Posted by PZ Myers
https://freethoughtblogs.com/pharyngula/2025/06/29/a-conserved-genetic-signal-for-building-neuronal-pathways/
https://freethoughtblogs.com/pharyngula/?p=76830
A new video! This one is just science, a cool paper I read way back when I was doing my post-doc. It left a strong impression on me, so I thought it would be worthwhile introducing it to all of you.
Today I want to talk about one of my favorite papers, this one:
It’s titled “Genetic Analysis of a Drosophila Neural Cell Adhesion Molecule: Interaction of Fasciclin I and Abelson Tyrosine Kinase Mutations”, by Thomas Elkins, Kai Zinn, Linda McAllister, Michael Hoffman, and Corey Goodman. Right there in the title you can see a few of my favorite things: genetics and the nervous system, and as we dig deeper you’ll find it’s also about development and relevant to evolution.
Those of you with sharp eyes may also notice that it was published in 1990 — it’s 35 years old. As an aside, I have to mention that I sometimes get students asking me if it’s OK to cite papers older than 5 years, which I find ridiculous. Of course you can. A good paper does not become obsolete, except perhaps in its interpretations. The data should hold up, although it can be improved upon. I cited papers from the 1830s in my Ph.D. thesis!
This paper holds up. I was impressed when it came out, and it affected how I think about genetics and evolution. At the time, it was incredibly relevant to the work I was doing at the time, on neuronal pathfinding. I still think it’s cool stuff.
In order to explain what’s going on in this paper, I have to provide a lot of background, so bear with me while I tell you a little bit about the technology we used in that ancient era (and still use today!).
Monoclonal antibodies
One of the methods used here was the generation of monoclonal antibodies. To do this, they took embryonic grasshoppers, ground them into a paste, added some adjuvant to stimulate the immune system, and inject the goo into mice. The mouse immune system reacted by generating antibodies against grasshopper antigens. To make them monoclonal, you cut out the mouse’s spleen, which contained the antibody producing cells, teased the tissue apart, and then fused individual cells with cancer cells to immortalize them, and then the cells are separated into individual vials, where they each make an antibody that binds to a different, single epitope. At that point you’ve got a lot of vials, each of which allows you to label a different molecule in the grasshopper.
That’s the relatively easy part. Then what you have to do is dissect hundreds of grasshopper embryos, paint each one with one antibody, and ask where the antibody sticks. The Goodman lab isolated a lot of antibodies and characterized what they bound to, and gave them names. They were specifically searching for antibodies that bound to the developing nervous system that might play a role in building the structure of the CNS.
NS structure
In the insect embryo, that structure is fairly straightforward. The nervous system is made of a chain of repeating ganglia extending the length of the animal.
These ganglia are connected by a pair of nerve pathways, one on each side of the embryo, called the longitudinal fascicles.
Each ganglion contains a pair of nerve pathways that connect the right and left sides; one is called the anterior commisure, and the other is the posterior commissure. It’s easy to visualize the early nervous system as a kind of ladder, with the longitudinal fascicles forming the side rails, and each ganglion containing two rungs, the commissures.
We also have peripheral nerves branching off to innervate the periphery.
These fascicle are like cable conduits. They contain multiple fibers, and they grow as new neurons mature and send processes into them, and each process is covered with specific molecules that act as recognition and adhesion factors. The specific combinations of these molecules is an important factor in navigation — how does a cell on the left side of the ganglion make connections with, for instance, a muscle in the right forelimb of the animal? One way is to follow the coded molecules set up by the earlier axons that had grown to form these pathways. So…turn left at the first longitudinal fascicle, follow it until you reach the posterior commissure, turn right, grow until you reach the next longitudinal fascicle, follow that until you bump into a peripheral nerve, and change tracks to grow until you reach muscle. You can build a complex nervous system that way, with fairly simple rules for each neuron.
You can see these fascicles and commissures easily with a microscope, but the molecules that label them are invisible. That was the beauty of making these monoclonal antibodies — you can use them to reveal the molecular coding present on each of the fibers running through the conduit, and you can finally see what signals the growing nervous system is employing. Some of the antibodies label everything in the CNS, like this one, called BP102, which allows you to clearly see the ladder-like structure of the embryonic nervous system.
Others label subsets of the processes, and those are the most interesting ones. The Goodman lab identified multiple antibodies that can be used to map out portions of the developing CNS, and some of them were named fasciclins, because they are expressed on the surface of axons in various fascicles. These are typically cell adhesion molecules, molecules that make the neuron ‘sticky’ to other cells, especially to some cells and not others.
It’s a molecule identified in grasshoppers, but it is homologous to molecules found in people. You won’t be surprised to learn that it’s also expressed in fruit flies, and the pattern of expression in flies and hoppers is almost identical. I spent a lot of time staring at hopper nervous systems through a microscope, and got so familiar with them that I could instantly spot and name individual cells — the terrain was like home to me. When I later got to look at the embryonic and larval fruit fly nervous system, it was a revelation. I could see the same landmarks and follow the same cells in both.
Flies and hoppers are separated by over 350 million years of evolution, so that nervous system ladder is a highly conserved structure in insects.
The one obvious difference is that flies are tiny. I was used to working with a microelectrode and micromanipulator to poke around the hopper nervous system, but no way was I going to be able to do that with flies — flies were miniaturized hoppers, and while hoppers were great for cellular work, flies were far outside my skill set. Fortunately, the Goodman lab was using genetic tools on flies, with far greater precision. They identified these promising molecules in hoppers, and then applied them to flies, where we can play all kinds of genetic games.
One of the molecules identified is called Fasciclin I, and that’s the focus of this paper by Thomas Elkins. FasI is a glycoprotein expressed on all peripheral axons, a subset of the axons in the commisures and longitudinal fascicles, and some non-neuronal cells. It has been genetically mapped — it’s in division 89D of the third chromosome of Drosophila, and it’s been isolated, cloned, and sequenced. If you’re vertebrate-centric, it has several homologs in humans: periostin and stabilins, for instance. Basically, it’s a small homophilic molecule on the surface of cells, that make cells expressing FasI stick to and follow other cells expressing Fas1.
The question Elkins and others were asking is specifically what Fas1 is doing in the nervous system, and it’s expression pattern is suggestive. It is found in “a ventrolateral cluster of neuronal cell bodies (arrows at top left), a fascicle in the posterior commissure (middle arrows at right), and the intersegmental nerve root (arrows at bottom left).” So, not everything, but a select subset of pathways. In the case of the commissures, what we see is a few thin fibers that cross the midline, so one hypothesis is that these are pioneer axons: one slender thread is sent across the ganglion, laying down a track of Fas1 signals that subsequent fibers could follow to build a more robust commissure.
But that’s just a hypothesis, an inference from an observation of the phenomenon of gene expression. The paper goes beyond that, and tests the hypothesis by deleting Fas1 and asking what the CNS of the embryo does in response. If Fas1 in the first axon to cross is the sherpa that leads all the other fibers across, then deleting Fas1 should leave all the following axons lost, and maybe the commissure would fail to form altogether. A significant part of this paper is an exercise in molecular genetics to knock out the Fas1 gene. They started with a transposable element inserted into the Fas1 gene that disrupted its function, and then to be really thorough they used gamma radiation to delete the transposable element and the adjacent DNA that contained the broken fragments of Fas1. They totally expunged Fas1 from the genome and asked what the CNS in homozygotes looked like.
The embryonic nervous system looked totally normal, as if Fas1 was totally superfluous.
Well, not totally. They looked at adult fly behavior and noticed that they were a bit uncoordinated in walking, so maybe it plays a more subtle role in fine tuning behavior.
It wasn’t a dramatic result, though, so I’m sure it was a bit disappointing. They didn’t see a clear association between Fas1 expression and gross changes in CNS structure.
Back to the drawing board. They had another interesting gene, Abelson tyrosine kinase, which is expressed in the central nervous system, much more widely than Fas1 is. Let’s make a non-functional abl mutation, and see how that disrupts the early fly CNS!
They did. It didn’t. Homozygous abl mutants produced a perfectly normal CNS scaffold of fascicles and commissures, although they did see an increased frequency of errors in 10% of the embryos. Maybe abl is a redundant accessory to building these pathways, increasing fidelity but not acting definitively to build them?
Still, it’s not a particularly exciting result.
But what if they made a homozygous double mutant, carrying both the Fas1 null mutation AND the defective abl mutation?
Now it’s getting interesting. The commissures fail to form! It looks like normal development requires cooperative interactions between at least two different gene products, fas1 and abl. Elkins dug deeper to look at how individual neurons behave in the double mutants.
In wild type embryos, a cell called RP1 grows across the midline in the anterior commissure, and then turns to grow for a short distance in the longitudinal fascicle, and then exits via the contralateral intersegmental nerve root to grow into the periphery. In the absence of both Fas1 and abl, though, the cell continues to grow, but it’s confused, and in different animals takes different, seemingly random routes. Sometimes, since it can’t cross the midline, it just exits out the ipsilateral nerve. Sometimes it just stops. And sometimes it just keeps growing down the longitudinal fascicle, looking for that labeled pathway that no longer exists.
To sum up the important points in this paper:
- There are signals that direct the growth of neurons. The developing nervous system is a matrix of various molecules that provide guidance cues.
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These signals increase in complexity over time. RP1, for instance, is just the first cell to send a process across the anterior commissure, but as each subsequent cell uses that process to navigate, it adds its own molecular signals to the mix. If the first pioneer gets lost, there is a whole series of later neurons that are confused, and the commissure entirely fails to form.
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There isn’t one gene that defines a specific morphological feature — neuronal pathways are the product of multiple interacting molecules and membranes and cells that cooperate to build a functional whole.
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There is a degree of redundancy in the assembly of the nervous system. Mutating one gene does not cause the whole structure to collapse — this fault tolerance allows the accumulation of mutations that can lead to more subtle variations in the wiring of the nervous system.
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Divergent species can be built on the same core structure, using the same molecules, but variations can produce animals as radically different in morphology as grasshoppers and fruit flies. Species that have been diverging even longer, such as humans and insects, still share similar core patterns of organization.
It’s a great paper. Unfortunately, it was published posthumously, and I have to mention that Tom Elkins died in 1989. We lost a good one.
That puts my plight in perspective. I seem to have snapped a ligament in my knee and have either been sitting in hospital rooms or lying in bed for the past few days, with no immediate relief in sight. Obviously, it could be worse. While my patreon supporters scroll by here (you can be one, too, just visit patreon.com/pzmyers), I’ll just say that I’m lucky, I live in a small, rural, and very Republican town, and we have a good local hospital with a helpful and attentive staff. I think I’ll be seeing more of them than I’d like in the next few weeks.
Unfortunately, the Republicans in congress are making changes that will hurt all the Trump-voting citizens of my county, and are destroying medicaid and medicare, and will lead to the closure of many of these rural hospitals all across the country. The leopards will be feasting on faces here, and one of the costs will be the loss of facilities that could help sew those faces back on.
While I’m not feeling great right now, I am getting the help I need, and it could be so much worse if this administration gets its way. Resist! Fight back by making sure these wicked rascal are kicked out of office! Save health care for everyone, even MAGA!
https://freethoughtblogs.com/pharyngula/2025/06/29/a-conserved-genetic-signal-for-building-neuronal-pathways/
https://freethoughtblogs.com/pharyngula/?p=76830
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loganberrybunny
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Unlike the Sugar Gem oranges, this watermelon was sweeter than a regular ol’ watermelon. Not only that, but the label boasts a rich, red flesh. I thought it may have been all talk, but lo and behold it was indeed very red! I bought this one for six dollars, which is pretty much the exact same cost as a regular watermelon, and it’s roughly the same size, so I’d say you should go ahead and buy this one over the regular ones if you are someone who prefers a juicier, sweeter watermelon.
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