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Monday, October 29, 2018

Domestication of Yeasts

Saccharomyces cerevisiae is known as the Baker's Yeast. It has helped us make bread, beer, and wine since before recorded history. These days we also use it to make fuel, pharmaceuticals, and for basic biology research. With the innumerable industrial, food, and research purposes we use it for, it is a thoroughly domesticated organism.

With the various mammals we've domesticated, researchers have identified a "domestication syndrome"; a set of features common across domesticated animals. They have shorter faces, milder temperaments, reduced weaponry (teeth, horns, claws), and color changes. In short, they've become cuter. To some degree these are traits that could have been actively selected for, but it turns out that if we only select on temperament, all of the other traits come along for free because all those traits are mediated by the action of neural crest cells throughout the body.

Now, yeast don't have neural crest cells, but they're still domesticated. It didn't evolve to have a more amenable temperament, but it did evolve to grow rapidly in the amenable conditions we provide for them. There's a different sort of "domestication syndrome" that it would have developed along the way. Any trait or ability it needed to live as a wild yeast, but did not need to live under our care, would be lost. This would happen because any lineage that dispensed with those traits would be able to grow faster without the energy drain they represent.

So. What traits would yeast lose under domestication? It's not entirely clear. We can't just look at the cells and see a difference. Nor do we exactly have the wild progenitor yeast around to make comparisons with.

Here we're going to take a bit of diversion.

My first major project in grad school was to figure out how to use flow cytometry to determine the genome size of a different yeast called Candida albicans. In the past, This analysis had proven difficult to do with this yeast for others. This difficulty had been generally blamed on the organism's ability to grow either as independent yeast cells or as elongated hyphal cells that get all tangled up in each other.

I started with protocols developed for S. cerevisiae. At three months in, I was testing yet another protocol variation and the data that came out of the experiment looked like the figure at right. Previous data had much broader, indistinct peaks. (I'm sure I have some of those early figures around somewhere, but I'm not going to spend a bunch of time digging for them.) I was amazed and quickly set up a repeat of the exact same experiment. It failed miserably.

I had made a mistake somewhere in the protocol which made things work. Because it was a mistake, it wasn't written down in my lab notes. You can only write down what you know you're doing.

It took me another frustrating month to figure out what it was I had done wrong. I had used way too much EDTA in the buffers for processing the cells. With this improved protocol, I could get good flow cytometry data from even the most difficult hyphal-growing strains of C. albicans. This disproved the previous theory as to why this species was difficult to work with while doing this assay.

Subsequently, the protocol proved effective with every random yeast species I was able to acquire for testing. I never tested them with the original S. cerevisiae protocol for comparison. In retrospect, I consider this to be an oversight.

The flow cytometry protocol has since then been used in numerous papers from several separate labs. The flow cytometry protocol and analysis tools I developed become the second chapter in my thesis. The idea of wrapping up the material into a paper did come up after I graduated, but I really didn't have the time/energy to dedicate to the process. Researchers should probably cite that chapter, but I know that thesis chapters tend to only get cited rarely. If you are interested in all the details, you are welcome to have a read.

I pretty quickly developed a working theory about what was going on. EDTA binds to divalent cations (Ca2+ and Mg2+) in solution, locking them up so other enzymes don't have access to them. Many enzymes require certain levels of these ions to function normally. For whatever reason, the endogenous nucleases of C. albicans were much less sensitive to low levels of divalent cations than those found in S. cerevisiae. Now, I couldn't think of any way to test this theory. I wasn't in a biochemistry or structural biology lab, so the techniques that would have been useful were well outside our wheelhouse.

This uncertainty has stuck with me for the roughly seven years since then. Just a couple days ago, I developed an idea that in some sense explains the results. Domestication.

S. cerevisiae is a thoroughly domesticated species. It hasn't had to fight for what it needs, so it could very well have evolved enzymes that are used to easier environments with more consistent levels of necessary ions. I strongly suspect the flow cytometry protocol for S. cerevisiae only works because of the domestication syndrome of traits found in S. cerevisiae.

I'm not sure how one would test this theory, but it sure seems to make sense of the observations so far.


Monday, October 1, 2018

The Color of Beans

I've been looking for some blue-colored beans for several years. Its easy to find beans in a range of colors (red, pink, white, yellow, green, black), but blues are a rarity in beans. Early on I found an Italian bean called "Nonna Agne's Blue Bean", but the only seller in my country was out of stock. Sometime along the way I received an offer of some French heirloom blue beans via a facebook connection, but no seeds ever appeared. (She offered them for free, so I can't complain too much.) Blue beans are around, but they're rare.

Last year I received some beans from an online collaborator after I had mentioned my interest in blue beans. She said one of her plants that season had turned out to be an unexpected hybrid that produced blueish seeds. The three seeds that arrived are shown at left. To my eye they were basically black, but with maybe the slightest blue cast. I wasn't optimistic, but after the difficulty I'd had finding blue beans I was going to give them a try.

Two of those three beans sprouted. This was kinda a dramatic time, as those two sprouts could easily have died and then another possible blue bean lead would have gone nowhere. Fortunately, both plants thrived.

A few months later I had a small pile of new beans. When I started shelling them I was very pleased to see some distinctive blue color. As the beans age and dry down, they start to produce some tan pigment which muddies up the pretty blue.

Next spring I'll plant enough of the more blue beans so I can grow enough to make a few meals of them. Right now I have too few to make a meal and have enough for planting.

How did I know that the biology of bean color should be able to produce a blue bean? The red color of beans is due to a group of biological pigments called anthocyanins. This same group of compounds is also responsible for the rare blue pigments we see in biology.

An analysis of black beans showed most of the anthocyanins to be delphinidin (at 56%), with lesser amounts of petunidin and malvidin (26% and 18%, respectively). Delphinidin and malvidin are responsible for blue color in various flowers. The petunidin is described as having a dark-red/purple color. All together, this suggests that black beans really are just super-dark blue beans. This is corroborated by references I've heard of black beans crossed to white beans sometimes producing distinctly blue beans in among the progeny.

So, why are blue beans so rare? I got nothing that explains it. Blue is such a lovely and generally rare color that I would have thought people would have been growing blue beans as much or more than the now-common red beans. Maybe I can help rectify the situation in time.

As I was writing this post I decided to look around again for vendors selling blue bean varieties. I found a European vendor that seems to have stock of the Italian "Nonna Agne's Blue Bean". I also found another unrelated blue variety called "Blue Shackamaxon Pole Bean". I might think about ordering some of each, but it'd be more fun to make my own now that I've got a start at it.


Monday, September 24, 2018

Botanizing in Hawaii: Hawaiian Pepper

Closeup of a pepper plant branch. Several leaves hang down, with a few small elongated peppers and small whitish flowers raised above the leaves.
One of the plants I really wanted to find on my trip to Hawaii is known as the Hawaiian Pepper. This semi-wild pepper plant is generally referred to as a type of Capsicum frutescens, though you will often find references to it as different varieties of C. annuum. The ancestors of these chiles were first introduced to the islands around 1815, but they have since been integrated into the local culture and are often described as native. The small size and non-aggressive growth of the plants has allowed them to integrate into the island ecosystem without being too disruptive. Like other wild peppers around the world, birds also help distribute the seeds.

You can order seeds for it (I have no affiliation with the linked company, but found them via a quick search.), but I wanted to find the species growing wild on the islands.

Wider view of a whole pepper plant, with dried grass and shredded wood mulch around the plant.The plants I found were... not exactly growing wild. As I walked along part of the resort where we were attending a conference, I glanced through a gap in some hedges and saw the characteristic look of chile plants. When I walked around behind the hedges, I found what looked like a little guerrilla garden someone had setup outside the watered and maintained landscaping of the resort. There were several Hawaiian Pepper plants of about the same age/size spaced about the area. I suspect someone who works on the resort planted them and would go by every now and again to tend to them.

I collected a few dried pods that had dropped to the ground around the plants. I didn't grow any this year, but I did send some seeds to a collaborator out in California. (They're on twitter as @ChaoticGenetics. Go check them out!) Last I heard the plants were growing well.


Monday, September 17, 2018

Botanizing in Hawaii: Railroad Vine

Green vines stretched out across the pale sand. There are a few pink flowers along the vines at left.
This is a plant that I knew from my childhood visits to the south Texas shore. Railroad Vine (Ipomoea pes-caprae) is a cousin of the common Morning Glory vine that is specialized to live on beach-side sand dunes. Its seeds are salt-water tolerant and are distributed widely by ocean currents. It grows on tropical and sub-tropical beaches worldwide. On Hawai'i, we only found it growing in one location. Most of the beaches we visited were too rocky for it to prosper.

Closeup of a pink flower with leaves around it.
Closeup of a single leaf. The leafe looks something like a round paper plate folded in half, with a stem at one end.The flowers seemed to wilt under the intense sunlight. If we had found them earlier in the day, they probably would have looked more like my childhood memories of them.

The leaves are thick and smooth, with a major crease down the middle. My recollection is that the common name, "Railroad Vine" has to do with the plant's habit of growing long strait vines along the sand, with evenly spaced leaves.


Monday, September 10, 2018

Botanizing in Hawaii: Solanum linnaeanum

Closeup of flower, leaves, and stem of plant. Flower is pink with four fused petals, forming a square, and has yellow anhers gathered in the center. The leaves are heavily lobed, with long sharp spines protruding from the underside. The stem too is covered in dramatic spines.
Hawaii has a long history of biological invasions. Plants and animals from all over the world have arrived and thrived there under the tropical sun. This can make it somewhat difficult to identify a random plant, because it could literally come from almost anywhere on the planet.

On one of my hikes, I found several plants I immediately recognized a member of the family Solanacea. This is the same family that includes tomatoes, peppers, and eggplants. I was pretty sure it was even a member of the genus Solanum, one of the several species sometimes referred to as "spiny eggplants".

Closeup of a few leaves and two small round fruit. One fruit is entirely brown. The other fruit is pale green with darker green stripes. The leaves are heavily lobed and covered in spines.
 I collected a few fruit, intending to secure some seeds for planting back home in Minnesota. A couple days later, while sorting through my collection at a motel, I finally identified the plant as Solanum linnaeanum. Among its various common names are: "Poison Apple", "Devil's Apple", and "Apple of Sodom". They're native to parts of southern Africa, but have naturalized in Hawaii and various other places around the world.

The plant and its fruit are chock-full of toxins. Enough so that very few animals are willing to eat it. At about this point I decided not to take seeds home for garden trials. I do have a few seeds in my collection that come from highly toxic, or otherwise dangerous, plants. I wasn't entirely certain how much difficulty I would have with trying to leave Hawaii with collected seeds. If some official asked me what they were and why I had them, they might not appreciate my responses. So, to limit that risk, I dumped the S. linnaeanum seeds in the garbage.

(Several days later when I left the state, I learned I would have had no trouble at all. Dried seeds in vials didn't concern the USDA officials at the airport at all. Bringing seeds into Hawaii gets their attention, not taking seeds away from Hawaii. Next time I'll be a bit more bold.)

Interestingly, it appears this species can cross with domesticated eggplant (S. melongena). Doganlar et al. studied an F2 population derived from a cross of the two species in order to map the genomic positions of genes for traits important in domestication. Unfortunately, their paper doesn't have any photos of what the F2 plants looked like. Even with the risk of dragging the toxic traits from S. linnaeanum into the progeny, this would be a fun cross to recreate and explore. The wild species probably has numerous disease/insect resistance traits that would be useful in a garden eggplant, so there is probably value to the experiment beyond simple personal amusement. There's a few online vendors offering seed for this species, so I won't have to make another trip to Hawai'i to start on this project.


Monday, September 3, 2018

Goats of Hawai'i

Group of five goats at the side of a curved road. Three goats are brown and two are black. Behind the goats are piles of dark brown lava stone and scattered clumps of dried grasses.
I visited Hawai'i last year for a horticulture conference. Well, my spouse was attending the conference. I was just going along for vacation. I spent a lot of time driving and hiking during the days when the conference was in session.

Much of the north-west side of the island where the conference was being held is dry-land, with exposed rock from several different ages of lava flows. I came across the bleached bones of pigs and other large animals among the lava, but rarely saw any sizable living creatures.

One day I was driving out to a nearby park to do some hiking and I saw a group of goats crossing the road. I lucked out and was able to capture a few photos like the one above. What immediately struck me about the goats was that they were colored just like many of the aged lava stones I had been seeing the previous few days. They didn't have any of the white markings so common on goats I've seen almost every where else.

It made me think the goats might have been under a pretty severe hunting pressure and that their colors represented adaptive camouflage, protecting them somewhat from visually-hunting humans. If the goats had been resting among the rocks as I drove by, I likely would have thought they too were just rocks.

Goat hunting on the Big Island is allowed year-round in some places, with defined seasons in other areas. There have been intense and largely successful goat eradication efforts in the larger fence-enclosed parks on the island. This represents a fairly high level of hunting pressure, which would definitely be expected to select for traits that help the animals avoid predation.

Unfortunately, I have been able to find no research on the topic of the evolution of wild goats of Hawai'i due to human hunting. This might be a nice topic for a PhD for some motivated student living on the island. Let me know if you come up with anything.


Monday, July 30, 2018

Growing Bur Oak Trees 2

Burr Oak acorn cap.
One of my long-running interests has been domestication of oak trees for food. Now, you can already prepare and eat acorns and there is a long history of native peoples around the world doing so, but rarely does it seem like the oaks have been transformed by the process. What would be ideal is an oak tree that produced very large acorns which were very low in the tannins that make most acorns inedible without intense processing.

Several years back I found a Burr Oak (Quercus macrocarpa) tree with huge acorns littering the ground beneath it. When I broke one open and tasted it... It was straight up sweet. I probably could have taken home a bag full and made a meal from them. Instead, I collected several that looked in the best condition and took them home (to Minnesota) to grow.


It's now been a couple years and the young trees have not only survived our winters, but they've been thriving. This wasn't a forgone conclusion. The seeds came from trees growing about two thousand miles south of where I planted them. This goes against the general guideline of planting tree seeds collected from somewhere near where you plant to grow them.

Each tree is distinct, with leave size and shape variations. They're all different heights too. I suspect these early differences in growth rate will continue.

Burr Oak seedlings, each photographed from above and the side.

The trees have only been outside for a couple winters, so it isn't guaranteed that they will survive long-term. Either later this summer or early next year I'll be transplanting the seedlings to cleared spaces in our woods where they can spend the rest of their lives. The local squirrels will be pleasantly surprised in about eight years when the seedlings should start making their first acorns.