The genetics of olfaction: In evolution, where there's one, or one thousand....there's more. But how much more? Part II.

Last Friday and today we're discussing the means by which you are able to detect and discriminate among infecting pathogens to which you may be exposed.  We reviewed the immune system and the idea of monoallelic expression, by which only one of the two copies of a gene you inherit is chosen for use in a given immune cell.  In fact, in the immune system, it's really only part of a cluster of gene segments that is used, and only one cluster of the two in your inherited genome.  It's called 'adaptive' immunity (you also have a second, very different, 'innate' immune defense unrelated to our subject here).

We went on to say that odor detection is a somewhat similar challenge:  of the countless odor molecules that you might want or need to be able to detect, your body can't know all in advance.  Since odorants are molecules, perhaps a molecular--a genetic--method of open-ended variation would serve as well as it does in our 'adaptive' immune system. In fact, we have something similar in our odor-detection sense.  But it works very differently from the immune system, using unrelated genes and unrelated (as far as is known) gene-selection mechanisms.

We have about 1,000 (yes, that's thousand, about 5% of our genome) different Olfactory Receptor (OR) genes in our genomes. (We've posted on aspects of this story in past years, e.g., here and here.)  Since we have two copies of all our genes (except for XY genes in males), that means we have a total of nearly 2,000 OR genes.  An OR gene codes for a protein that sits in the surface of olfactory neurons, hanging out in the nasal passage, in contact with the air we breathe, and the odorant molecules it carries.  The OR genes are in clusters, large and small arrays of adjacent, related OR coding genes, scattered across most of our chromosomes.  Each OR gene is the result of a past duplication event, so is closely related to some other OR's often right nearby on the same chromosome; 'closely' means a few mutations different from the 'parent' gene that have arisen as part of the duplication or since that occurred.

OR locations in the genome (red); obtained from http://www.leffingwell.com/olfact2.htm

In some mammals that rely heavily on smell, most of the OR genes, in most clusters, are working.  In primates and humans, we've still got as many identifiable OR genes but more of them are dysfunctional, having been mutated out of usability; the protein code just doesn't generate a functional OR protein.  But for many or most, that means that ORs that may be dysfunctional in one of my copies may be functional in the other, or in other people.

Olfactory neurons develop as part of the nasal cell lining.  They express OR genes on their surface, that dangle into the nasal airway.  But as a future olfactory neuron develops, it picks one OR gene, from one cluster, and of that only from one of the two copies of that cluster, to express.  The other copy is inactivated as are all the other 1,999 OR genes!  But this is not so easy to explain as the comparable monoallelic expression of the antibody genes that we described on Friday.

Source

The two copies of them, at least, are in the corresponding place on the two copies of the respective chromosomes (one inherited from each parent).  That makes them in principle easier to be put into correspondence by which one is chosen and the other inactivated.

In the case of the OR genes, the developing olfactory neuron must first pick one of the many clusters to use, and then use only one OR gene from that cluster, and then inactivate the other genes in all other clusters, both copies, on all other chromosomes!  Pick one gene, and then go find all the others and silence them.

This is a remarkable feat and the very explicable outcome is that each olfactory neuron uses only one OR protein, and can detect only odorants that, wafting by, are recognized by that OR protein and trigger the neuron to send an "Aha!"  message to the brain. The brain collates the messages sent at any given time by all the neurons in both nostrils, and assembles and remembers a catalogue: "These neurons respond together and we'll call that 'lemon';  next time this same collection of neurons fires, I'll know to get out the glass, water and ice cubes--lemonade!"

Because they are being duplicated rapidly and they mutate rapidly, the 2,000 OR genes provide a huge repertoire of potential responders, and their combinatorial signals, which provides the kind of open-ended cataloging of odorants--including mates, predators, food and prey.  This is an extensive form of monoallelic expression that seems obviously to have been useful to our ancestors' survival.

Current status
In 2005 I spent a sabbatical leave at the Sanger Genome Center near Cambridge, England, trying to find DNA sequence motifs that might help account for this very selective monoallelic expression.  I didn't succeed....but neither has anyone else since.  A recent review by Ivan Rodriguez in Cell describes what is known about this system, and an article in the same journal describes one facet of that mechanism.

Rodriguez notes that some cluster-specific regulatory regions have been found on some chromosomes, and there are instances of those having similar sequences; but they are far down the chromosome from the OR genes themselves, they only affect their local cluster, which makes such regulatory sequence elements hard to find on other clusters.   Previous ideas of global all-cluster control sequence signals have not panned out.  But a mechanism has been found that, in part at least, explains how a cell that happens at first to pick a defunct OR gene for use fails to develop further but instead is returned to the OR-picking stage.  It's not known what happens if a cell mistakenly picks more than one OR gene, but such cells are rarely seen so they must degrade somehow, or fail to reach the relevant part of the brain (the olfactory bulb).  The part of the DNA including a chosen cluster region is open for expression, but how the rest of the OR-cluster areas on the other chromosomes are silenced isn't known.

Other related systems have similar one-gene selection. These include the V1R and V2R genes that code for pheromone receptors and two other groups, the FPR and TAAR genes, that are each expressed in a one-only way but in a different part of the nose, called the vomeronasal area. 

An older idea that the cluster-containing parts of chromosomes were located near the outside of the neuron cell's nuclear membrane was shown to be wrong; instead these areas may be clustered toward the center of the nucleus.  This may bring various OR regions closer together so they can be activated or shut down.....but how are these areas shepherded there?  And since an olfactory neuron is also expressed in many hundreds of other genes, how are they kept open for expression?

This is considerable advancement in our understanding of how the chosen OR gene leads to a functional neuron or how a dysfunctional OR choice leads to a re-selection. It explains some aspects of how an OR cluster's genes are opened for expression, and how the coded protein is processed outside the cell's nucleus. But it doesn't answer the $64 question:  how is the choice made in the first place and the unlucky 1,999 lottery losers shut down?  How is this partially related to the neighboring cells' choice, presumably because they are recent cellular descendants of each other in the growing nasal epithelium?  And how is the choice totally independent in other cells?

The OR genes are members of a large, ancient family of genes that code for all sorts of cell-surface receptors that are used in all sorts of other functions, but as far as is known (or at least as far as we're aware) their expression is not monoallelic and they are not used in a choose-one/exclude-the-others mode.  So mystery upon mystery still attends the monoallelic expression, the very nice bookkeeping strategy, by which we can tell when we smell a rat.

What is curious is that the same mechanism does not seem to apply to the various other known forms of monoallelic expression, described on Friday, each of which is very deep in evolutionary terms.  And more curious is the question: how widespread is monoallelic expression, if it exists, in other systems where choosing among many genes is not the 'trick' but in which there may be reasons for only one copy of a gene to be used in a given cell in a given context?  If recent evidence is any guide, we will find many more examples.

This is yet another way in which our rather stereotypical, and one may say superannuated Mendelian concepts are in need of some serious rethinking.

The genetics of olfaction: In evolution, where there's one, or one thousand....there's more. But how much more? Part I.

The way we're taught genetics in school, we have two copies of every gene and both are expressed in their appropriate tissues.  That expression is based on regulatory sequences near to the gene, and when specific proteins (call them Transcription Factors, or TFs) stick to those sequences, the nearby gene is transcribed into messenger RNA which is then used to make the protein the gene codes for.

Thus, like the pair-by-pair march onto Noah's Ark, your two sets of globin genes march on--are expressed--in red blood cells, your digestive enzyme genes in the gut, neurotransmitter genes in brain cells, and so on.  So if you have an 'aa' genotype both 'a' alleles are expressed, but if you're 'Aa' your cells get a comparable dose of each....just as Mendel told us.

Well, not exactly!  Each copy is expressed at a level determined by its nearby regulatory DNA sequences.  These could be very different (due to inherited variation in their sequence details, among other things), so even if both copies are being used, they may not be being used as much.

But that's not all.  Sometimes we find monoallelic expression: only one of the two copies is being used!  Our knowedge of this once-strange exception to the rule has been growing rapidly in recent years.  Now we know there's nothing exceptional about it.....but how 'nothing' is that?

X marks the un-used: monoallelic expresssion and sex determination
Female humans and other mammals are specified by having two X chromosomes.  Since males have only one X and a very different Y chromosome, one might expect the delicate cell environment to be impaired in males,  with only a single dose of all the X-linked proteins, when for the rest of their genes on all the 22 other chromosomes they had a double dose.  But instead, it has been found that in females, for about 85% of the genes on the X-chromosomes, only one of the copies that she carries is actually used.  The other is silenced.  Early in a female embryo's development each cell picks one of its X's to use, and inactivates the other (except for about 15% of genes, when both are used).  Thereafter, descendent cells express only the chosen X.  Since this is random, about half a female's cells use a gene on one X chromosome, the other half genes on the other X.  The use of only one of two available genes in a given cell is known as monoallelic expression (the two copies being 'alleles' of the same gene).

The patchy distribution of color on tortoiseshell cats results from the random inactivation of one X chromosome in females. Scitable, by NatureEducation,

Monoallelic expression, beyond the X
A specific mechanism for this was worked out, starting decades ago, and today much is known about how it works.  It was long thought to be a unique random-selection-inactivation phenomenon. But things in evolution seem to pop up again and again, sometimes in very different guises.  For example, some decades ago it was found that the antibody genes that are expressed in white blood cells and work against microbial infections have something similar to X-inactivation....but also quite different.  Each white cell selects from a large array of possible gene sections, chains them together to form the messenger RNA for an antibody protein, then cuts the un-used sections from the DNA and discards them from the cell.  Not only that, but the other of the two copies of this entire gene is inactivated.

The protein coded by the activated genecopy is shepherded to the surface of the white blood cells, and as the cell circulates in the blood or lymph, it bumps into whatever is around.  If it happens to bind to something it recognizes, like a nasty virus particle, a cascade of events is triggered.  Among other things, the excited white cell divides rapidly, making more of its kind (that remember and continue to express the chosen antibody gene), so they can detect other copies of the infecting villain that are in the body.  When successful, this triggers a cascade of hunt-and-destroy activities that rid the body of the infection.
 

B Cell development; Weiss and Buchanan; Genetics and the Logic of Evolution, 2004

I've omitted many details, but it is in this basic way that you fight the kinds of infection your inheritance can't have directly predicted--that is, that you didn't evolve specifically to be able to ward off. It works because basically every white blood cell lineage is using a somewhat different antibody molecule, and this  diversity makes it possible to molecularly 'recognize' unpredictable foreign molecules.  There are several gene complexes, located on different chromosomes, that are used in essentially the same way in the formation of various white blood cell types. How these choices are made and the antibody gene regions identified is not fully known.

A given pathogen or alien substance, pollen, virus, bacterium etc., has its molecular surface characteristics.  If one or (usually) more than one part of its surface can be recognized by at least one lineage of immune system cells because of the particular variant receptor they have on their surface, then the pathogen can be grabbed and surrounded by such cells or antibody molecules, and destroyed.

Is that all there is to monoallelic expression?  By no means!

Olfactory receptors: monoallelic expression big-time
How do you smell?  We're not talking about your BO or your Chanel No. 5 sex appeal.  No, we mean how can you smell?  How can you tell it's bananas, lemons, or a dead rat you smell when you walk into a dark room?

The world is drenched in molecules wafting through the air, all around us, and of an essential infinity of different structures.  If detecting food or predators has always been important, one would expect the system to be quite specific, or at least quite sensitive.  In fact, while there may be some specific smell senses (see, pheromones, in Monday's post), in animal evolution it has apparently been generally safer to generate a large array of more or less random odorant detector molecules so that your chances that at least one of them would be sensitive to any odor molecule you may come across, and be sensitive to, and able to remember and discriminate among such molecules.   Your ancestors needed to be able to tell if they smelled a rat or a ratatoulle.

As with the unpredictability of potential sources of infection, you can't know what odors you may be exposed to or want to identify--and remember.   In fact, this has been achieved in mammals and even in insects by a kind of combinatorial mechanism very similar to that used in immune defenses.....but with entirely different mechanism!  We'll talk about that next week.