subdivisions. Next, pair-rule genes establish pairs of body segments. Finally, segment-polarity genes set up the front-to-back axis of both halves of the paired segments. At this point, the boundaries and orientation of each of the embryo’s segments are permanently established. The segmentation genes’ protein products, for the most part, are transcription factors that advance the step-wise system of body plan formation; in turn, they activate sets of genes that activate other sets, ending with the homeotic genes that determine the specific anatomy of the embryo’s individual segments.
The sum total of an organism’s development is in the delicately balanced timing and sequence of these master genes being switched on and off, and the duration of their activation. Molecular biologist Sean Carroll, one of the early leaders in the field of evodevo, provides glimpses of the remarkably sophisticated logic employed—yet another exemplary instance of Natural Design on display:
A gene not only may have multiple switches for different subpatterns of expression at a given time, but will frequently have different switches that control entirely different states in development. Tool kit genes are rarely, if ever, devoted to a single developmental operation. Rather, these tools are used and reused again and again in development in different contexts to shape the growing embryo. Switches endow individual tool kit genes with great versatility. Virtually every tool kit gene is controlled by multiple switches. Ten switches or more is not uncommon, and we don’t know what the upper limit, if any, may be…. The developmental steps executed by individual switches and proteins are connected to those of other genes and proteins. Larger sets of interconnected switches and proteins form local “circuits” that are part of still larger “networks” that govern the development of complex structures. Animal architecture is a product of genetic regulatory network architecture.
Researchers involved with the Drosophila studies that resulted in the initial isolation and elucidation of Hox genes were thrilled by their discoveries. Curious to see if other organisms might possess something analogous, they were shocked to find Hox genes in other animals—barnacles, worms, fish, frogs, mice…and humans. They found that these genes were positioned on chromosomes in the same order as in the fly. Researchers were astonished to realize that these remarkable genes performed the same jobs in each case: specifying the developmental fate of each embryonic segment in the same head-to-tail order. And it was ascertained that in each organism, parts of the Hox gene sequences were virtually identical.
In one famous experiment performed in the 1990s by Swiss developmental biologist Walter Gehring and his colleagues in Basel, the Hox gene responsible for overseeing the construction of a fruit fly’s eyes was manipulated so as to be expressed in different parts of the fly’s body. In this way, eyes were induced to grow on different flies’ legs, wings, and antennae. Gehring’s team then repeated the experiment on Drosophila but used a mouse’s eye-forming gene instead; remarkably, these flies were able to grow normal eyes. In a similar experiment, the eye-forming gene in a frog was replaced by the analogous gene in Drosophila and the frog was also able to grow completely normal eyes. This means that a single gene product is able to function as a switch that turns on eye development in a diverse array of animals.
As a result of these amazing experiments, it now appears that all types of eyes arose from a common ancestor and that the genes involved are highly conserved and have been virtually the same for hundreds of millions of years. Until this discovery, it had been assumed that vision had arisen independently, scores of times, in creatures as varied as echinoderms, mollusks, and vertebrates. Apparently, though, the genetic tool kit was fully emplaced in ancient organisms—distant relatives of all animals alive today—and that the extremely rapid radiation of forms during the Cambrian period was merely awaiting favorable ecological conditions that would “light the fuse.”
©2016 Tim Forsell
10 Apr 2016
 Campbell. p 990. (A zygote is a just-fertilized egg cell.)
 The breakthrough —another classic case of the phenomenon of multiple discovery (see Shubin, pp 182-83)—was made independently by researchers at the University of Basel, Switzerland; at Stanford in California, and Indiana University.
 Confusingly, the root “homeo” refers to the sequences’ similarity—not because they’re part of homeotic genes. And “box” is microbiologist slang for a sequence of identical (or nearly identical) nucleotide bases found within multiple genes.
 Carroll (2005) . pp 123, 129.