As Many Exceptions As Rules: January 2013

Wednesday, January 30, 2013

Carp Diem - Polyploid Fish Seize The Day

Biology concepts – polyploidy, invasive species

There are 60,000 different species of weevils, a type of

beetle. And almost all of them are polyploid! Polyploidy

benefits speciation, so maybe them being polyploid is

why there are 60,000 species. On the left is

Trachelophorus giraffa, named for obvious reasons, and

on the right is Rhigus nigrosparsus. You’ll have to go to

Brazil to see one in person.

We think that polyploid animals are the rare exceptions, and they certainly are in the case of mammals, but there are other groups of animals don’t think twice about being polyploid. Arthropods are notorious for developing polyploid lines, while amphibians and reptiles are probably the most well studied polyploids. But there are more - that cedar planked salmon you enjoyed the other night – it was probably triploid as well.

We pointed out last week that polyploidy in plants has done a lot to promote speciation events, and this seems to be the case in fish as well. While some families have a few polyploid members, like the loaches, or the carps and minnows, other families are completely polyploid, like the Salmonidae (salmon). Wat is more, the families with the greatest number of polyploid members also have the highest number of species overall. Of the 28,000 known species and >60 orders of fish, 63% fall into the 9 orders that include polyploidy – coincidence? I don’t think so.

Remember that polyploidy in plants is well behaved, not much genome restructuring goes on even though there can be subfunctionalization and neofunctionalization leading to speciation at the molecular level. In contrast, fish polyploidy seems to induce a tolerance of change, and gene ordering and genome restructuring seem to run rampant. This seems to be at least one reason for high rates of new species development in fish that are polyploid.

The effects of polyploidy on fish are similar to those we have talked about previously. Polyploid fish tend to be larger, ie. the gigas effect, and they tend to live longer and grow faster, ie. heterosis. Inductions of triploidy or formation of auto- or allopolyploid species tend to have fewer diseases. For some reason, sexual maturation in fish is linked to higher infection rates – most likely due to stress. Finding a mate and having kids is stressful, ask any adult. Stress is directly related to infection rates, as one of the effects of the stress hormone cortisol is to turn down the immune system.

The sunshine bass on top is a diploid female which

is filled with eggs (gravid). In contrast, the female

on the bottom is triploid. She is bigger, even

compared to a gravid fish. The gigas effect in

polyploids is real, and effects sport fishing. Everyone

wants to catch a bigger fish.

Whether or not ploidy level itself has an effect on immune fitness is up for argument. A 2012 opinion paper from three prominent researchers states that increased gene numbers could lead to expression of more immune proteins, and antibodies to more different parasites, so it could increase resistance. They also offer that the mere increase in genes could end up producing more immune cells in total, therefore conferring more resistance.

In plants, a recent study indicates that a disease resistance cluster of genes in soybeans indicates that production of new disease resistance genes could develop by polyploid development. In an autopolyploid soybean, the number of disease resistance genes doubled, but they didn’t produce twice as much protein. It seems that they have begun to evolve independently. This may in turn produce newly functional resistance genes, or on the other hand, may eliminate one of the clusters. It appears that specific immune function and polyploidy may be interpreted only on a case by case basis.

But there negative effects that are similar to plants as well. Triploid species are often less reproductively active, due either to difficulties in gamete production or to aberrant sex steroid levels as a result of dosage imbalance.

In some cases though, sterility has been used to the advantage of humans - triploid salmon are less likely to return to spawning grounds, which means they stay in the ocean longer, growing fat and happy. For wild salmon fisheries, this means a greater number of bigger fish. For salmon hatcheries and commercial growers, it means less stress on the animals and a greater harvest. Triploidy can be induced in the salmon (and other species) by cold shocking the eggs near the time of fertilization or using chemicals to prevent chromatid separation during meiosis.

Triploid oysters on the west coast are raised so that they can be

harvested year round. They are bigger and taste better

than spawning diploids. They are also more disease

resistant, and this might affect pearl formation, since most

natural pearls are induced by parasites that bore through the shell.

For a reason completely different than organism size or stress, oyster farmers have also induced triploidy in their product organisms. It seems that spawning reduces the sweetness and size of the oysters. They taste “spawny.” Since triploid oysters do not spawn, they retain their size and sweetness throughout the summer, when diploid oysters would be less tasty and are not harvested.

Pacific oyster species use up to 80% of their body weight for production of sperm and eggs – not good for food harvesting. This can last for most of the late spring and summer, so the triploids allow for harvesting when people are accustomed to avoiding oysters – typically, the rule is don’t eat oysters in any month without an “r.”

But if we harvest fewer diploids, and introduce more triploids – could we end up with a glut of oysters? The diploids that would have been caught are free to reproduce and we end up eating the triploids that wouldn’t have been reproducing anyway. What ecological niches might be disturbed by too many oysters? You can discuss amongst yourselves whether this is a good idea in the long run – I render no opinion one way or the other.

You can argue both sides of the polyploid introduction argument; human efforts to enhance (alter) the zoological face of the planet have met with some disastrous failures, but remember that majority of foreign introductions have been ecologically moot. This point is often overlooked, but we have talked about it before.

Polyploidy in wild salmon is extremely common, so would there really be that great a change? The use of triploid induction is more common in commercial fisheries and in the shellfish industry because they believe it provides a hedge against escapement and breeding with wild populations. Triploid fish and shellfish are sterile, so even if they did escape into the wild, they would be unlikely to breed with the wild type populations.

But Mother Nature always finds a way, doesn’t she? There have several cases of reversion to diploidy in triploid oysters. These shellfish are then free to breed with wild species. And what is more, induction of triploidy is not 100% efficient in fish, so some organisms will remain diploid. The incomplete induction of triploidy has been illustrated brilliantly by the invasion of the asian carp.

What we call the asian carp is actually four different

species. But they all get big. O.K. usually not this big!

They are moving up the Mississippi River and

threaten to enter the Great Lakes. If they do, they could

destroy a multibillion dollar a year fishing industry.

What we refer to as asian carp is actually a mix of four species, the bighead carp, the black carp, the grass carp, and the silver carp. The grass carp was introduced into Geogria from China and the USSR in the 1960’s as a way to control overgrowth of grass and weeds in local ponds – that's what they eat. The interested parties did consider escapement and breeding, so they instituted a program of triploid induction. However, since some eggs escaped triploid development and some fish escaped the ponds, they became invasive. In the late 1980’s a program was introduced to assure that all released fish were triploid, but by that time the damage was done.

The bighead carp and silver carp were introduced into the US in sewage treatment plants and aquaculture ponds as a way to produce clearer water. These two species eat zooplankton and the waste of other animals, so naturally they were a good choice to improve water quality. But as with the grass carp, they ended up in the Mississippi and now have become a great problem. As far as I can tell, bighead carp and silver carp were not required to be tested as triploid before release until 2005 or later.

Nobody wants to bite into their striped bass fillet and find a

yellow grub. Black carp were introduced to destroy the

snails that serve as one life cycle stages of the grub. My Gosh!

Did this guy actually circle the grub with his wedding ring?!

The black carp was also introduced to help aquaculture farms. In raising striped bass, the yellow grub had become a major problem. The grub arrives in the waste of wading birds and can then wreak havoc by multiplying in snails and then attacking the striped bass fry. They burrow into the fish and cause large cysts to form. You can’t sell a food fish that releases a worm when you cut into it.

Black carp love snails, so they were introduced into fish farm ponds in the 1990’s to interrupt the yellow grub life cycle. The plan worked, and worked well; too bad the work to induce sterility did not work as well - the black carp has ended up in the Mississippi as have the other species of asian carp.

The result of these escapes is that rivers in 23 states are choked with asian carp, to the point that many native fish die off. True - the fish are big, very big, so they could provide a source of food. But they haven’t caught on as a food fish, and some places (like Canada) won’t even allow them to be sold for food. Fishing them for sport isn’t going to work as well, as their diets don’t help. How would you bait a hook with a piece of grass or a zooplankton?

The numbers have grown so large in recent years that other problems have developed. The silver carp has a strange habit of leaping out of the water when a boat motor approaches; there have been hundreds of instances where people have been struck by the fish. Noses have been broken, boats have been damaged, and this is all on top of losing the native species in the rivers. Check out this video of the silver carp problem and the birth of a new sport, aerial bowfishing.

Jeff Goldblum said it in Jurassic Park – life finds a

way. There is no way of predicting which turn life

will take, or the lengths evolution will go to help

life persist in the face of huge obstacles. Too often,

we are that obstacle.

There are no exceptions to two rules of nature: one - life will find a way to exist in every form you can imagine and using strategies that you can’t even imagine; and two – altering nature through anything other than natural selection is going to have unintended consequences. Thus, polyploidy is a strategy that fish have employed to diversify and fill niches, and polyploidy used by humans has been both a benefit and a bane.

But we haven’t even talked about one of the most interesting exceptions in nature that is related to polyploid development; the link between extra sets of chromosomes and the abandonment of sexual reproduction. To illuminate this exception, we will focus on the insect, lizard and amphibian polyploids next time.

Ashfield, T., Egan, A., Pfeil, B., Chen, N., Podicheti, R., Ratnaparkhe, M., Ameline-Torregrosa, C., Denny, R., Cannon, S., Doyle, J., Geffroy, V., Roe, B., Saghai Maroof, M., Young, N., & Innes, R. (2012). Evolution of a Complex Disease Resistance Gene Cluster in Diploid Phaseolus and Tetraploid Glycine PLANT PHYSIOLOGY, 159 (1), 336-354 DOI: 10.1104/pp.112.195040

King, K., Seppala, O., & Neiman, M. (2012). Is more better? Polyploidy and parasite resistance Biology Letters, 8 (4), 598-600 DOI: 10.1098/rsbl.2011.1152

For more information and classroom activities, see:

Polyploidy in aquaculture –

http://www.sciencedirect.com/science/article/pii/S0044848609004104

http://www.nature.com/scitable/content/the-advantages-and-disadvantages-of-being-polyploid-1554633

http://books.google.com/books?id=UaT8gZaOA04C&pg=PA216&lpg=PA216&dq=polyploidy+aquaculture&source=bl&ots=Y4vQHYI5I1&sig=2l9WIstm3A1E_KNGtBQdS1kKRI0&hl=en&sa=X&ei=91f0UM_gEcfg0gGnvYDwCA&ved=0CD0Q6AEwAg#v=onepage&q=polyploidy%20aquaculture&f=false

http://theoystersmyworld.com/tag/polyploidy/

http://www.thefishsite.com/articles/869/sterile-salmon-a-sustainable-future

http://www.organicconsumers.org/Patent/frankenfish0302.cfm

http://www.kokaneepower.org/projects/projects_salmon_triploid.php

http://www.foodsafetynews.com/2010/09/frankenfish-created-with-creepy-science/#.UPRY2YVf13I

http://www.imr.no/nyhetsarkiv/2011/mai/gode_resultat_med_steril_oppdrettslaks/en

Asian carp –

http://www.ideastream.org/lsi/enemy

http://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=4&ved=0CEEQFjAD&url=http%3A%2F%2Fhcpssenhancingalgebra2.wikispaces.com%2Ffile%2Fview%2FAsian%2BCarp%2BInvasion%2BLesson%2BPlan.doc&ei=9Vv0UNbJFtGF0QHo_YDYCg&usg=AFQjCNHhLw_I1Yy5YIJkOozHm1uhqXdxHA&bvm=bv.1357700187,d.dmg

http://www.scholastic.com/teachers/article/invasion-asian-carp

http://www.nwf.org/Wildlife/Threats-to-Wildlife/Invasive-Species/Asian-Carp.aspx

http://www.glfc.org/fishmgmt/carp.php

http://www.watershedcouncil.org/learn/aquatic%20invasive%20species/asian-carp/

http://asiancarp.us/

http://oklahomainvasivespecies.okstate.edu/silver_carp.html

http://mdc.mo.gov/discover-nature/field-guide/silver-carp

http://www.carpbusters.com/profile_black.shtml

http://deltafarmpress.com/black-carp-threat-aquaculture

http://www.ecy.wa.gov/programs/wq/plants/management/aqua024.html

Wednesday, January 23, 2013

An Evolutionary Ploy Employing Polyploidy

Biology concepts – polyploidy, autopolyploidy, allopolyploidy, gigas effect, heterosis

The Sixth Day was a cheesy science fiction thriller

about cloning. But plants do have different versions

of themselves in many cases, produced not by

cloning, but by polyploidization. I have no idea

what cloning Arnold had to do with the funky

lights in the eyes; maybe it is a vitamin D thing.

Imagine that there are three different versions of you, each with different strengths and weaknesses, living in different places and surviving in different ways. Sounds like a strange Arnold Schwarzenegger sci-fi movie; maybe one version of you has really big muscles and an accent.

For some organisms, this isn’t science fiction, it is science fact. In the last two weeks we discussed how one mammal manages to survive while being polyploidy in all its cells. We have also discussed how our bodies have discrete sets of polyploidy cell types. While these cells are crucial for human development, they are tightly regulated; indiscriminate polyploidy in humans is deadly- it's called cancer.

Now we can talk about whole groups of organisms that use polyploidy as a key to their evolution. Not only can they survive as polyploidy beings, they thrive on it.

A study from late 2012 highlights the importance of polyploidy in plants. It turns out that plants can tolerate being polyploid much better than most animals can. In fact, being polyploid is the reason for much of their success in colonizing different habitats.

The researchers in the 2012 study were looking at a plant called Atriplex canescens, a drought resistant shrub that lives in the Chihuahuan Desert of the American Southwest. A. canescens has three versions of itself, called cytotypes. One is diploid in all its cells (except the ovule and pollen sperm of course). Another is tetraploid (4n), and the third is hexaploid (6n). It turns out that each cytotype lives in a slightly different habitat in the desert, depending on how much water is available.

The hexaploid version lives in the clay, the type of soil that is most likely to be water-poor. The diploid cytotype lives in the sandy soil nearest the regular sources of water, and the 4n shrub lives in between. Therefore, it was hypothesized that the different ploidys result in different physiologic and structural characteristics. This turns out to be so.

When plants have more than two copies of each chromosome, it changes the structures of their leaves and stems. Polyploid plants tend to have larger, but less densely packed pores in their leaves. We talked about these pores, called stomata, in an earlier post. They are responsible for releasing water and oxygen to the outside world. This regulates the movement of water in the plant. As more water evaporates from the stomata, more is drawn up from the roots by negative pressure, called transpiration.

Embolisms are bad for plants in the xylem and for human in the

arteries. Divers have to ascend slowly from deep dive so that

the gases in their blood has time to adjust to the pressure

change. If the rise too quickly, the gases come out of

solution and forms bubbles in their vessels. This is called

decompression sickness or the bends, and it can kill.

Polyploid plants also tend to have thicker epidermis layers on their leaves, and this, together with the lower density of stomata means that polyploid plants tend to lose less water than diploid version of the same species. That could be helpful in low water environments.

Polyploid plants also have changes in their xylem. The xylem is the vessel-like tissue that moves sugars and nutrients throughout the plant. In time of drought, low water levels can cause an air pocket to form in the xylem. This stops the xylem flow, much like an air or solid object embolus can stop the flow of blood when it gets stuck in a blood vessel. You wonder why the nurse takes such care to remove the air from the syringe when she gives you a shot? An air bubble getting stuck in an artery in your heart, lung, or brain could very well kill you.

Emboli formation is less likely in polyploid xylem, because the channels are bigger. This is good for safety and remaining alive in drought conditions, but it is not good for growing fast when more water is available. Therefore, the diploid versions of a species are more likely to live where there is more water, and the polyploid versions where there is less water.

This is exactly what the researchers found out. The hexaploid cytotype had the high measured water resistance, with the largest stomata, thickest leaves and widest xylem channels. The opposite was true for the diploid version, and the 4n cytotype was in the middle. Therefore, they show that water conservation and movement is different in the different ploidy plants and this accounts for their different habitats.

The gigas effect isn’t just seen in the watermelon fruit.

The leaves and flowers of the tetraploid version (on

the right) or bigger than those of the diploid watermelon

plant. More DNA means a bigger nucleus, and a bigger

nucleus needs a bigger cell. So all the structures get

bigger as well.

One species being able to live in several habitats is quite the evolutionary advantage. They don’t compete with one another and they can colonize a larger portion of the land. Being polyploid is quite the boon for some plants.

The advantages all seem to come from size; bigger stomata, thicker epidermal cells, wider xylem. If a cell has more DNA to house, the cell is necessarily going to be bigger. This leads to the bigger plant structures, and their size leads to less water loss. If the conditions arise where water is not available in a certain area, these characteristics will be advantageous and selected for by evolution.

But larger cells are supposed to be one of the disadvantages of polyploidization. Called the gigas effect, larger cells leads to higher energy needs and altered surface area to volume ratios. These change can inhibit interactions between the plasma membrane proteins and cytoplasmic elements can be disadvantageous, even lethal. However, for some things in plants, like fruits, huge increases in DNA, up to 126n or more work just fine.

Do you like watermelon? More watermelon is better then, right? Melons grow large because of the gigas effect. Many watermelon species are triploid or higher. The strawberries that come coated in chocolate and are as big as your palm are very likely to be octaploid (8n).

Autopolyploids can arise from genome duplications, or from

hybridizations between a diploid gamete and a haploid

gamete, with later stabilization of the genome by

polyploidization. But all the genes come from one version of

the organism. Allopolyploidization (on the right) come from

hybridization of two different organisms, often with a sterile

first generation (F1), and polyploidization to return fertility.

This plant has several copies of different genes, so it has quite

the chance to become a new species.

Many crops are polyploid, the results of hybridizations and crosses over many years. These crosses have been meant to increase yields reduce disease susceptibility and expand the environments in which the crops can be grown. For hybrids of two different species, this is called allopolyploidy (allo = different). Using this method, we have developed strong wheat (hexaploid), apple (tetraploid), cotton (tetraploid), and sugar cane (octaploid) crops.

Many crop hybrids are often sterile in first generation, especially if they come about from autopolyploidy hybridizations. “Auto” means same, so these are crosses between variants of the same species, and are often associated with endoreplication events (see When Too Much Is Just Enough) giving a diploid gamete mating with a haploid gamete to give a triploid organism. Triploids are often sterile. This is how you have things like seedless watermelons and you know those little black dots in your banana, those are the undeveloped seeds. You have to propagate these plants by cuttings (called vegetative reproduction), not by seeds.

When you induce polyploidy in the triploid hybrids, they become fertile again, and they (and allopolyploids) also display another feature, called heterosis, also known as hybridization vigor. This heterosis is another reason why most of the cash crops of the world are polyploid. While the crosses are meant to alter traits, the resulting polyploidization increases heartiness. Still think GM crops are a bad idea – you’ve been eating them your entire life.

When plants undergo polyploidization, they have more

copies of each gene, called redundancy. This represented

by the green circles, only showing two here for convenience.

The plant may get rid of some copies, as in the left panel, or

may compartmentalize some of the functions in each allele

(subfunctionalization, on the right). In other cases, some alleles

may drift genetically, until they have new functions –

neofunctionalization, as show in the middle panel. New functions

could lead to a new species, if environmental changes make

them advantageous.

But heterosis could also have unwanted results. In the late 1800’s, hybrids of different spartina bush species were carried out in England in hopes of breeding a species that would better prevent erosion of the tidal mud flats. It turned out that the offspring underwent allopolyploidization and became too strong a species. The new species, Spartina anglica, underwent significant and rapid genetic changes and became invasive in salt marshes. It can crowd out other species and can grow dense enough to prevent some animals from moving from land to water.

The new talents of S. anglica are related to its polyploidization. When plants become polyploid, they may have lots of DNA with the same functions; therefore they tend to try and reduce their genetic load. This can occur by getting rid of some gene copies, or letting mutations run wild in some alleles, as others will still be around to perform the needed function.

This can lead to subfunctionalization (altered functions) or neofunctionalization (new functions) in the changing genes. New functions + change in environment can lead to new species, ie, speciation. Speciation due to polyploidy is apparent in 15% of angiosperms and 31% of ferns. In fact, 40-100% of flowering plants have some polyploidy in their past.

The sweet corn in your low country boil is a direct

effect of polyploidization. The sucrose produced in

polyploids is higher than in diploid corn, so

naturally we can’t get enough of the polyploidy

versions of corn. Sweet corn with shrimp, sausage,

and potatoes – can’t beat it.

But not every polyploid development is so simple. Sometimes the new cytotypes cannot quite overcome the problems inherent in having many more copies of genes all working at once. In corn for instance, some polyploid numbers are better tolerated than others. In 1996, Guo, the same primary researcher involved in the Atriplex work cited above, was working on haploid, diploid, triploid, and tetraploid versions of maize. He found that some gene products (proteins) did increase with increasing gene copy number, but others didn’t.

For example, sucrose synthase levels were twice as high in the 4n version as in the 2n version of maize as expected, but mRNA levels were 3x higher in the haploid plants and 6x higher in the triploid versions! Obviously, some regulatory pathways were not controlled as well at some of the polyploidy levels. In these plants, fully 10% of the genes had an “odd-ploidy” effect. This leads to less than stable cytotypes and poor endurance in the environment.

Next time, we will see that fish are one of the exceptions of the animal world. They tolerate polyploidy well, and we have even used that fact to increase our harvests, but also our headaches.

Hao GY, Lucero ME, Sanderson SC, Zacharias EH, Holbrook NM. (2012). Polyploidy enhances the occupation of heterogeneous environments through hydraulic related trade-offs in Atriplex canescens (Chenopodiaceae). New Phytol.

SALMON, A., AINOUCHE, M., & WENDEL, J. (2005). Genetic and epigenetic consequences of recent hybridization and polyploidy in Spartina (Poaceae) Molecular Ecology, 14 (4), 1163-1175 DOI: 10.1111/j.1365-294X.2005.02488.x

Guo M, Davis D, Birchler JA. (1996). Dosage effects on gene expression in a maize ploidy series Trends in Genetics, 12 (8) DOI: 10.1016/0168-9525(96)81463-6

For more information or classroom activities, see:

Polyploidy in angiosperms –

http://www.vrml.k12.la.us/7th/7SC_By_Unit/unit6/7SC_Un6Act6.htm

http://evolution.berkeley.edu/evolibrary/article/speciationplants_01

http://projects.coe.uga.edu/plantbreeding/index.php?title=5.Polyploidy

http://books.google.com/books?id=qoKahqZ2E0gC&pg=PA60&lpg=PA60&dq=%22gigas+effect%22&source=bl&ots=1--rHb7KiH&sig=ZwfKhGmut8X_K3Aj-UYLiedl0ME&hl=en&sa=X&ei=6KHoUNfNA8jCrQGK8YFY&ved=0CHUQ6AEwCTgK#v=onepage&q=%22gigas%20effect%22&f=false

http://www.nature.com/scitable/topicpage/polyploidy-1552814

http://milkweedgenome.org/?q=system/files/Straub_CV_2011.pdf

http://www.as.wvu.edu/~kgarbutt/QuantGen/Gen535_2_2004/Polyploidy.html

Polyploidy in crop plants –

http://www.biologie.uni-hamburg.de/b-online/e12/t1.htm

http://128.192.141.98/CottonFiber/pages/outreach/professional.aspx

www.eeob.iastate.edu/.../Udall&Wendel_CropScienceSupp_2006.pdf

http://www.sciencedaily.com/releases/2012/01/120107151855.htm

http://potatogenome.berkeley.edu/nsf5/potato_biology/polyploidy.php

http://lifeofplant.blogspot.com/2011/02/polyploidy-and-aneuploidy.html

Autopolyploidy and allopolyploidy –

http://www.youtube.com/watch?v=49pVJRkvUAs

http://ezinearticles.com/?Types-of-Polyploidy---Autopolyploidy-and-Allopolyploid&id=5316974

http://erdmannevolution.blogspot.com/2010/04/allopolyploid-and-autopolyploid.html

http://wps.prenhall.com/wps/media/objects/1143/1170856/Chapter_07/Present/Activities/7_2/7_2_1a.swf

http://books.google.com/books?id=yegDAAAAMBAJ&pg=PT87&lpg=PT87&dq=autopolyploidy+allopolyploidy&source=bl&ots=k1k8_24g_a&sig=oQGjfn-Yrcx2Nlt9iv3y6K43m6Y&hl=en&sa=X&ei=H6PoULuNEcj1rAHt9YCAAQ&ved=0CIYBEOgBMA0

Wednesday, January 16, 2013

When Too Much Is Just Enough

Biological concepts – endoreplication, endocycling, endomitosis, decidualization, trophoblast, megakaryocyte

Last week we learned that polyploidy plays a role in cancer development and is the number one cause of spontaneous abortions in humans. Polyploidy is just no darn good.

There’s alot to fret about once you hit the

atmosphere. But take heart, you’ve already found

a way to make a cancer-like pathway work for you.

Don’t worry about the details, you’ll get it all in

Biology class.

But what if I told you that this same evil process is crucial for the birth of every baby that has ever come kicking and screaming into this cold, cruel world? Without some very specific polyploid cells, none of us would be here. Many of the signaling pathways that contribute to cancer polyploidy also function in normal development, although they are dysregulated in the former and tightly regulated in the latter.

For example, osteoclasts (osteo = bone, and clast = to break) form from the fusion of two or more precursor cells. Since each precursor cell has its own nucleus with a 2n set of chromosomes (n=23 for humans), the fused cell may have 4n, 6n, 8n, or more chromosomes, in one or more nuclei. New evidence shows that not only can they fuse, but they can also fission to form more osteoclasts when needed. This had not even been hinted at before.

Osteoclasts eat bone; you are forever tearing down bone and replacing it with new bone. If you lift weights and build bigger muscles, you need bigger bones onto which you can attach your now stupendous guns. About every ten years or so, you have an entirely new skeleton!

Polyploid cells can be formed when diploid cells fuse, but it is more interesting when they are formed by the processes of endoreplication (endo = within). Normally, most cells just hum along, growing (G1), then replicating their DNA (S), then growing some more (G2), and finally dividing into two daughter cells by mitosis (M). The two new cells then repeat the process. This is called the cell cycle, and is abbreviated as G1, S, G2, M.

The mitosis portion of the cell cycle itself has several parts that we all learned in biology class – shout them out with me - prophase, metaphase, anaphase, and telophase! At the end of telophase, the two daughter cells finally decide they can’t be roommates any longer, and they divide up their belongings.

The phases of mitosis finish up by dividing the cytoplasm and

nucleoplasm. You can see that the cytokinesis starts first, with

the appearance of the cleavage furrow, but karyokinesis is

completed before cytokinesis is done. Therefore, you can’t have

complete cytokinesis with defective or incomplete karyokinesis.

The replicated chromosomes (each having two sister chromatids) had already separated in anaphase, so now the rest of the nuclear contents split in half and a nuclear membrane forms around each new nucleus – this is termed karyokinesis (karyo = nuclear, and kinesis = in motion). The last thing they do is divide up their cytoplasmic contents and pinch off a new membrane between the two of them, becoming two separate cells – cytokinesis.

In endoreplication, one or both of these processes is turned off, so the two daughter cells continue to share a room, but now the room has twice as much DNA (4n instead of 2n). The cell skips at least a portion of M phase, and the cell cycle becomes G1, S, G2 ----G1, S, G2, etc. It may occur just once, producing a tetraploid cell, or it may occur several times, forming huge cells with 32n or more chromosome sets.

If the cell skips mitosis all together, the process is called endocyling. In this case, the chromatids don’t separate in anaphase, and you end up with chromatids that remain stuck together at their centromeres. If they replicate again in the next S phase, you end up with an octopus-looking chromosome with several arms sticking out – called a polytene chromosome.

The left side of the cartoon shows endocycling.

Skipping mitosis altogether keeps the chromatids

connected and forms polytene chromosomes.

Endomitosis is on the right, where the cell goes

through part of mitosis, then skips the part where

the two cells separate, either by skipping

cytokinesis alone or karyokinesis and cytokinesis.

Polytene chromosomes occur naturally in some animals, like the huge (1 mm) chromosomes in the salivary glands of larval fruit flies (Drosophila melanogaster). They benefit the fruit fly larva in that the cells can produce more proteins from the many copies of the genes. This allows the fruit fly larva to make enough of the proteins that are important in forming the pupal case when it undergoes metamorphosis. All due to endocycling and polyploid formation.

On the other hand, if a cell starts through mitosis and separates its chromatids, AND THEN decides to not divide, this is called endomitosis. Cells that have undergone endomitosis have many sets of chromosomes. Endomitosis without cytokinesis results in large cells with multiple diploid nuclei because karyokinesis separated the nuclei. Endomitosis without karyokinesis and cytokinesis results in large cells with a single polyploid nucleus. You can see that polyploidy would need to be highly regulated to keep it from getting out of control.

So how is that polypoloidy is crucial for our survival? It turns out that that some specialized cells of the embryo undergo polyploidization as the embryo implants into the wall of the uterus.

The embryo has an outer layer of cells called the trophoblast; these cells become the placenta, attach the embryo to the uterine wall, and create the blood vessel connection between mama and junior. The trophoblast is the first set of cells to differentiate in the embryo and they become several different types of trophoblasts.

One type in particular, the extravillous cytotrophoblasts (ECTs), spread out from the developing placenta and burrow into the uterine wall. This creates the tight attachment between mom and embryo. The ECTs also send out hormones to rearrange the mother’s blood vessels, forming the umbilical cord and vessels. This is how the growing baby gets all its nourishment until delivery.

ECTs have been studied most in rodents; they weren’t recognized in humans until just recently. However, a late 2012 study has shown that ECTs are released from the placenta and can be studied by collecting them at the cervix. The cells were sufficient to determine the sex of the child after only 5 weeks of gestation, and were generally of 4n-8n ploidy. Interestingly, female fetuses tended to form ECTs at a rate almost 7x higher than male fetuses – you’re guess is as good as mine as to why that might be.

In panel A there is a bunch of abbreviations. VT is the villous

trophoblasts that make the connection to the decidua (DD).

In the black box which is enlarged in panel B, you can see the

extravillous cytotrophoblast (EVT here) cells invading the

decidua. Both the EVT and the decidua are polyploid.

The most amazing thing about the polyploid trophoblast cells is that they also regulate polyploidization of some of the cells of mom’s uterus. When the endometrium of the uterus prepares to accept the fertilized egg, it undergoes several changes that are together referred to as decidualization. Differentiation of stromal cells into decidual cells and other cellular differentiations make the uterus able to support embryonic growth.

The reason that cells of the decidua must be polyploid is unknown, but the fact that polyploidization begins at the point of implantation and spreads to a greater part of the uterus tells you that they are necessary. A new study points to a few possible reasons. Comparing polyploid decidua to non-polyploid decidua showed that many genes were up-regulated or down-regulated.

The up-regulated genes had to do with metabolism, especially the mitochondrial energy production. On the other hand, down-regulated genes had to do with apopotosis and immune function. These results suggest that polyploidization of the decidua is meant to increase cell functions for the benefit of the embryo, and this takes energy (so more mitochondrial function), while at the same time making sure the cells survive to support the fetus until delivery (reduced apoptosis gene function) and protection of the fetus from the mother’s immune system (the baby is a foreign body after all).

So baby has polyploid cells that mediate joining with the mother, and mom has polyploid cells that also work in the formation of the link between the two. Everyone has to bring polyploidy to the party, or ain’t nobody getting born!

However, polyploidy in fetal development is only part of the story. You don’t abandon polyploid cells altogether once you are born or give birth. All of us have polyploid cells in our bodies right now. Take megakaryocytes for instance.

When you cut yourself, or there is a leak in a blood vessel, platelets arrive to help close the hole and stop the bleeding. Platelets are of irregular shape and are sticky, so they tend to get stuck along the edges of broken blood vessels. Then other things stick to them, a few dozen enzymatic reactions take place with myriad proteins, you form a clot (called a thrombus in the medical world).

I have described how platelets are important for coagulation.

This cartoon shows a break in the cell on the bottom, and

ALL THE STUFF that has to happen to forma thrombus (clot).

Platelets are central, but they are certainly not all the story.

Platelets are not cells, they are actually just fragments of megakaryocytes that pinch off and travel around in the circulation looking for holes to plug. A healthy adult will produce 100 billion platelets each day! Megakaryocytes can afford to give away lots of cytoplasm and membrane because they are large. And they are large because they are polyploid, with lobulated nuclei due to incomplete karyokinesis and no cytokinesis.

Hepatocytes (liver cells), smooth muscle cells in blood vessels, heart muscle cells – these can all be polyploid. In hepatocytes, polyploidization occurs in cells that are done dividing and specializing (terminally differentiated) and are now just doing their job. Fetal and newborn liver cells are exclusively diploid, but 30-40% of adult hepatocytes are polyploid.

Polyploidy may be a way to increase liver metabolism and function without going through cell division. Or it may help to protect the cell from the effects of individual mutations. Since the liver is involved in breaking down toxins, it’s a good guess that some genes will mutate. Having extra copies around would prevent a mutation from inhibiting cell function. One mutated gene can be compensated for by an additional normal gene.

On the other hand, smooth muscle cells seem to undergo polyploidization as a prerequisite to senescence; they are aged and they just stop working. This interesting, since we said last week that cancer cells are more likely escape therapy induced senescence by becoming polyploid. Once again, biology can turn the ordinary on its head.

We have discussed the appearance of a polyploid mammal and crucial sets of polyploid cells in humans. These are the exceptions in higher vertebrates. But in other organisms, polyploidy is a key to evolution. Next time we’ll talk about the exceptional role of polyploidy in the development of plants.

Biron-Shental, T., Fejgin, M., Sifakis, S., Liberman, M., Antsaklis, A., & Amiel, A. (2012). Endoreduplication in cervical trophoblast cells from normal pregnancies Journal of Maternal-Fetal and Neonatal Medicine, 25 (12), 2625-2628 DOI: 10.3109/14767058.2012.717999

Ma, X., Gao, F., Rusie, A., Hemingway, J., Ostmann, A., Sroga, J., Jegga, A., & Das, S. (2011). Decidual Cell Polyploidization Necessitates Mitochondrial Activity PLoS ONE, 6 (10) DOI: 10.1371/journal.pone.0026774

Jansen, I., Vermeer, J., Bloemen, V., Stap, J., & Everts, V. (2012). Osteoclast Fusion and Fission Calcified Tissue International, 90 (6), 515-522 DOI: 10.1007/s00223-012-9600-y

For more information or classroom activities, see:

Osteoclasts and bone remodeling –

http://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=13&ved=0CHoQFjAM&url=http%3A%2F%2Fteachhealthk-12.uthscsa.edu%2Fcurriculum%2Fbones%2Fpa12pdf%2F1203D-all.pdf&ei=gNDPUKddqOTLAfOggYAD&usg=AFQjCNHwwn6yU3j4aoRDM1xr6CtGKsSgbQ&bvm=bv.1355534169,d.aWc

http://www.youtube.com/watch?v=78RBpWSOl08

http://www.rndsystems.com/cb_detail_objectname_FA04_OsteoclastCoStimulation.aspx

http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/B/Bone.html

http://www.youtube.com/watch?v=0dV1Bwe2v6c

http://depts.washington.edu/bonebio/ASBMRed/growth.html

Endoreplication –

http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/E/Endoreplication.html

http://www.news-medical.net/news/20111031/Researchers-shed-light-on-inner-workings-of-endocycle.aspx

Trophoblast and decidualization–