“In my day” i.e. when I started my PhD back in 197<cough>, the first few weeks were spent joining the Enzyme Club. This encompassed all the biomedical researchers at the University of Leicester. Each new student would prepare a batch enzyme for recombinant DNA work. In my case, I made Hsu I (an isoschizomer of Hae III but allegedly easier to prepare). Since it was years ago, I can’t remember how many litres of the organism I grew up, but I remember very clearly doing the first assay on two litres of crude extract, and figuring out I was holding £40 million pounds worth of enzyme at the then current market prices. The first affinity column cut it down to £15 million, and a quick gel filtration to couple of millions pounds worth – still pretty good for two weeks work, especially when you remember that two million pounds was enough to buy you a house back in the 1970s!

Why did the Enzyme Club exist? Because these reagents were scarce in the 1970s, and rationed both by price and availability. Only a few years before, the only way to get hold of any of these enzymes was to make your own. This type of open science made sense. Why did the Enzyme Club cease to exist? Gradually, it became clear that the batch of enzyme I made wasn’t very good. It had a persistent exonuclease activity which meant it was fine for restriction analysis but rubbish for cloning, and it went off very quickly in storage, so that after three months there wasn’t much activity left. And although I’ve always been a rubbish protein chemist, that was a pretty common experience. Gradually, the companies dropped their prices and improved both the quality and availability of commercial enzymes. The day came when the Enzyme Club didn’t make sense any more, and it quietly died. It’s probably still moldering in the back of a coldroom over in the MSB.

So boys and girls, this is a story of the economics of open science, which made sense in response to scarce resources. When the availability of enzymes was limiting, this open approach made sense. When time became limiting, we all retreated back into our laboratories and got on with whatever we needed to do to get a PhD. The moral of this story is that open science pops up it’s head when times are hard and resources are scarce, but retreats quickly as the balance changes.

Viruses provide attractive models to study simple developmental decisions. In prokaryotes, much has been learned using bacteriophage λ to investigate how the lysis-lysogeny decision is made. In eukaryotes, latent herpesviruses exist in one of two developmental states within their hosts and must resolve an analogous question of whether to remain latent or initiate productive viral growth. Different herpesviruses permanently colonize distinct specialized host cell-types. Those that establish residency in dividing cell populations, exemplified by members of the γ-herpesvirus subfamily that includes Kaposi’s sarcoma associated herpesvirus (KSHV/HHV8), express a limited subset of viral genes that stimulate cell proliferation, allow for viral minichromosome replication and segregation, and evade antiviral defenses. In response to poorly understood environmental cues, these viruses can switch to a developmental program that results in productive replication. This alternate pathway involves activating a temporally coordinated cascade of viral lytic gene expression, which in turn results in massive viral DNA amplification, progeny virus production, and ultimately host cell destruction. To effectively switch its gene expression program, all herpesviruses produce a new population of viral mRNAs transcribed by the cellular RNA polymerase II, which are mostly capped and polyadenylated like their host counterparts, and these must successfully engage and reprogram the host cell translational apparatus. This is a critical component of the developmental switch because viruses are absolutely dependent upon the translational machinery resident in their hosts. Manipulating host translation initiation factors to ensure that nascent viral mRNAs successfully recruit ribosomes will therefore determine the overall level and efficacy with which the newly transcribed developmental instructions are executed. While we have a general understanding of how the viral transcriptome is altered for many viruses, a role for translational control in the developmental switch from a latent to a productively replicating state has not been described.

Kaposi’s sarcoma-associated herpesvirus (KSHV) is an important human pathogen and, like all herpesviruses, establishes a state of permanent residency in the infected host called latency. Major sites of KSHV latency are cells of the immune system and cells lining blood vessels. In individuals with weakened immunity, inappropriate growth of these cells driven by the resident virus can give rise to primary effusion lymphoma and Kaposi’s sarcoma, respectively. These life-threatening cancers are most common in patients with HIV/AIDS and have become a major source of mortality in parts of sub-Saharan Africa. Under appropriate stimuli, herpesviruses change their relationship with the host cell and begin to manufacture proteins required to assemble new infectious virus particles that can be released and spread. To achieve this, the virus hijacks key processes within the cell and conscripts them into producing viral proteins. A recent study describes for the first time how KSHV carefully manipulates the host protein synthesis machinery during the switch from latency to this specialized infectious virus production mode. The results show that although overall protein synthesis is diminished, key components of the host’s protein manufacturing machinery are actually stimulated, presumably to accelerate virus protein production.

Activation of Host Translational Control Pathways by a Viral Developmental Switch. PLoS Pathog 5(3): e1000334. doi:10.1371/journal.ppat.1000334
In response to numerous signals, latent herpesvirus genomes abruptly switch their developmental program, aborting stable host–cell colonization in favor of productive viral replication that ultimately destroys the cell. To achieve a rapid gene expression transition, newly minted capped, polyadenylated viral mRNAs must engage and reprogram the cellular translational apparatus. While transcriptional responses of viral genomes undergoing lytic reactivation have been amply documented, roles for cellular translational control pathways in enabling the latent-lytic switch have not been described. Using PEL-derived B-cells naturally infected with KSHV as a model, we define efficient reactivation conditions and demonstrate that reactivation substantially changes the protein synthesis profile. New polypeptide synthesis correlates with 4E-BP1 translational repressor inactivation, nuclear PABP accumulation, eIF4F assembly, and phosphorylation of the cap-binding protein eIF4E by Mnk1. Significantly, inhibiting Mnk1 reduces accumulation of the critical viral transactivator RTA through a post-transcriptional mechanism, limiting downstream lytic protein production, and impairs reactivation efficiency. Thus, herpesvirus reactivation from latency activates the host cap-dependent translation machinery, illustrating the importance of translational regulation in implementing new developmental instructions that drastically alter cell fate.

Related:

• Cellular Genes Targeted by KSHV MicroRNAs
• Modulation of the immune system by Kaposi’s sarcoma-associated herpesvirus

In recent years, the Internet and its users have undergone a fundamental transformation. Originally, the Internet was designed to allow access of information provided by the publisher of a given website. Although websites have become very sophisticated in how this information is presented and while the information given can be tailored to the individual user, the information stream is predominantly unidirectional from the website to the user, and the content of the site is determined by its owner. Examples include sites that aim to convey general information about businesses or organizations, for example, the website of the Centers for Disease Control and Prevention, or allow for minimal manipulation of personalized information, such as online banking or bill payment. In Internet terms, this use of the Internet is referred to as Web 1.0. By contrast, Web 2.0 comprises Internet applications in which the information stream is more or less reversed. Here, the content of a website is mostly driven by the users of the site. This information can take multiple forms, including a variety of uploaded file formats (text, graphics, audio, video), blogs (web-logs), vlogs (video logs), chats, etc. Social networking sites, including YouTube, MySpace, and FaceBook are among the prime examples of the Web 2.0 applications that have revolutionized Internet use in the last decade.

In this article, the authors discuss the implications of the shift from Web 1.0 to Web 2.0 technology on sexual health from three perspectives: the Internet as an STI risk environment, the Internet as a venue for STI prevention, and, finally, the Internet as a tool for STI service and prevention providers. The growth of the Internet as a communication medium has had far-reaching consequences for STI/HIV prevention ranging from a venue for partner recruitment with potential risk as well as prevention benefits, to the use of the Internet as a place to deliver STI/HIV prevention services in a variety of more or less interactive formats, and finally as a tool for the development of a prevention work force. However, while the Internet has great potential as an important STI/HIV prevention medium, it appears that the greatest potential is yet untapped and that the providers of these services are considerably lagging behind their target audience in the creative and innovative uses of the new medium.

Web 2.0 and beyond: risks for sexually transmitted infections and opportunities for prevention. Curr Opin Infect Dis. 2009 22(1): 67-71
The continued growth of the Internet as a communication medium has had major implications for the transmission and prevention of sexually transmitted infections (STIs). The purpose of this review is to describe recent developments in this rapidly changing environment. The interface between the Internet and STIs is described from three perspectives: the Internet as a risk environment, that is, a place where prospective, potentially STI-infected, sex partners can be recruited; the Internet as a venue where public health prevention interventions aimed at STIs and HIV prevention can be placed; and the Internet as an increasingly important work environment for all STI prevention disciplines. The review highlights recent developments and identifies potential avenues for future research and program development. The increasing interactivity of the Internet, known as ‘Web 2.0′, especially the user-driven social networking sites that allow users to share near limitless amounts of personal information with their peers in the network, is compounding the potential of the Internet as an environment for both STI risk and prevention.

Related:

• Health Risk Behaviors on MySpace
• You’ve Got Mail

Unfortunately, by this time, it is too late for the antibody (‘‘humoral’’) immune response to clear HIV from the body. Indeed, the humoral immune response to HIV is very slow; for most viruses, neutralizing antibodies appear within days of infection. To help them design an effective HIV vaccine, scientists need to understand how the virus delays humoral responses to HIV infection (and how it later causes the production of HIVspecific antibodies to decline). Little is known, however, about the early effects of HIV infection on B lymphocytes. These cells are born and mature in the bone marrow. ‘‘Naive’’ B lymphocytes, each of which carries an antigen-specific receptor (a protein that binds to a specific antigen), then enter the blood and circulate around the body, passing through the ‘‘peripheral lymphoid organs’’. Exposure to antigens in these organs, which include lymph nodes and gut-associated lymphoid tissues, activates the subset of B lymphocytes that recognize the specific antigens that are present. Finally, with the help of activated T lymphocytes, the activated B lymphocytes proliferate and change (differentiate) into antibody-secreting cells and memory B lymphocytes (which respond more quickly to antigen than naive B lymphocytes). In this study, the researchers investigate the effects of early HIV-1 infection on B lymphocytes in blood and in gut-associated lymphoid tissues.

Although the depletion of gut-associated CD4+ T lymphocytes in early HIV-1 infection is well known, these new results demonstrate the effects of early HIV-1 infection on gut-associated and circulating B lymphocytes. The results of this study are limited by the methods used to analyze the antibodies induced by HIV infection and by only taking tissue samples from one region of the gut. Nevertheless, the findings of polyclonal B-cell activation and damage to gut-associated lymphoid follicles soon after HIV-1 infection may have implications for HIV-1 vaccine design. Specifically, these findings suggest that an effective HIV-1 vaccine will need to ensure that significant levels of neutralizing antibodies are present in people before HIV-1 infection and that other protective immune defenses are fully primed so that, in the event of HIV-1 infection, the virus can be dealt with effectively before it disables any part of the immune system.

Polyclonal B Cell Differentiation and Loss of Gastrointestinal Tract Germinal Centers in the Earliest Stages of HIV-1 Infection. PLoS Med 6(7): e1000107. doi:10.1371/journal.pmed.1000107
The antibody response to HIV-1 does not appear in the plasma until approximately 2–5 weeks after transmission, and neutralizing antibodies to autologous HIV-1 generally do not become detectable until 12 weeks or more after transmission. Moreover, levels of HIV-1–specific antibodies decline on antiretroviral treatment. The mechanisms of this delay in the appearance of anti-HIV-1 antibodies and of their subsequent rapid decline are not known. While the effect of HIV-1 on depletion of gut CD4+ T cells in acute HIV-1 infection is well described, we studied blood and tissue B cells soon after infection to determine the effect of early HIV-1 on these cells. In human participants, we analyzed B cells in blood as early as 17 days after HIV-1 infection, and in terminal ileum inductive and effector microenvironments beginning at 47 days after infection. We found that HIV-1 infection rapidly induced polyclonal activation and terminal differentiation of B cells in blood and in gut-associated lymphoid tissue (GALT) B cells. The specificities of antibodies produced by GALT memory B cells in acute HIV-1 infection (AHI) included not only HIV-1–specific antibodies, but also influenza-specific and autoreactive antibodies, indicating very early onset of HIV-1–induced polyclonal B cell activation. Follicular damage or germinal center loss in terminal ileum Peyer’s patches was seen with 88% of follicles exhibiting B or T cell apoptosis and follicular lysis. Early induction of polyclonal B cell differentiation, coupled with follicular damage and germinal center loss soon after HIV-1 infection, may explain both the high rate of decline in HIV-1–induced antibody responses and the delay in plasma antibody responses to HIV-1.

Related:

• Antibodies against persistent viruses
• Re-awakening old genes to help in the fight against viruses
• HIV is evolving rapidly to escape the immune system

On MicrobiologyBytes I’ve often discussed bacterial communication – the ways in which bacteria talk to each other using chemical signals. If Bacillus subtilis cells are growing in a nutrient-poor area, they release chemicals into their surroundings which tell their neighbours “There’s not much food here, so clear off or we’ll both starve.” In response to these chemical messages, the other bacteria move away, changing the shape of the colony.

Many single-celled organisms can work out how many other bacteria of their own species are in their vicinity – something known as “quorum sensing“. Each individual bacterium releases a small amount of a chemical into the surrounding medium. If there are lots of other bacteria around, all releasing the same chemical, levels can reach a critical point and trigger a change in behaviour of the whole population. This “voting system” can be used to decide when to launch an attack on a host. Once they have grown to sufficient numbers to overwhelm the immune system, they collectively launch an assault on the body. Jamming these signals might provide us with a way to fight back.

Bacteria form communities known as biofilms, familiar as the thin layer of slime that coats the insides of water pipes, or surgical implants. Many different species live side by side in these “bacterial cities”, consuming each other’s wastes, cooperating to exploit food sources, and safeguarding one another from external threats such as antibiotics.

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Many microbes can accelerate the rate at which their genes mutate. This allows them to obtain new abilities that may be helpful when conditions get tough. Escherichia coli mutates more rapidly when under stress (Stress-induced mutagenesis in bacteria. Science. 2003 300(5624): 1404-9), and yeast can perform the same trick (Adaptive mutation in Saccharomyces cerevisiae. Crit Rev Biochem Mol Biol. 2007 42(4): 285-31).

Microbes are also pretty good at navigation. The single-celled algae Chlamydomonas swim towards light, but only if it is of a wavelength that they can use for photosynthesis. Some bacteria move according to the presence of chemicals in their environment – a behaviour called chemotaxis. Another group of bacteria align themselves to the Earth’s magnetic field, allowing them to head directly north or south, and more importantly, up or down for optimum photosynthesis.

When the amoeba Dictyostelium searches the surface of a Petri dish for food, it makes frequent turns. But it does not do so randomly. If it has just turned right, it is twice as likely to turn left as right on its next turn, and vice versa. It remembers which direction it last turned.

Remarkable though these behaviours are, we have probably only scratched the surface of what single-celled organisms can do. With so many still entirely unknown to science, there must be plenty more surprises in store.

Related:

• Quorum Sensing
• Biofilms
• It’s good to talk
• Communication, cooperation, competition and cheating – bacteria are just like us

Bacteria capture iron from heme by keeping tetrapyrrol skeleton intact. PNAS USA June 29, 2009, doi: 10.1073/pnas.0903842106
Because heme is a major iron-containing molecule in vertebrates, the ability to use heme-bound iron is a determining factor in successful infection by bacterial pathogens. Until today, all known enzymes performing iron extraction from heme did so through the rupture of the tetrapyrrol skeleton. Here, we identified 2 Escherichia coli paralogs, YfeX and EfeB, without any previously known physiological functions. YfeX and EfeB promote iron extraction from heme preserving the tetrapyrrol ring intact. This novel enzymatic reaction corresponds to the deferrochelation of the heme. YfeX and EfeB are the sole proteins able to provide iron from exogenous heme sources to E. coli. YfeX is located in the cytoplasm. EfeB is periplasmic and enables iron extraction from heme in the periplasm and iron uptake in the absence of any heme permease. YfeX and EfeB are widespread and highly conserved in bacteria. We propose that their physiological function is to retrieve iron from heme.

Related:

• Iron Uptake and Virulence
• New drug targets for urinary tract bacterial infections
• Bacillus anthracis gets iron from hemoglobin
• Utilization of iron by Campylobacter jejuni
• Bacterial sensors of oxygen

One of the immediate problems facing evolutionary studies of viruses is the evident fact that viruses are hugely diverse in size, appearance, even the nature of their genetic material (DNA or RNA). From this, it is reasonably clear that they are a not a single evolutionary group, and cannot be easily added as a single unit to the tree of life with its three main divisions (Bacteria, Archaea and Eukarya). By the same token, it seems likely that different virus groups (e.g. animal RNA viruses, retroviruses, large DNA viruses, bacteriophages) may indeed have entirely separate evolutionary origins. In this article I will describe two areas where recent discoveries have produced tantalizing new insights into the origin and ubiquity of some of these groups. Through the application of new, mass-sequencing techniques and scope for large-scale environmental sampling for virus genomic sequences, we may finally be able to understand the extent and complexity of the “virosphere” in which we live, and the extraordinary diversity of viruses that infect us.

Read more

Related:

• Evolutionary history and phylogeography of human viruses
• One virus particle can be enough to cause infection
• Coevolution with viruses drives bacterial mutations