On BioScience and Life and Such

Archive for February, 2008|Monthly archive page

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Tests that make me sad (updated)

In Uncategorized on February 29, 2008 at 10:46 am

In a previous post (“Clarifying misuse of Science“) I expressed concerns over Prenatal testing for familial hypercholesterolemia. Now it seems I should have included two more (and in my mind, – more controversial) tests approved in the UK for preimplantation testing preceding IVF. These are tests for breast cancer (BRCA1) and early onset Alzheimer.

Especially the case of BRCA1 worries me. True, the prevention (mastectomy) is a horrible ordeal for the patient, but is this sufficient grounds for excluding eggs for IVF ? Can’t you live a reasonably happy and normal life after such a procedure (see breast reconstruction) ?

Why haven’t there been more fuzz around this ? Is everyone going to be using prestested eggs in IVF now ? And what are the limits on acceptable tests, will testing for athletic performance become an exclusion criterion soon ?

I must have been naive to so strongly oppose the slippery slope argument in genetic testing discussions up until now. I wish we could restrain ourselves a bit more, but fear that we can’t.

The solution is to come up with treatments for most of these conditions (although treating athletic performance may constitute a problem in terms of the number of people needing treatment as well as treatment alternatives…..).

Treatment alternatives need to appear soon if we’re to avoid the brave new world future that critics of the genetics era have been promoting …..

The “find a cure” process however, is not going to be fast and in the 10-25 years to follow, those concerned couples that have the option of BRCA1 and Alzheimer testing will most likely not take the chance that a treatment will be found in time for their child to be cured…….., – and opt for the safer pretested egg and IVF.

Seeing that the slope is becoming slippery I have decided not to argue for genetic testing any further, and if this trend continues, – argue against, at least until treatment options appear (the next 25? years) or a sensible limit for testing is drawn (right now, – please).

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On race and genetics

In Uncategorized on February 12, 2008 at 3:10 pm

In the previous post “A refreshing view on James Watson“, I referred to the website HonestThinking. The background was a post on this website pointing out the lack of scientific evidence for the unison discreditation of Watson. This has spurred a debate in some (local) newspapers about the validity of the term “race”. Race it is argued, is a social construct used to stigmatize groups. In addition there are claims that there isn’t any biological/genetic evidence to indicate that we are more different between races than within a race. Race is clearly then, a controversial term.

HonestThinking has replied to, and rejected, these arguments elegantly and has referred to the brilliant website Edge. Edge has a piece written by Mark Pagel on this issue which is very worth reading (see quote below).

As a reminder that knowledge is key to avoid prejudice, enlightened biologists do not view race as controversial. We are familiar with genetic diversity. We know that there are significant differences between ethnic groups. Our community does not associate these genetic differences with political or racist issues. James Watson on the other hand, did, – and that was hopefully, the real reason for his job-suspension. Our challenge is to communicate the scientific truth in a manner exactly opposite to Dr. Watson’s.

The scientific truth is that there are genetic differences with biological implications. These differences are larger than we previously thought. Not only are there differences in protein encoding regions of DNA, there are also differences leading to differential gene expression and in addition, there are differences in genetic insertions/deletions in the human genome. Add to this a variation in gene copy number and the multitude of ways we can be different at the genetic level becomes apparent. Altogether the 99,9 % genetic similarity we thought existed between humans is probably much lower.

On top of the genetic differences there are epigenetic differences that can contribute to phenotype variation (difference in appearance). Thus, we are all substantially different, but that does not mean we are of unequal value. It just means we’re not the same, – let’s be thankful and appreciative of that. I know I am.

Ending quote from Mark Pagel in Edge’s “The World Question Center”:

“What this all means is that, like it or not, there may be many genetic differences among human populations — including differences that may even correspond to old categories of ‘race’ — that are real differences in the sense of making one group better than another at responding to some particular environmental problem. This in no way says one group is in general ‘superior’ to another, or that one group should be preferred over another. But it warns us that we must be prepared to discuss genetic differences among human populations.”

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Hsp90 to end controversies in evolution theory (final chapter, blogging in Just Science 08)

In Uncategorized on February 8, 2008 at 9:42 am

Previous posts have shown Hsp90 to be a molecular buffer allowing rapid morphological changes in times of stress. As will be discussed below, such a buffering function supports the evolutionary theories of punctuated equilibria, hopeful monsters and canalization.

So…, this last post will end with the final conclusions based on the arguments presented in the previous 4 posts. But, first….Two fundamental questions:

1. Even if Hsp90 can promote rapid changes in phenotype (appearance) how is this change retained (fixed) for future generations ?

This fixation has been demonstrated to occur (see Sangster TA et al.), and the traits become independent of Hsp90. The exact mechanism(s) however remains to be elucidated.

Nevertheless, temporarily compromising Hsp90 function (either by drugs or by temperature rise) is sufficient for fixing new traits. Simulations seem to show that knocking out the genes for key proteins (not necessarily heat shock proteins) lead to increased phenotypic diversity, and thus the underlying cause may be genetic fixation. However, interplay between epigenetic and genetic mechanisms has been suggested and been backed up by experiments. Thus fixation probably happens through yet to determined genetic as well as epigenetic mechanisms, or a combination of both. A model for epigenetic fixation is given in the thumbnail below:

Epigenetic evolution through Hsp90

Models for genetic fixation follows the theory of canalization with Hsp90 functioning as the Waddington’s widget (see Semin Cell Dev Biol. 2003 Oct;14(5):301-10). This is discussed further under the next bulletpoint, the second question…..

2. Does these aspects of Hsp90-function fit into current models of evolution ?

Yes, although some of these theories are controversial. First we have the idea of punctuated equilibrium and hopeful monsters discussed in my previous post. To expand on these ideas let’s also include the theory of canalization. Canalization explains punctuated equilibrium by referring to an organisms buffering capacity (to counter the potential deleterious effects of mutations). The theory was put forward by C. H. Waddington more than 50 years ago, but is still controversial it seems. Hsp90 is a molecular explanation of the canalization concept in that organisms with different genotypes express the same phenotype until times of stress. There are also indications that other heat shock proteins or other “signaling hub”-proteins or even miRNA can serve such buffering functions (see references within this review).

Taken together, these controversial evolutionary theories and the experimental evidence on Hsp90 supports one another, and a paradigm shift in evolutionary biology is in place. Darwins theories are correct up to the point of gradual and constant evolution of traits. Evolution instead, occurs in bursts. This series of blogposts have conveyed the molecular evidence for such punctuated equlibria and canalization, which comes from studies on the molecular chaperone Hsp90. I hope I have enlightened and convinced at least some evolution biologists into believing that Darwins theories can be expanded to include these (no longer controversial) theories.

There are however, a lot to work out in terms of the underlying molecular mechanisms for Hsp90 (and/or other buffering bioactive molecules ?) in canalization. To end this blogpost-series I will therefore quote the closing remarks from Salathia N and Queitsch C‘s review in 2007:

“Clearly, organisms have succeeded in integrating multiple canalization mechanisms into robust wild-type phenotypes which can respond appropriately to environmental perturbations and evolve new shapes and functions over time. Now it is up to us to determine how molecules as diverse as a molecular chaperone, chromatin remodeling proteins, and the RNAi machinery interact coherently to achieve such synergy, a truly fascinating and worthy field of future inquiry.”

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Evidence for Hsp90 involvement in rapid evolution of new traits (chapter IV, blogging in Just Science 08)

In Uncategorized on February 7, 2008 at 10:39 am

Previous posts have attempted to demonstrate that there is a potential role for (Heat Shock) proteins that mask mutations, to enable rapid evolutionary changes. The Hsp90 protein has been presented and the basic problems one face to explain bursts in evolution have been outlined. Now the time has come to show real examples of Hsp90 influencing the evolution of traits.

The following are very short summaries of key papers. For details, please see the referenced papers.

1. Hsp90 and Cancer

In 1993 Yang Xu and Susan Lindquist showed that Hsp90 associates with v-src and inhibits its activity in an concentration-dependent manner. Hsp90 was not merely an on and off switch for v-src, but exhibited transient inhibition, dependent on intracellular concentration of Hsp90. This was a clue to understanding Hsp90’s role in cancer (as well as in evolution). After this, many cancer-related proteins have been identified that interact with Hsp90 (see table).

Table from a review in Nature 2005 by Whitesell and Lindquisthsp90-table-2005-review.jpg

The mechanism one speculates, is similar at the molecular level, to the mechanisms postulated for morphological change. Hsp90 stabilizes the otherwise unstable oncogenic proteins, to aid in tumor growth in an environment hostile to tumor development. In other words: the heat shock proteins protects the oncogenic cells from stress. When the tumor cells subsequently attain further mutations and protein alterations, inherent to oncogenic growth, the heat shock proteins are unable to stabilize all of the altered proteins and the tumor can progress into accelerated growth and/or metastasis. The role of Hsp90 in cancer development has been widely accepted and inhibitors of Hsp90 activity is currently undergoing clinical trials for cancer treatments.

2. Drosophila

The key paper on Hsp90 and Drosophila evolution is the Rutherford and Lindquist paper in Nature 1998. This paper has been mentioned on several previous posts here on SciPhu and also in the introductory Just Science post. Again, the take home message is that reducing levels of Hsp90-activity leads to changes in phenotype. The reason for such dramatic effects is probably that Hsp90 stabilizes proteins that are key elements in intracellular signaling pathways. Often these are kinases, phosphatases or transcription factors, see this table for full list. The common feature of these affected proteins is that they regulate the activity of other proteins downstream in the signaling cascade. Thus, changes in the activity of one master protein acts on the stability and function of many other “executive” proteins ultimately resulting in massive changes. The phenomenon has further been elucidated in other species……….

3. Yeast

In yeast, a reduction in Hsp90 levels potentiates drug resistance and this resistance has multigenic determinants working through Hsp90. Hsp90 thus helps yeast evolve to counter the stressful effect of the drug. Interestingly, this effect is diminished by temperature rise. Increasing the stress (by adding heat) therefore, titrates Hsp90 away from the drug-resistance and makes the yeast vulnerable again (could this effect explain why fever has developed ?).

4. Arabidopsis

Evidence for the same mechanisms occurring in plants comes mainly from two publications on Arabidopsis thaliana (Queitsch C et al. and Sangster TA et al.). These images from the latter publication show the extensive morphological changes seen in the plants.

journalpone0000648g002.png
Figure 2. Similar morphological phenotypes of seedlings with reduced HSP90 function by RNAi or pharmacological means (GDA). (a) and (b): purple pigment accumulation; (c) and (d): organ number defect; (e) and (f): narrowly-shaped deformed true leaves; (g) and (h): twisted rosettes; (i) and (j): lobed cotyledon. RNAi plants are T3 generation with from line RNAi-A3. Size bar 2 mm for b and g–i, 1 mm for a, c–f, and 3 mm for normal phenotype. (b) and (f) originally published in [5].

These effects can also be induced by increasing the temperature. Demonstrating the generality in the stress response. Since the genetics of these plants is easier to trace in these plants than in Drosophila or Yeast, the evidence for buffering genetic changes is even more clear-cut in this organism.

An excellent illustration to summarize Hsp90-buffering comes from Sangster TA et al.:

Hsp90-buffering

In the last post I will present published models on how Hsp90 can act in evolution, – welcome back for the last post in Just Science 08, tomorrow.

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How can chaperones act in evolution (chapter III, blogging in Just Science 08)

In Uncategorized on February 6, 2008 at 11:15 am

Moving away from the specifics of Hsp90 for a while, this post shall focus on the general principle of chaperones in evolution. What supports such an hypothesis:

To begin lets look at the problem of rapid evolution. Darwinian evolution is based on a constant rate of random mutations in the genome of any evolving organism. This implies that mutations happens constantly, by chance, regardless of the external environment. Adaptations consequently arise by chance. Evolution of new traits one may think, is therefore slow and gradual.

Gradual evolution

Gradual evolution (reproduced with permission from Dr. Dennis O’Neil)

Since such a view, does not fit with the bursts of evolution observed in fossil material, alternative explanations have been put forward (see more below). However, even when using a constant random rate of mutations one would expect “bursts” or rapid transitions. This is elegantly illustrated in this simulation of an evolving clockwork. Since a beneficial mutation can have a profound impact on fitness, then there should be no surprise that the transition between av less fit form and a more fit one, happens quickly. Thus even with a constant mutation rate one would probably not see a slow, gradual evolution of species, – basic math skills on exponential growth should make this clear. Why this notion of gradual evolution is prevailing I cannot understand.

In addition there are those that believe that the mutation rate may not be constant. Thus, with an increased mutation rate and rapid transitions one can start to explain the observed bursts of evolution.

Further explaining bursts of evolution we have the theory of ‘punctuated equilibria’, associated with Niles Eldredge and Stephen Jay Gould, which states that organisms go through short periods of rapid evolution from time to time, against a background of relative stasis (see picture below, and this genomicron post as a starting point for more on punctuated equilibrium ).

Punctuated equilibrium
Punctuated equilibrium (reproduced with permission from Dr. Dennis O’Neil, for his tutorials on evolution go here and here).

This has further led to the theory of hopeful monsters. These theories account for non-linear rapid evolution within the boundaries of Darwinian principles, but they have been heavily criticed. One of the main criticisms of these theories, as far as I can understand, is the improbability of a single mutation to give rise to radical morphological changes, and further that this change, if it happens, is most likely deleterious, and if it against all odds is beneficial, its even more improbable that this individual is able to produce offspring with the same trait(s).

So we are still left with some problems: External environment changes can happen really quickly. Is random mutation events, occurring at a slow rate (even if it’s sped up in larger populations or even if monster are hopeful in times of stress), sufficient to explain the effectiveness of adaptations seen in nature ? Does an organism rely on (slow) random mutations to evolve a trait to help the species adapt to the new environment, or are there additional mechanisms in place to speed up this mutation rate and perhaps guide mutation events towards selected genes that allows rapid changes in phenotypes ?

Enter heat shock proteins…….

The hypothesis is the following: If there is a way to mask (deleterious) changes in proteins under normal conditions, one may accumulate such changes without exposing them.

Hsp90 evolution
Illustration from Sangster TA et al. (more on Waddington will follow in the last post).

Thus, with Hsp90 acting as a buffer: one could potentially get a lot of hopeful monsters, under times of stress, as these traits were exposed. This would drastically increase the chances of a beneficial change to occur at the right time. And since the chance of mating with other monsters with similar traits (there are more than one monster, in fact very many), the chance of keeping the trait(s) in subsequent generations is also increased.

Now missing……..evidence, which will follow in the next post.

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Presenting….Hsp90 (chapter II, blogging in Just Science 08)

In Uncategorized on February 5, 2008 at 10:44 am

Chapter I gave an introduction into the role of Heat Shock Protein 90 (Hsp90) in evolution. The main point was its ability to mask/hide transforming mutations until needed in stressful times. This chapter describes its physical and biological properties.

Firstly, what the protein looks like. A really good introduction into Hsp90’s structure is found at the Sandwalk blog. Here’s a picture I stole (with permission) from his blog post on Hsp90:

Hsp90 structure
Picture from: Dollins, E.D., Warren, J.J., Immormino, R.M. and Gewirth, D.T. (2007) Structures of GRP94-Nucleotide Complexes Reveal Mechanistic Differences between the hsp90 Chaperones. Molec. Cell 28:41-56.

From Sandwalk: The complete protein is a dimer of two identical subunits. Each monomer has three distinct domains; an N-terminal domain (N); a middle domain (M); and a C-terminal domain (C). The ATP hydrolysis site sits at the interface between the N and M domains. The C domains interact to form the dimer. The presumed site of binding for misfolded proteins (“client” site”) is in the V-shaped pocket formed when the C domains come together. The mechanism of action of Hsp90 proteins is not known although it presumably involves a conformational change induced by ATP hydrolysis.

Secondly, it’s mode of action. Hsp90 works in concert with many other proteins to form protein-complexes ultimately activating the target protein. The molecular mechanism is as mentioned, still unresolved despite the presence of crystal structures. However, the following illustrations are taken from The Jackson Laboratory at University of Cambridge UK, to depict the overall events:
Model of the activation of a client protein by chaperone machines.
The chaperone Hsp70 system targets the client protein, in this case the steroid receptor, to the Hsp90 chaperone via the organising protein HOP which binds both chaperones. After transferring the steroid receptor to Hsp90, Hsp70 dissociates and is replaced by co-chaperones such as p23 and the high molecular weight immunophilin FKBP52. It is only in this complex that the steroid receptor is activated to bind ligand with high affinity.

 

And thirdly, which proteins does Hsp90 interact with. Again from The Jackson Laboratory:

 
Hsp 90 and Proteins it interacts with.
The abundant protein Hsp90 is thought to assist in the activation and assembly of specific proteins. Many of these proteins are critical for signal transduction and cell division. As a result, Hsp90 is a target for anti-tumour drugs.

Other examples are: PDK1, PKC-gamma, vSRC/cSRC, PPAR-alpha and p53. A comprehensive list of interacting proteins compiled by Picard Laboratory can be found here.

Thus, looking at all the proteins that can be affected by Hsp90, there is no surprise that the protein is highly conserved and that effects are profound when Hsp90 is fiddled with.

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Revolution Evolution (chapter I, blogging in Just Science 08)

In Uncategorized on February 4, 2008 at 10:01 am

My contribution to JustScience 2008 will be a review on a protein with the potential to transform evolution theory as we know it today. The review will be divided into 5 separate blog posts:

1. Introduction to Hsp90 and evolution (this post)

2. Presenting the Hsp90 protein

3. How can chaperones act in evolution

4. Evidence for Hsp90 involvement in rapid evolution of new traits

5. Summary

Here’s the teaser: In one generation you can go from this

Drosophila wild type

to this (a hopeful monster ?).

Hsp90 reduced expression in drosophila
(from Rutherford SL and Lindquist S, Nature 1998, to see more “monster”-pictures, do a google image search on Hsp90 and evolution)

The protein in focus, Heat Shock Protein 90, is otherwise as normal as a protein can get. It is ubiquitously expressed in all cells and across species, and its function is the same as other heat shock proteins, it’s a chaperone.

A chaperone is a protein that helps other proteins fold correctly (or prevents them to aggregate into non-functional protein junk). Without the chaperone the protein would not achieve an active conformation and end up being degraded. For an excellent video illustrating chaperone assisted protein folding go here.

Caperone illustration
Illustration of chaperones in action. Picture from Nurse Minerva

Now, it turns out that the Hsp90 chaperone function is important for development and evolution of new traits. To illuminate this, the first paper I will discuss is from 1998 by Susan Lindquist’s lab at the Whitehead Institute.

In this paper they take fruit flies and reduce the expression of Hsp90. As I shall come back to later, this is the experimental equivalent to a stressful condition (like for instance high temperature). They can’t knock out Hsp90, because a complete lack of it is not compatible with life (which demonstrates the importance of this protein). When Hsp90 levels are reduced, the fruit flies are born with a number of different defects ranging from defects in the legs and bristles to defects in the eye (see image above). Now, this could be expected when knocking out a protein, but such extensive morphologic changes are not expected from just reducing the levels of a protein (unless maybe if it’s a transcription factor, which it isn’t). The hypothesis explaining this goes as follows……Since Hsp90 is a heat shock protein it is helping other proteins fold. When stressful conditions occur, there will be more proteins that are in need of folding-help, thus some of the proteins that under normal conditions got their help from Hsp90 will now be left on their own. Since, under such conditions, you see these extensive morphological changes, these proteins must be doing something out of the ordinary when on their own, or if they aggregate, their absence causes abnormality. The theory states that these in-need-of-help-proteins must have accumulated mutations that potentially causes abnormality, but under normal conditions they are still able to perform normally due to the action of Hsp90. The implications for evolution are breathtaking since this means that an organism can accumulate a number of mutations and still function normally, but when exposed to stressful conditions, the changes at the protein level are suddenly exposed in their offspring and appear as physical abnormalities. This allows for extremely rapid evolution and could potentially enable a species to change in just a couple of generations.

Illustration of the process and further evidence for this theory will be presented in the next 4 posts the next 4 days, in SciPhu for just science 08.

Next week is Just Science 2008 – week

In Uncategorized on February 1, 2008 at 3:22 pm

Next week is devoted to science only, as I will be posting in Just Science 2008.

Hope many of you will still stop by. I am going to try to write so that the posts will make sense also to those outside of the field. I can’t guarantee though, that everything will be instantly comprehensible as this is dependent on my writing and presentation skills (where there’s definitely room for improvement). Please, do not hesitate to use the comments field for any questions.

The subject of the posts to come will be a mini-review on Hsp90 and evolution. I am grabbing this opportunity to present some scientific evidence for what I believe is the most underrated phenomenon in evolution biology.