NK Cells: Do these cells have Memory? (Part 1)

Introduction: A Primer


Memory

The ability of the immune system to recognize previously encountered pathogens and initiate a better immune response is the hallmark of memory. Responses to previously encountered organisms are characterized by, among other things, a large clonal population of cells that are uniquely specific for the offending pathogen. As an oversimplification, this large population then out-competes the rate of pathogen spread and brings the infection under control.

These memory populations have been thought to come from the adaptive immune system, consisting of both B and T cells. Upon primary infection, naive B and T cells specific for the pathogen exist at low precursor frequency. In order to be activated and clonally expand into a large effector population, these cells must be activated by the innate immune system, which provides the necessary co-stimulation in order to fully induce an adaptive response. Immune system responses can be thought of as a game of escalation: the innate arm recognizes pathogen-associated molecular patterns common to most pathogens and responds early in the infection process. If, however, the innate system is unable to control pathogen spread, it activates the adaptive immune system. Once activated, B and T cells expand into large number of cells directed against the specific pathogen, proceeding to contract into a small population of memory cells after clearance of antigen. These memory cells protect against future infection (or damage to the host) through a variety of ways

Inducing memory (and thus a B and T cell response) is the mechanism by which vaccines work.


NK cells

Natural killer cells are part of the innate immune system. Like all innate cells, NK cells are characterized by their ability to recognize patterns and respond early in infection. However, unlike macrophages and dendritic cells, NK cells do not interact directly with pathogens.* Instead, they recognize patterns on host cells associated with cellular abnormality, which can either be induced by viral infection or if the cell has become cancerous.

NK cells have both activating and inhibitory receptors on their surface which provide the ability to surveil the state of the host. These receptors interact with ligands on other cells and the combined signal from both types of receptors ultimately determine the response from the NK cell. If there are more inhibitory signals than activating ones, the NK cell does not respond. If, however, there are more activating stimuli, any inhibitory signal is overrode and the NK cell carriers out its effector function (death of the target cell, cytokine production). Examples of activating ligands include cellular stress molecules expressed during virus infection. In another example, the absence of an important inhibitory molecule called MHC is an indication that there is something wrong with the cell.

Because NK cells possess a limited set of receptors that respond to patterns of positive and negative signals instead of specific pathogens, it makes sense that these cells should not have memory. An expanded subset of relatively nonspecific NK cells (as compared to B and T cells) might actually be detrimental to the host in certain circumstances. When control can be handled by the innate arm of the immune system, inducing a large memory population of powerful NK cells might cause more damage to the host than the offending pathogen.

The Current Picture

However, within the past few years, the idea of what cell types constitute memory has been challenged. While this area is still largely the domain of B and T cells, recent evidence points to memory-like properties of NK cells such as: increased immune response to secondary encounter with antigen, adoptive transfer of NK cells providing pathogen-specific responses, and long- lived** subsets of NK cells induced by primary infection with pathogens.

In the following posts, my goal is to give a very brief introduction to this new finding.




*There are some virus-specific receptors, and we will talk about one of them in the near future.
**long-lived in this case meaning up to 90 days

Personal: Passed Quals!

So I passed my qualifying exam and am now an official Ph.D. candidate! That means I can get back to work on research. It also means that I can start posting more science-related articles. I've got a few that I think are really interesting and pave the way for breakthroughs in immunology.

The exam itself was a humbling experience. I mean, I knew I know 0.0001% of the total amount of immunology there is to know, but I found out I could work on some areas in addition to that.

Overall, I'll remember to work harder from the experience, but also to be confident in what I do know

Passing on a link: Michael Palm Basic Science, Vaccines, and Prevention Project Blog

No, I haven't gotten hit by the swine flu, I've just have been busy studying for my qualifying examine in immunology. Hopefully, I'll be a more regular poster around June.

In the meantime, I wanted to pass along this link to Michael Palm's blog for the Treatment Action Group (http://www.treatmentactiongroup.org/). It's a great resource for everything HIV related, and is current on HIV research findings.

If you really want to understand the immune system, just look at a virus which directly disrupts it.

His blog can be found at http://tagbasicscienceproject.typepad.com/

Passing on a link: Mystery Rays From Outer Space

Mystery Rays

If you like anything I put up on this blog, you'll definitely like Mystery Rays. Dr. Ian York does a great job of presenting new and interesting aspects of immunology/virology. Topics range from virus evolution to cancer and tend to center around immune escape. In addition to good reading, I have also used this site to find background information and paper suggestions. Mostly, this is because Dr. York covers some of the stuff they don't mention much in introductory immunology courses (for example, immunodominance). And, he does it in a clear and informative manner.

He has a new post up concerning the possibility of HIV actually increasing its virulence.

So, if that sparked your interest, go check it out!

Mystery Rays from Outer Space

http://www.iayork.com/MysteryRays/2009/04/16/is-hiv-becoming-more-virulent/

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The Dawning Age of Robot Scientists

Introduction

Friday has become my favorite day. Not because it's the weekend, oh no. No, Fridays are my favorite because I'm such a geek that I'm excited when The Journals come out. And even if you don't regularly scan their table of contents, you'd be hard pressed to miss the headlines in the science sections from the general news media: The first steps to a robot scientist are here people!

(All on the same story, but from different sources)

http://www.nytimes.com/2009/04/07/science/07robot.html
http://news.bbc.co.uk/2/hi/science/nature/7979113.stm
http://sciencenow.sciencemag.org/cgi/content/full/2009/402/1
http://blog.wired.com/wiredscience/2009/04/robotscientist.html

The actual scientific paper in Science is called The Automation of Science by King et al. at Aberystwyth University. There is also a comment on it and a related paper in the same issue (though the news outlets do a good job as well.)

Essentially, this robot, termed Adam, is able to complete the scientific process. Whereas computers and automation today are capable of collecting vast amounts of data, they have so far been unable to process it. Human scientists usually need to look at the data and determine what's useful. However, King et al. designed a system that can not only interpret the data and form a hypothesis about it, but can also perform followup experiments to test this hypothesis. To do this, they designed and implemented several different programs, ranging from hardware interaction to data collection and data analysis.

Just a note on the hardware: Adam is pretty beautiful, consising of multiple fridges, incubators, and robot arms. And, it never gets tired. The experiments involved growth curves of various yeast strains, which Adam can do hundreds of times a day. Just thinking about a human doing that amount of pipetting gives me thumb arthritis.

What Did Adam Actually Do?

Being a machine, Adam is great at hundreds of simple tasks and measurements. After taking specific yeast strains from the freezer, Adam can inoculate them in media and measure the ODs at various time points to establish a growth curve.

In order to test Adam, the authors decided to look at orphan enzymes, or enzymes for which the corresponding gene(s) haven't been identified. These enzymes have been well characterized in terms of their biochemical actions, so Adam is able to screen for enzyme activity by recording growth curves in specific media. Adam was also able to take advantage of yeast knockout strains for which the gene was known. Thus, Adam is able to take yeast that lack a gene and see if it is still able to grow when tested in a variety of limiting medias supplemented with certain metabolites. From this data, Adam is able hypothesize about the unknown gene encoding the orphan enzyme and is able to run further tests to confirm this hypothesis.

But Wait, There's More! (Bioinformatics)

One of the software packages I didn't talk about was their relational database of proteins and genes known to be present in yeast. Although the authors aren't descriptive, this is probably something similar to the DAVID Bioinformatics database (http://david.abcc.ncifcrf.gov/), Gene Ontology(http://www.geneontology.org/), or Pathways Analysis(http://www.ingenuity.com/products/pathways_analysis.html%20to)-- to name a few. I think this is the most important part of Adam, its "memory". Without such databases, it would be hard to understand gene and protein interaction and formulate an educated hypothesis.

Conclusion

Mass data collection, expanding bioinformatics tools, increasingly sophisticated programming, and efficient hardware are converging at the biological sciences, extending capability and knowledge at an exponential pace. Adam is only the beginning.

(If you don't believe me, read Ray Kurzweils The Singularity is Near, which, though on a slightly different topic, is on the same general trend. Or read about Eve at the group’s website, below.)

The paper is:

King et al. The Automation of Science. Science, vol. 324, April 3 2009.
http://www.ncbi.nlm.nih.gov/pubmed/19342587

The group's website can be found at:

http://www.aber.ac.uk/compsci/Research/bio/robotsci/

There are videos of Adam in action and other interesting things on the site

Passing on a link: The Virological Synapse

Virogical Synapses and HIV Transmission

A new article at MIT's Technology Review highlights a recent paper about the way HIV is transmitted between CD4 T cells. Using a GFP-expressing virus, the authors ( show direct cell-to-cell transmission of HIV via formation of what's known as the virological synapse. Check it out at http://www.technologyreview.com/blog/editors/23244/

Mini Introduction to the topic:

Viruses (as well as bacteria) can infect cells in a variety of ways. Normally, we think of the route of entry as being mediated by cell surface receptors. The virus attaches to the cell through this method and is taken into the cell, where it begins to replicate itself. Later, the cell bursts, spilling thousands of viral progeny into the host; alternatively, the virus can continually bud off from the plasma membrane. (The virus can either do this right away or form a latent infection, where viral DNA is present but little viral protein is made. In this method, the virus rides along with the cell until conditions are right to start producing viruses again.)

The key point here is that at some point in their lifetime, viruses are thought to go extracellular; viral particles have to spread out and infect cells. And when they're out in the open like that, the host immune system can see them better; more arms are available to fight the virus. For example, a virus can be neutralized by antibody or a lipid envelope can be degraded by complement (to name only a few things). If a virus wasn't exposed to this arm at all, the immune system is at a disadvantage.

Relatively recent work has proposed the existence of a virological synapse which effectively transmits virus between cells without progeny having to ever enter the extracellular space. Essentially, the virological synapse is an immune synapse (in this instance, without the priming stage). MHC II, as well as adhesion molecules, are upregulated upon T cell activation. The adhesion molecules attach to other T cells and MHC II interacts with TCR, collectively forming a tight seal between T cells. Usually, this is good for T cell priming, in addition to probably playing a pertinent role in cytotoxic T cell ability.

However, CD4 T cells are a main target for human immunodefiency virus (HIV). Instead of a protective immunological synapse forming, infected CD4 T cells interacting tightly with uninfected CD4 T cells can pass on the virus through the immunological/virological synapse. This might happen a variety of ways (i.e. trogocytosis, active infection, etc) but the thing to keep in mind is, again, that this exchange is thought to keep out antibody and other effector molecules. Incidentally, if the immunological/virological synapse is presenting cognate antigen, it will help to activate (or keep activated) the uninfected CD4 T cell, which provides an excellent environment for the virus to replicate in.

Not having read the paper (sorry to so uninformed, more pressing papers to read), I don't know whether the authors prove this beyond a doubt. (For example, notice how long it takes for the uninfected CD4 T cell to become infected. Usually an infection is quicker- maybe there was virus in the supernatant that was the cause of the late GFP explosion in that CD4 T cell.) But regardless, it certainly strengthens the argument for a virological synpase and is another avenue scientists have to explore if we are ever going to effectively combat this virus.

It’d also be interesting if they took a look at FAS/FASL on these cells. If they are going to interact that strongly, what happens when activation-induced cell death (AICD) starts to really get going? (…Or does HIV inhibit this, I just don’t know.)

The actual paper can be found at:

http://www.ncbi.nlm.nih.gov/pubmed/19325119

Prions are What?

Normally, I think of prions as a bad thing. Anyone hear of bovine spongiform encephalopathy(BSE)? BSE, or Mad Cow, was featured prominently in the news a few years ago and probably made a vegetarian out of a few people. That's because BSE is caused by prions, which are hard to detect, hard to prevent (prions are highly resistant to heat and other sterilizing devices), and ultimately result in death. So what are prions?

A prion consists of a single infective protein. Unlike other pathogens, nucleic acid isn't necessary to make future generations. Instead, a prion is capable of making more of itself by converting a particular host protein (called prion protein- PrP) into a prion. In essence, the prion protein functions to change the conformation of PrP such that it now takes on the function of a prion; these converted prions in turn recruit others. Most of the pathogenicity occurs because these prions also form aggregates in cells of the brain, ultimately resulting in death.

Prions, as we know them, are dangerous, but because PrP can be so easily converted into a prion, many groups have begun asking, "What function does normal PrP have inside a cell?” Or more generally, what are these potential prions doing there?

Eleven prions have been identified in yeast so far and are a good place to start for finding out what these things do. Interestingly, a new paper published in Nature Cell Biology (http://www.nature.com/ncb/) by Dr. Susan W. Liebman's group at the University of Illinois Chicago has discovered another method of non-Mendelian inheritance via prion function (note: this is my take, the paper is more scientifically conservative and accurate in that its title is The yeast global transcriptional co-repressor protein Cyc8 can propagate as a prion).

It's a good paper and there's a insightful comment on it in the issue, so I won't go into much detail here, but I did want to list a few key points.

Cyc8 (called OCT in its prion form) is a global transcriptional co-repressor protein that, among other things, regulates yeast growth on lactate. This protein suppresses other proteins important for growth on lactate plates. However, by transiently overexpressing the putative prion domain of Cyc8 (and thus increasing the chance of misfolding for the naturally occurring Cyc8) , the authors were able to show increased colonies growing on lactate plates versus controls. The authors were then able to do further experiments using this Lac+ phenotype

One of the most important things they did was an experiment to show that these Lac+ colonies were not spontaneous mutations resulting in revertants. (i.e. mutated nonfunctional Cyc8). Overexpression of functional Cyc8 did not restore the Lac+ phenotype.

However, as an aside, there was always the possibility of a gain-of-function mutation. Perhaps the gene required for lactate growth lost its ability to be repressed by Cyc8. I feel that simple sequencing of the most important genes involved in the Lac+ pathway could have been useful, but the authors’ main concern was elsewhere and they focused their remaining experiments in proving that OCT has all the characteristics of a prion.

This was the most fundamental, and convincing, aspect of the paper. The OCT+ mutation was dominant in an OCT- cross (not characteristic of Cyc8 loss mutations), cytoplasm of OCT+ cells could transfer the Lac+ growth phenotype, and fluorescence of OCT fused to YFP showed aggregates within the cells.

This paper continues to extend the knowledge of prion function. They ain't just fer disease anymore! Instead, proteins with prion-like characteristics might serve to regulate cell function in a rapid and inherited manner, without the need to continuously monitor transcriptional and translational levels of mRNA. (In fact, this paper cites other work with the prion Swi, a protein involved in chromatin remodeling.)

While this paper involves propagation of OCT and didn't look at any possible regulation of Cyc8 to OCT conversion, the fact that there are 11 putative prions in yeast is suggestive. Instead of relying on mutation and unchangeable consequences, perhaps some prion-like proteins can be converted between the two states. (The paper might have a citation on this; I'll need to check it out further)

Again, if you're really interested, check out the paper:


Patel B.K., Gavin-Smyth J., and S.W. Liebman. The yeast global transcriptional co-repressor protein Cyc8 can propagate as a prion. Nature Cell Biology. March 11, 2009. DOI: 10.1038/ncb1843

http://www.ncbi.nlm.nih.gov/pubmed/19219034