Biophysics 2024 in Philly
Ctenophores provide an example independent experiment in potassium channel evolution
Comb Jellies, ctenophores, have been in the spotlight recently because genome analysis pretty convincingly says that they are the oldest animal lineage still around today. What is cool about this is that they have nervous systems. That means animals like sponges and placozoans lost a nervous system that was present in the common ancestor of all animals alive today! But close analysis of ctenophore genomes suggests those nervous systems are very different from “ours”, ours being all other animals with nervous systems – from sea anemones and jellyfish to flies and humans. They are missing a lot of the neuronal gene families that all the rest of us have – so many in fact that some people argue comb jelly nervous systems evolved independently. We don’t think that is case, but they certainly split from us at a time when nervous systems were pretty simple. That means ctenophores represent a partially independent experiment in nervous system evolution. We are channel people, so one of the questions we are interested in is what ion channels were there already in those earliest nervous systems when we split from comb jellies and was that enough for them? One of the cool things about us is that we have eight functionally independent types of voltage-gated K+ channels. These channels control how electrical signals are patterned as they move through our neurons. Having 8 different types that we can express in the same cell without crosstalk allows us to build neurons with very complex and regionalized electrical properties. Its key to our behavior. We can trace these 8 types to a common ancestor of humans and sea anemones, but when it comes to comb jellies, we only find 2! Comb jellies do have a lot of genes for these channels, as many as humans, but almost all are a single type: Shaker channels. So, does this mean comb jelly neurons mostly just express one type of voltage-gated K+ channel? That could limit their electrical complexity and the behaviors their neural circuits could support. We are functionally characterizing comb jelly Shaker channels to find out, and Ben’s first paper on them (see publications page) came out last month and gives a nice preview. It looks like their big Shaker family has independently evolved into multiple functionally independent types that could be expressed in the same cell. So, ctenophore neurons are probably electrically complex, they just had to do some of the evolutionary work to get there on their own. Ben also showed some comb jellies Shaker currents look an awful lot like ours. Stay tuned for his next paper where we get into the diversity of comb jelly Shakers in a bit more detail. The short story is they have proliferated to cover a lot of the functional diversity of our eight gene families. So maybe we aren’t so different after all?
The new Textbook of Ion channels is out!
I wanted to put in a plug for the new Bible for our field, The Textbook of Ion Channels, edited by Jie Zhang (UC Davis) and Matt Trudeau (University of Maryland). This is a 3-volume set that touches on almost everything you need to about Ion channels, from history of the field and modern electrophysiogical and biophysical techniques to detailed descriptions of most of the major classes of channels found in animal models. Its written at a level that serves beginners and experts alike, and would be a great choice for an upper level class. Jie and Matt brought together a ton of channel experts to put this thing together and it turned out great! Check out our Chapter “Taxonomy of Ion Channels” where we present an overview of ion channel diversity and evolutionary relationships across the breadth of the tree of life. When I was training way back in the 90s, we always assume that animals had the most complex ion channel sets because of our nervous systems and that seemed to be supported over the next decade as we observed many fewer channel types in other Eukaryotes like plants and fungi. But that turns out not to be the whole picture – we highlight in this chapter that ancestral eukaryotes already had all the major structural classes of ion channels we recognize today. Animals are unusual because we kept everything while many other eukaryotic lineages lost some! One of my other favorite things in the chapter is showing how the Kv2 voltage-gated K+ channel gene subfamily evolved from a single channel in ancestral deuterostomes into a set of 12 channels that provide a diverse set of delayed rectifier currents to our nervous systems. You can’t get from that single ancestor to our 12 genes by simple addition: the whole story includes gene locus duplications, two genome duplications, and a bunch of gene losses. We can pin down all this evolutionary history now because we have genomes from all the key species from echinoderms to the full diversity of vertebrates!
Cnidarian nervous systems not so simple after all
Genome-Scale Analysis Reveals Extensive Diversification of Voltage-Gated K+ Channels in Stem Cnidarians. Lara, A., B. T. Simonson, J. F. Ryan, and T. Jegla. 2023. Genome biology and evolution. 15(3), doi: 10.1093/gbe/evad009.
Bilaterian animals have centralized nervous systems and complex signaling needs and have thus always been viewed as are more complex than cnidarians such as sea anemones and jellyfish that have only diffuse nerve nets. But does this translate to more genomic complexity in the molecules that fast neuronal signaling? We took a look at this question by comparing the ancestral set of voltage-gated K+ channels in cnidarians to that of bilaterians. These channels play key roles in determining action potential threshold, shape and firing frequency, and their molecular diversity is a major contributor to the electrical diversity of neurons. We surprisingly find it is ancestral cnidarians that were the champions of K+ channel diversity, with almost 4X as many functionally distinct channel types as ancestral bilaterians. Electrical signaling in cnidarian nervous systems is likely to be far more complex than their comparative simplistic anatomy would suggest. It could be that cnidarians are much more reliant on creating behavior from circuits where neurons have diverse intrinsic firing properties, while bilaterians rely more on the complexity of synaptic connections. Anyway, I am particularly proud of this paper because it is the culmination of a project I started in grad school 30 years ago when I discovered the first cnidarian ion channels.
External Cd2+ and protons activate the hyperpolarization-gated K+ channel KAT1 at the voltage sensor J Gen Physiol (2021) 153 (1): e202012647. https://doi.org/10.1085/jgp.202012647
Yunqing’s first paper showing that plant and animal CNBD superfamily ion channels adopt similar voltage sensor structures in the down state at hyperpolarized voltages. Both have divalent and proton bindind sites in the external voltage sensor in the down state. Interestingly, KAT1 is a hyperpolarization-gated channel so divalents and protons, which inhibit activation of animal EAG family K+ channels, activate it. A big question in the field is understanding why these similar voltage sensor conformations oppositely influence pore opening in depolarization- and hyperpolarization-gated channels. We also thought it was cool that the plant channel KAT1 responds to a much lower pH range than animal EAG channels; plants typically have low extracellular pH compared to animals. Both channel types seems tuned to respond to the pH range found in their native environment.
Cytoskeletal and synaptic polarity of LWamide-like+ ganglion neurons in the sea anemone Nematostella vectensis. J Exp Biol, 2020 Nov 10;223(Pt 21):jeb233197. https://journals.biologists.com/jeb/article/223/21/jeb233197/226253
One of the big questions in evolutionary neurobiology is figuring out how and when functional neuronal polarity evolved. In particular, we want to know whether functionally distinct axons and dendrites predate the evolution of polarized nervous systems in bilaterian animals. We have been developing the sea anemone Nematostella vectensis to address this question. Cnidarians like Nematostella are cousins to bilaterians, but split off when nervous systems were still comparatively simple diffuse nets. This is Michelle’s paper showing that a major characteristic type of the cnidarian nerve net is non-polar and has axon-like processes that likely signal bidirectionally. We don’t think that is the end of the story however, so stay tuned – there may be other cell types that point to a more complex story in which at least some important aspects of neuronal polarity evolved in early nerve nets. We are particularly proud of this paper because it took years to develop the tools we needed to enable live imaging of transgenic sea anemones.
The S6 gate in regulatory Kv6 subunits restricts heteromeric K+ channel stoichiometry http://jgp.rupress.org/content/150/12/1702
This paper was the core of Aditya’s thesis and is a great example of how you can use evolution to highlight key features of proteins. K+ channels are tetrameric and can typically form as homotetramers composed of four identical subunits. However, some subunits have evolved a “regulatory” phenotype in which they can no longer make homotetramers and instead depend on mixing with other closely-related K+ channel subunits to form functional channels, often in an unusual asymmetric 3:1 ratio. We identified sequence changes that correlated with evolution of the regulatory phenotype and found that the interface between subunits in the lining of the pore itself played a key role in determining channel composition. Previously, it had been thought that a cytoplasmic sorting domain, T1, was fully responsible for channel subunit composition. We proposed a two-step model of how you can evolve asymmetric channels that involves sequential mutation of T1 and the pore. This is one of those typical JGP papers with heroically complicated biophysics experiments. We also received a great assist from the Hancock lab who worked with us to count subunits in individual channels using fluorescent tags.
Evolution and Structural Characteristics of Plant Voltage-Gated K+ Channels http://www.plantcell.org/content/early/2018/11/01/tpc.18.00523
This a review Greg and I wrote with Sally on the evolution of Plant Voltage-Gated K+ channels and the CNBD channel superfamily to which they belong. We managed to squeeze a lot of new analyses into it, so its sort of a hybrid review/research article. Here is my favorite thing in the article: 1) CNBD superfamily channels, which include the plant channels and a wide range of animal channels that control sensory perception, neuronal excitability and heartbeat, come from a prokaryotic ancestor that is common in eubacterial lineages but appears to be absent (at least so far) in the Archaea. Since eukaryotes evolved from the Archaeal lineage, that could mean we picked up the CNBD channel superfamily by lateral gene transfer. There of course are plenty of documented cases of lateral gene transfer, but I still think its amazing that the channels that we use to see and smell and the channels that plants depend on to open and close their leaf pores may have been “borrowed” from bacteria. The Plant Cell is of course nothing new to Sally, but it was nice for me to bag another top journal for my “life list”.
Research Opportunities
Its hard to believe the semester officially starts next week! Welcome to new students and welcome back to those who are returning! Our lab is looking for new members at both the undergraduate and graduate levels. We have a wide variety of projects examining the function and evolution of neurons and neuronal signaling, and we are looking for highly motivated students to expand our team. If you are looking for research experience in cellular and molecular neurobiology and are interested in a career in the biological or biomedical sciences, consider joining us. All inquiries should be directed to Tim Jegla (tjj3@psu.edu)
Discovery rewriting the evolutionary history of the nervous system
Here is a link to the Penn State News story on the Axon Initial Segment paper:
Rewriting the evolutionary history of the neuron
Almost all features of our neurons, from the ion channels that underlie electrical signaling to cellular structures such as axons, dendrites and synapses are ancient and are therefore shared among all living bilaterian animals. However, one critical feature that appeared to have a much more recent evolutionary origin was the axon initial segment (AIS), a specialized compartment at the beginning of the axon that serves as a barrier for maintaining axon identity and as the site of action potential initiation. The AIS was believed to be a recent vertebrate innovation for precision signaling, because the giant ankyrins which link AIS ion channels to the cytoskeleton and are required for barrier formation were believed to be vertebrate-specific. We collaborated with the Rolls lab to show that giant ankyrins instead have a much earlier origin in an ancestor of all bilaterians. They appear to organize an AIS-like domain in the axons of fly sensory neurons, suggesting that the AIS itself is part of that ancient, shared bilaterian neuronal heritage. We think this work will establish Drosophila as a model system for detailed molecular genetic dissection of AIS function. This is important because AIS biology has many important open questions and AIS dysfunction plays a role in a variety of nervous system disorders.
http://journals.plos.org/plosgenetics/article?id=10.1371/journal.pgen.1006457
Nicotinamide (vitamin B3) activates invertebrate TRPV sensory channels
Check out the link below for Wendy Hanna-Rose’s story on identifying nicotinamide as the first endogenous activator of sensory TRPV channels in invertebrates. The most notable role for our TRPV channels are as heat sensors, and compounds such as capsaicin in chili peppers can activate them. Invertebrates seem to use TRPV channels mainly for mechanosensation – they have been studied for years in C. elegans, but nobody had ever been able to functionally express them until Wendy’s lab did a series of elegant genetic studies that suggested nicotinamide could be an agonist for worm TRPVs. Her grad student Avni Upadhyay found that in worm mutants with elevated nicotinamide levels, cells expressing two TRPV channel subunits OSM-9 and OCR-4 die. This phenotype could be rescued by knocking out either TRPV subunit. We confirmed that nicotinamide directly activates both worm and fly TRPVs. We also worked with Will Hancock’s lab (and Keith Mickoajczyk in particular) to figure out that nicotinamide-sensitive worm TRPVs form functional channels as a 2:2 heteromer of OSM-9 and OCR-4 subunits.
Complex nerve-cell signaling traced back to common ancestor of humans and sea anemones
Check out the Eberly College news story on our PNAS paper exploring the evolutionary origins of voltage-gated K+ channels. We find that many of the key channel types that shape electrical signals in our neurons were actually missing in the first nervous systems. However, they are still ancient and we can trace them all back to a common ancestor of humans and cnidarians (animals like sea anemones and jellyfish). We want to know why – what important event was going on in nervous system evolution at that time?
http://science.psu.edu/news-and-events/2015-news/Jegla2-2015