Colinearity of Hox Genes

The word "homeotic" was invented by William Bateson (1861-1926), the famous early geneticist, to mean substitution of one structure at the normal location of a serially-homologous structure, such as an insect forming an extra pair of legs where the antennae should be ("antennapedia") or a mammal forming an extra pair of canine teeth from the tissue that normally develops into pre-molar teeth, or forming an extra pair of ribs. Incidentally, this is the same Bateson who invented the word "Genetics", and was one of the 4 re-discoverers of Mendel's research papers. One of the books Bateson wrote ("Materials for the study of variation...") amounts to a catalog of birth defects, most of which turned out not to be caused by mutations. It has been reprinted, and I own a copy. He was seeking examples of discontinuous variation, because most scientists then assumed that big jumps in phenotype would be needed to produce different species. Actually, it is still a topic of active research how new species branch off from existing species.

Several homeotic mutations were later discovered in Drosophila. The two most famous were "Antennapedia" (which causes legs to form from the imaginal disks that normally become antennae), and "Bithorax" (which causes an extra pair of wings to form from the imaginal discs that normally become much smaller structures called "haltares". At least 8 such mutations were discovered in flies.

Notice that the Wikipedia article about homeotic genes says that they "cause the development" of specific organs, i.e., rather than controlling which organ will form from specific tissues. This is an example of how difficult it is to think logically about how genes cause anatomical structures. The relation between base sequences and amino-acid sequences is understood. Before that, Beadle and Tatum deduced the causal relation between genes and enzymes. Some examples are known of mutations that alter self-assembly of proteins; but beyond that scientists know very little about how genes affect larger-scale anatomical structure, although hundreds of mutations have been found that cause abnormal anatomy. Many were discovered in flies by studying the anatomy of larvae that died because they were homozygous for particular genes. These genes were given whimsical names, based on whatever the abnormal embryos looked like: hedgehog, armadillo, etc. Nearly all of them code for transcription factors, i.e. proteins whose function (or one of whose functions, in the case of armadillo) is to bind selectively to specific DNA sequences, and either stimulate or inhibit transcription of other genes.

Would you say that buttons "cause" elevators to move to different floors of buildings? Cables and pistons also have something to do with movements of elevators. Which would you say are more fundamental? The buttons, or the cables and pistons? Do buttons activate the genes for being in a different altitude? Choice of vocabulary often limits scientific progress. The words you choose will limit your concepts. This aspect of the philosophy of science doesn't get nearly enough attention. The best way to study it is to read early scientific papers on topics about which major breakthroughs have since been made. (For example, "The Dreaded Erroneous Paper Report", that you should now be thinking about.)

Incidentally, the peculiar flower structure of cauliflower and broccoli (the parts that you eat) result from mutations that cause each petal, stamen, pistil etc. of their flowers to try to develop into an entire flower (all the petals, etc. of which then try to become whole flowers), and this goes on and on.

Do you see the analogy to homeotic mutations in animals? Imagine a mutation that caused each finger to develop into a whole hand, each finger of which also tried to become a whole hand, except that each of the fingers of these extra hands developed into a whole hand, etc. without limit.

Although no animal genes have been found that produce such branching hands, the normal development of lung alveoli results from epithelial out-foldings, each of which branch into two outfoldings, with both of these themselves branching into two, and all four of these branching into two more. 2, 4, 8, 16, 32, 64, 128 etc. up to about a million alveoli per lung in humans.

The branching structures of salivary glands, the pancreas, and kidneys, also develop by sequential splitting of out-foldings into two, which themselves branch into two, etc. The resulting structures are examples of what mathematicians call "fractals", because the component parts are scale models of the whole, which results in being considered to have two and a half dimensions, more or less. Fractional dimensions are less important than the parts being miniatures of the whole.

A Professor of Biology at Cal. Tech. named Edward Lewis (1918-2004) did the most important research on homeotic genes in fruit flies. Among his discoveries was that these genes are located in two closely-linked bunches, and that the relative locations of the homeotic genes along their chromosomes matched the relative anterior-posterior locations of the anatomical positions of the abnormalities produced by mutation of the different homeotic mutations. This correspondence of relative gene locations, relative to anatomical locations of abnormalities is called co-linearity, and is unique to homeotic mutations (as far as I know).

I learned many interesting facts from the biographical web site
(, one of which was that the first chromosome regions to be completely sequenced were those where homeotic genes are concentrated in Drosophila. Another was that deletion of these whole sets of genes resulted in development of flies that looked like centipedes, in the sense that all their segments had the structure of the most posterior segment.

From the base sequences, it was soon realized that the homeotic genes all code for transcription factors, and that their binding sites have almost identical amino-acid sequences, 61 amino-acids long. Very similar amino-acid sequences were soon discovered in other transcription factors, including many that are coded by genes that are not homeotic. The famous "bicoid" gene is an example: it contains a "Hox Box", but mutations produce two tail ends. Or do you want to regard that as a homeotic phenotype? Logically, you could argue in favor of that; a second tail develops from the tissue that normally forms the head.

Every multicellular animal whose DNA was sequenced turned out to contain many (or at least some) genes that coded for almost this same "Hox" amino-acid sequence. Many of these were in closely-linked bunches. Humans and mice have four such bunches, called A, B, C and D (each on a different chromosome). The individual "hox genes" are called A1, A2, A3..., and B1, B2, B3, and D1, D2, D3 up to D13, although some of them are missing, in the sense that the similarities of amino-acid sequences are so extremely similar, for example between all the number fours. The term parologous groups refers to those assigned the same number. Orthologous groups refers to those with the same letter (unless I got it backwards; I am writing this from memory, and eventually will find out for sure.)

In mammals and fish, colinearity of hox gene expression includes (at least) two phenomena. A correct theory ought to predict both (and probably a third or fourth aspect).

One: "Spatial Colinearity): The anterior borders of where each hox gene is expressed (i.e. where it is transcribed) is farther forward for the lower-numbered hox genes.

Two: "Temporal Colinearity": The lower numbered hox genes are transcribed sooner during development.

Three (Please invent a good name): Deletions and other mutations in the lower-numbered hox genes tend to produce structural abnormalities toward the front of the animal.

Theories that scientists have proposed already:

* Very long range enhancer regions, located near the low-numbered end of the hox gene groups, that for some reason act at short ranges early, and in anterior tissues.

* Some gradual removal or loosening of histone binding, first at the low numbered hox genes.

* A diffusion gradient of retinoic acid, down the length of the body, with the hox genes differing in how much they are stimulated by retinoic acid (which doesn't require the hox genes to be adjacent)

* Control by the "clock and wave-front" mechanism that controls somite formation.

* Mechanical tension on the chromosome changes hox gene expression (I had nothing to do with this theory, despite my research on tensions.)

* (This one I invented) Each cell can (somehow!) detect which hox genes are being transcribed in adjacent cells, and compares them with which hox genes that cell itself is transcribing, and responds to large differences by activating transcription in the hox genes with intermediate numbers.

Lest this one seem too crazy, it has long been known, but unexplained, that if you graft embryonic tissues to locations next to tissues that are normally distant from them, both tissues respond by increased growth and mitotic divisions, and by changing their development to be more like the cells that should have been in-between.

This is equivalent to intercalary regeneration (As a student in the course asked).

But by what criteria do cells distinguish whether other cells "Should" be next to them? Nobody knows. If their spatial criterion were which hox genes their adjacent cells transcribed, and whether its hox genes were next to your hox genes (heaven knows how!), then that would kill two birds with one stone. (i.e. would "explain" how cells decide if they are "supposed to be" next to particular cells; and would also "explain" colinearity).

A major question for every theory is whether and how cells can detect which hox gene proteins (or mRNA) are present in the cells next to them, and how they could compare them with whichever hox genes are transcribed in adjacent cells.

An effort to test this will be described in class. The key ideas were

(A) to dissect cells out of anterior and posterior parts of chick embryos.

(B) To culture and subculture these cells, using in situ hybridization to test if they were transcribing hox genes corresponding to where on the chicken embryo the parent cells had been dissected from.

(C) You figure out what to do next


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