The recipient of the 2018 Crow Award reveals details of flu evolution at the smallest —and largest—scales.

For many viral diseases, a vaccine can provide lifelong protection. But for flu, you need a new shot every year. The influenza virus evolves so fast it presents a constantly moving target for both our immune systems and public health authorities, fueling epidemics like the particularly bad season we just endured. With over 30,000 people hospitalized in the United States alone this season, the flu provides a dramatic reminder of the importance of understanding evolutionary dynamics.

Katherine Xue, a graduate student at the University of Washington, is revealing the mechanics of influenza evolution on scales ranging from an individual person up to the entire planet. Xue was awarded the 2018 James F. Crow Award for Early Career Researchers for her doctoral work on the subject after a presentation at the Population, Evolutionary, and Quantitative Genetics Conference in May.

In a special session of talks by Crow Award finalists, Xue spoke about using deep sequencing to examine diversity in flu virus populations.

“Up until recently, we were only able to look at the average genetic identity across the millions or billions of flu viruses in a single infection,” says Xue. “We’ve used deep sequencing to show that within a single infection there are fast evolutionary dynamics that have been invisible to previous technologies.”

Xue approaches clinical topics with the conceptual tools developed by evolutionary and population geneticists. Linking ideas across fields is characteristic of Xue, says her mentor Jesse Bloom (Fred Hutchinson Cancer Research Center / University of Washington), whether it’s between medicine and evolutionary theory or between science and the humanities.

“What makes Katherine stand out is her ability to think about big scientific concepts and connect ideas,” says Bloom. “She’ll see and connect ideas in ways that I can’t.”

Viral cooperation

When Xue first rotated in Bloom’s lab, she was working on a molecular virology project about how a particular viral protein binds to a cell. In the course of examining sequence databases, she noticed the mutation she was studying was often ambiguously annotated.

Inspired by this hint of population diversity, she wondered whether the two viral variants might interact with each other. She was able to establish that the mutation tended to occur alongside the wild type version within a population; the mutation was deleterious to virus reproduction on its own but beneficial when mixed with wild-type. This example of cooperation suggests that interactions between different variants within flu populations can be important factors in virus evolution.

A glimpse of evolution in action

But can evolution be detected within the virus population of a single individual? Xue was drawn to the question of how global flu evolution traces back to the founding infections in which each mutation must first arise.

“I was intrigued because it was hard to imagine how this works,” says Xue. “Flu infections are very short; there’s not a lot of time for a new mutation to reach frequencies large enough to ensure it makes it over to the next infected person.” In the language of population geneticists, flu populations are repeatedly subjected to extreme bottlenecks. But observing such rapid evolution in action is extremely challenging.

Xue and her colleagues in the Bloom lab used a unique approach to get around this problem. They partnered with clinicians Michael Boeckh and Steve Pergam at the Fred Hutchinson Cancer Research Center, who had collected samples from four immunocompromised patients over the course of their months-long flu infections. Deep sequencing these samples gave them an in-depth view of a process that would normally be finished within days in a person with healthy immune defenses.

“I initially had doubts that this project would show us anything interesting or be worth doing,” says Bloom. “But Katherine is very independent and persistent, and she kept going despite my occasional words of discouragement.”

The results were dramatic. Over the span of about two months, there was a substantial amount of flu evolution within each patient. Mutations arose regularly, fluctuated in frequency, and even became fixed in the population in a few cases. They also saw evidence that some of these changes are due to selection. The same mutations would often arise independently and then rise to substantial frequencies in multiple patients, suggesting these particular changes were adaptive.

Remarkably, the mutations that arose repeatedly in different patients were sometimes the same mutations that spread through the global flu population within the next decade. The immunocompromised patients seemed to be microcosms of global evolutionary patterns. “We were astonished,” said Xue.

Most of these recurring mutations affect the part of the flu haemagglutinin protein that is most recognized by the host immune system, so the team hypothesizes that the changes help the flu escape host defenses.

These results raise many questions about how evolutionary dynamics interact across scales. How far do the conclusions generalize? Where and when do natural selection and genetic drift act? How do normal week-long infections generate enough diversity to fuel rapid global evolution? Could understanding these processes translate to better flu season predictions? Xue’s graduate research continues to explore flu evolution with these questions in mind.

Connecting ideas

The scientific big picture is never far from Xue’s mind, it seems. Alongside her thesis research, Xue is pursuing a certificate in science and technology studies, with a capstone project on the history of flu research. “I have really loved being part of the history, philosophy, and sociology of science community here,” she says. “It’s given me a lot of perspective that has been really enriching.”

A few years ago Xue helped start the UW Genomics Salon, which is a group of students and postdocs who take part in freeform discussions about the intersections of science and society. These discussions touch on policy, advocacy, communication, education, representation, law, art, and a host of other topics.

Bloom thinks Xue’s broad interests are yet another reflection of her creativity and ability to link ideas across fields. Before graduate school, she spent a few years working as a science writer for the Harvard Magazine. “She’s an exceptionally good science communicator and very dedicated to creating connections between science and other fields. It’s pretty awesome to have someone like that around!”

[youtube https://youtu.be/fTdaAwqdt0k&w=500&rel=0]

Watch #PEQG18 presentations from all the other  outstanding finalists for the Crow Award here.

GC content alone is associated with distinct functional classes of human enhancers.

Because enhancers can be located hundreds of kilobases away from their target genes, it can be challenging to accurately predict their functions. A new report in GENETICS uses sequence composition to distinguish two enhancer classes that have distinct functions and spatial organization in humans.

Enhancers are regulatory DNA sequences that aid in transcription initiation. In some ways, enhancers are like promoters, since both are bound by transcription factors as part of transcription initiation. Unlike promoters, which are located near the transcriptional start site of the genes they regulate, enhancers are sequentially far away from their targets, typically coming into long-distance contact with gene promoters via 3D DNA looping. Since it is difficult to identify enhancers through sequence information alone, our understanding of them is somewhat primitive compared with other DNA regulatory elements.

Lecellier, Wasserman, and Mathelier were interested in classifying enhancers based on their sequences. The percentage of a given sequence that is guanine and cytosine (the GC content or %GC) can be used to classify promoters, so they investigated whether a similar approach could be useful for enhancer classification. To perform this analysis, they took advantage of the FANTOM5 project, which recently cataloged tens of thousands of enhancers across the human genome.

The enhancers were divided into two simple groups: those with higher %GC and those with lower %GC than the median overall. The authors compared the properties of the two groups, finding that different transcription factors were predicted to be associated with each group. Each group was also associated with different DNA shapes (e.g. bending) and distinct localization in chromatin loops, suggesting that the enhancer sequence composition is linked to the 3D architecture of the chromatin.

The authors then examined whether the two groups of enhancers had distinct biological functions. By consolidating previous reports, they compiled lists of thousands of genes predicted to be targets of each class of enhancer, and they analyzed these genes as proxies for the biological functions of the enhancers across different cell and tissue types. They found that enhancers with a higher %GC were associated with ubiquitous gene expression, whereas enhancers with a lower %GC were associated with specific patterns of expression in particular subsets of cells.

In particular, lower %GC enhancers were linked to immune response genes. To test this association against experimental data, the authors used data obtained from dendritic cells infected with Mycobacterium tuberculosis. This data tracked changes in chromatin accessibility, which can be mediated by enhancer activity. They found that lower %GC enhancers were significantly more activated in infected cells, providing experimental support for their observations.

CITATION:

Human enhancers harboring specific sequence composition, activity, and genome organization are linked to the immune response

Charles-Henri Lecellier, Wyeth W. Wasserman, Anthony Mathelier

Genetics August 2018, 209: 1055-1071; https://doi.org/10.1534/genetics.118.301116

http://www.genetics.org/content/209/4/1055

Beer, doughnuts, and genetics textbooks have one thing in common: they were all made possible by collaborations between humans and yeast. Our fungal ally Saccharomyces cerevisiae resides not only in breweries, bakeries, and laboratories, but also sometimes in our own bodies—where, on rare occasions, it betrays us.

S. cerevisiae is increasingly being reported as an opportunistic human pathogen, sometimes even in non-immunocompromised people. Infections remain rare overall, but studying them presents an exceptional opportunity to understand how benign fungi can turn destructive. Such lethal transformations are more common in some other fungi, but those species are often difficult to grow and manipulate in the lab. In contrast, the long history of S. cerevisiae use in laboratory research means there are many powerful tools for working with the species. If baker’s yeast and other fungi become pathogenic through similar mechanisms, understanding S. cerevisiae infections could have broad implications.

With this in mind, Phadke et al. searched for traits that allow S. cerevisiae to invade a model host: the larval form of the greater wax moth Galleria mellonella. This moth is often used in studies of innate immunity, and because it doesn’t naturally cohabitate with S. cerevisiae, the yeast shouldn’t have any preexisting adaptations that would allow it to infect the moth. This means investigating how yeast infect it can inform researchers about fungal pathogenesis in general.

Comparing yeast strains derived from clinical samples to ones from environmental samples revealed that strains from both groups were equally likely to be pathogenic in the moths. Although this conflicts with findings in mice that clinically isolated strains are more likely to be pathogenic, the studies do concur that pathogenic strains are more likely to be able to form pseudohyphae—elongated strands of cells that remain joined after cell division.

The researchers used a library of yeast deletion mutants to screen for strains that grow better or worse in the moth larvae than in standard conditions. They noticed that one strain that struggled to infect the moths lacked the gene slt2, which encodes a protein that helps maintain part of the yeast’s cell wall. This part of the cell wall normally shields fungal molecules that the host could recognize. If these molecules were noticed by the moth immune system, it could cause a defensive response against the yeast, providing a possible explanation for the strain’s inability to infect the moths. This finding aligns with previous observations that cell wall integrity genes are important for other opportunistic fungal pathogens to infect hosts.

Several genes encoding mitochondrial proteins were also more important for yeast growing in the moth larvae than they were for cells growing in vitro, but since mitochondria have so many roles in the cell, it’s difficult to say how exactly these genes affect pathogenesis. The screen also turned up genes involved in metabolism of some aromatic compounds. In most cases, strains with one of these genes deleted grew better in the moths than they did in vitro, indicating that the genes aren’t important for pathogenesis—but two strains suffered dramatic fitness hits. Both strains were mutant for genes required for the synthesis of two aromatic amino acids, tyrosine and phenylalanine. In mice, S. cerevisiae infection is greatly attenuated if the yeast cannot synthesize aromatic amino acids, and infection in the moths may be similar in this regard.

This work asserts that some of the genetic underpinnings of pathogenesis are shared among S. cerevisiae and other fungal opportunistic pathogens and fuels hope for development of a broad-spectrum treatment targeting numerous fungal infections. It also demonstrates the power of our alliances with laboratory models—even when they go on the attack.

CITATION:

Phadke, S.; Maclean, C.; Zhao, S.; Mueller, E.; Michelotti, L.; Norman, K.; Kumar, A.; James, T. Genome-Wide Screen for Saccharomyces cerevisiae Genes Contributing to Opportunistic Pathogenicity in an Invertebrate Model Host.
G3, 8(1), 63-78.
DOI: 10.1534/g3.117.300245
http://www.g3journal.org/content/8/1/63

Taking probiotics might be the latest health fad, but for people with inflammatory bowel diseases, the microbiome is a more serious matter. With these autoimmune diseases, the composition of the gut microbiome can have critical health consequences. In the June issue of GENETICS, Mistry et al. use fruit flies to determine whether immune system activity affects the composition of the gut microbiome. They found that flies with compromised immune systems had a similar microbiome to their healthy relatives, but flies with overactive immune systems had distinctly different microbiome compositions and more gut microbes overall. This work suggests that the interaction between the immune system and the gut microbiome is a two-way street.

As a fruit fly ages, the species composition of its gut microbiome changes Young flies have diverse microbiomes that vary with their genotype and environment, but as they mature into adults, the composition converges on a typical makeup. In this study, Mistry et al. surveyed microbiome composition across the lifespan of flies with healthy, compromised, or constitutively active immune systems while controlling for bacterial transmission from the mother—the most important initial source of microbiome bacteria.

Overall, their results revealed that a constitutively active immune system has a much larger impact on gut microbiome composition than an underactive immune system. In normal conditions, flies with an immune deficit develop the typical adult microbiome more rapidly and end up with a higher density of microbes but are otherwise similar to healthy flies. Only flies whose maternally introduced microbes were eliminated showed a difference in microbiome composition.

In contrast, flies with an overactive immune system maintained very different microbiome diversity to normal, even for those exposed to their mother’s microbes, indicating that constitutively active immunity has a powerful impact on the microbiome. The authors also found that when these flies were co-housed with healthy flies, the microbiomes of the healthy flies become more similar to those with overactive immune systems.

One possible explanation is that constant immune activity triggers inflammation in the gut, creating an aerobic environment that cultivates a different variety of bacteria than a healthy intestinal tract. Human conditions like Crohn’s disease and ulcerative colitis are characterized by constant inflammation and have been linked to changes in the gut microbiome. In fact, some types of bacteria commonly found in these patients were the same as those identified in the flies with overactive immune systems, suggesting Drosophila may prove a useful model for these debilitating diseases.

Interaction Between Familial Transmission and a Constitutively Active Immune System Shapes Gut Microbiota in Drosophila melanogaster.

Rupal Mistry, Ilias Kounatidis and Petros Ligoxygakis

GENETICS. June 1, 2017 vol. 206 no. 2 889-904; https://doi.org/10.1534/genetics.116.190215

http://www.genetics.org/content/206/2/889

The arms race between pathogens and their hosts leaves clear genetic marks: the most quickly evolving parts of host genomes often include immune genes. But are these fast-movers the exception among immune genes, or do most genes in this class bear the genetic signature of strong selection? In the January issue of GENETICS, Early et al. address this question.

In their comparison of five Drosophila melanogaster populations using information from FlyBase (a database of information on fly genes and genomes) and genetic data from a recently sequenced set of Drosophila lines called the Global Diversity Lines, the researchers found that viral defense genes are evolving at a remarkably fast pace. Genes that protect against viruses through the RNA interference pathway, which involves using RNA to shut down pathogens’ gene expression or translation, showed particularly elevated rates of change.

The evolution of genes responsible for resisting bacterial and fungal invaders, in contrast, is much more sluggish. And the changes are not as far-reaching—rather than broadly altering the regulation of the immune response, the adaptations to bacterial and fungal threats tend to involve making minor tweaks to recognition and effector genes.

Early et al. also found that some types of immune genes have changed more rapidly at the population level than at the species level and vice versa. For instance, phagocytosis and recognition receptor genes showed signs of rapid adaptation at the population level but not the species level, while genes in the Toll and immune deficiency (IMD) pathways had the opposite pattern.

This discrepancy may have been found because the population data came from samples collected at one specific time and thus can’t capture the dynamic fluctuations in selection and adaptation over a longer period. The fact that the researchers observed significant alterations to viral defense genes at both the population and species level, then, is a testament to the powerful directional selection driving those changes.

These extreme cases may lead to the impression that immune system genes in general are evolving more rapidly than the rest—a view that, the researchers conclude, doesn’t capture the whole story, at least in the fruit fly.

CITATION:

Early, A.; Arguello, R.; Cardoso-Moreira, M.; Gottipati, S.; Grenier, J.; Clark, A. Survey of Global Genetic Diversity Within the Drosophila Immune System.
GENETICS, 205(1), 353-366.
DOI: 10.1534/genetics.116.195016
http://www.genetics.org/content/205/1/353

In 2014, the GSA journals launched a contest inviting image submissions related to genetics and genomics. The winning entry was created by Jian Han, of the HudsonAlpha Institute for Biotechnology, and appears on the cover of the May 2015 issue of G3. We talked with Dr. Han about the striking image:

What does the image represent?

This image, called “The Picture of Health,” is an Im²print of a healthy immune system. An Im²print is one of many ways to decipher the large amount of data that comes from studying an individual’s immune repertoire. It is a tree map, with each rectangle representing a unique gene combination coding for B or T cell receptors, and the ability to defend against a particular antigen. The larger the rectangle, the more expressed the gene combination. In theory, a healthy individual would have a diverse immune repertoire; thus, the tree map would show many differently colored rectangles all relatively equal in size. In unhealthy patients, however, the immune system is compromised and can be less diverse, so the individual is less prepared for defense  against disease. The tree map of an immunocompromised individual would most likely be dominated by a single rectangle or small group. Im²prints are not only a quick graphical representation of the overall diversity of an individual’s immune repertoire, but are also personalized artwork.

Why are imp²print tree maps useful?

Im²prints are visually stunning, especially when people realize that the image was generated from data collected from an individual’s immune system.  By comparing side-by-side images of immune systems of different people, the health of each person’s immune system is vividly visualized: The larger the rectangles, the less healthy the immune system, while greater diversity indicates a healthier system.  This idea is not only useful in research and the clinic, it is also pleasing to the eye of people interested in science and health and the use of science as art.

What is your current research focus?

My laboratory focuses on developing integrated solutions for molecular differential diagnosis. We are developing an integrated technology platform that allows multiplex molecular differential diagnoses carried out in a high-throughput, semi-quantitative, automatic, closed system. This high throughput diagnostic system relies on a multiplex polymerase chain reaction. We have developed a novel multiplex amplification strategy called amplicon rescued multiplex PCR (arm-PCR) that can be easily automated. The arm-PCR technology is roughly 100 times more sensitive than the previously developed tem-PCR method and is equally specific, providing an opportunity for the entire test procedure to be automated. This technology will allow physicians to diagnose microbial infection in just a few hours replacing current technology which sometimes takes several days.

For example, we’re developing a panel for influenza diagnosis. Rapid identification of the specific type of influenza virus causing infection is one of the most critical steps for controlling an outbreak and managing a pandemic. Such typing information provides healthcare professionals with data that can aid them in deciding which patients should be isolated and treated with Tamiflu, which should be vaccinated, and which should be released. These measures can limit the spread of the disease, ease public panic, and better allocate limited resources.

Another focus of my laboratory is mapping the personalized immunorepertoire.  By combining the last 30 years of improvements in monoclonal antibody development, multiplex PCR technology, high-throughput sequencing, and bioinformatics, our new technology has successfully overcome past challenges of studying the immunorepertoire.s. We are using this technology to analyze the immunorepertoires of autoimmune diseases, such as systemic lupus erythematosus, and various cancers through an international collaboration called R10K, or Repertoire 10,000, partnered with HudsonAlpha Institute for Biotechnology. By studying the immunorepertoire, we hope to uncover possible disease mechanisms, identify new biomarkers, and develop new therapeutics.

About Jian Han:

Jian Han, M.D., Ph.D., is a faculty investigator at the HudsonAlpha Institute for Biotechnology in Huntsville, Ala.. A native of China, he has a medical degree from Suzhou Medical College in JiangSu Province, China, and a Ph.D. from the University of Alabama at Birmingham. In 1996, Han founded biotech company Genaco, the first company to introduce a Down syndrome prenatal screening and diagnostic service to China. During the SARS outbreak of 2003, Genaco developed technology and products for molecular differential diagnoses of infectious diseases. Han created a novel multiplex PCR technology, tem-PCR, that allows multiple molecular targets to be amplified in one reaction. He developed another multiplex method called arm-PCR for infectious disease diagnosis and immune repertoire analysis. In 2009, Han founded two biotech companies — iCubate and iRepertoire — to commercialize arm-PCR based applications.

The most diverse of all human genes encode a set of proteins at the frontline of our immune system. Many different Human Leukocyte Antigen (HLA) proteins are encoded by genes clumped together in one portion of the human genome known as the major histocompatibility complex region. HLA proteins sit on the surface of cells and bind the chopped-up fragments of other proteins (antigens), presenting them for inspection by immune cells. If the presented antigens are recognized as foreign, the immune system may be triggered to attack, whether the invaders are pathogens, cancer cells, or transplanted tissue.

Remarkably, most HLA genes have dozens, or even hundreds of alleles present in the human population, so across the genome region as a whole there are thousands of different alleles. This variation can affect individual susceptibility to infectious and autoimmune diseases, and is of great interest to geneticists studying human evolution and population history.

But despite the functional and evolutionary importance of HLA genes, sequencing data from this region is biased in many population genomics studies. As a consequence, the results from this region are often treated as suspect, and in many cases are discarded from subsequent analyses.

The reason is that it’s difficult to make sense of HLA data generated by the next-generation sequencing (NGS) methods that are now standard for population genomics studies. NGS methods generate short sequence reads, and when these reads come from highly polymorphic genes like the HLA genes it can be challenging to correctly align them to the genome reference sequence. This problem is even worse when the gene is just one of a group of related polymorphic genes, as is the case for many of the HLA loci.

Genotyping errors for a highly polymorphic gene: The left hand side represents a case where sequence reads come from an individual who is heterozygous at a SNP, but where the rest of the gene is relatively similar to the reference for both haplotypes. The reads from both haplotypes can be aligned to the reference, and the SNP genotype is “called” (i.e. determined by the analysis software) correctly. The right hand side represents a case where one of the haplotypes is different to the reference sequence at more than one position. Reads from this haplotype won’t align with the reference and the genotype will be incorrectly called as homozygous at the SNP of interest. Image credit: Vitor R. C. Aguiar.

Though HLA loci are the worst-case scenario for this problem, other examples of polymorphic genes that come in related groups might suffer similar issues (such as the killer-like immunoglobulin receptor (KIR) and olfactory receptor genes). But because the degree of polymorphism in other gene families is less extreme than in the HLA genes, the analysis issues may be less obvious and therefore less likely to be accounted for.

In the latest issue of G3, Brandt et al. demonstrate the scale of the challenge using HLA data from the 1000 Genomes project, which is a collection of high-coverage exome and low-coverage whole-genome sequences from 1092 people generated by NGS. The authors compared the NGS data to a parallel dataset in which 930 of the samples from the 1000 Genomes project were re-sequenced using the “gold-standard” of Sanger sequencing, which doesn’t suffer from the same problems of short read alignment (the Sanger data were generated by Gourraud et al.)

Using the Sanger data as a benchmark, Brandt et al. showed that approximately 19% of single nucleotide polymorphism (SNP) genotypes for HLA genes in the NGS data were incorrect. And around a quarter of HLA SNPs had allele frequency estimates that differed between the two datasets by more than 0.1, with a bias towards overestimation of allele frequency in the NGS data. They also found that the most “unreliable” SNPs in NGS data were those with the highest heterozygosity. In other words, the SNPs at which people were mostly likely to be heterozygous were those that were most difficult to genotype correctly.

The results also suggest the NGS problem probably can’t be solved by boosting the intensity of sequencing efforts (i.e. increasing coverage). Rather, the authors’ argue that better computational analysis is the way forward. For example, they suggest that a major part of the problem is that standard approaches align reads to a single reference sequence. For HLA genes, and perhaps other polymorphic genes, alignment to a database of multiple reference sequences (for example, Boegel et al. and Dilthey et al.) can greatly improve genotyping accuracy by accounting for the different alleles possible at each gene.

A computational fix would be a boon to the many genetic studies that currently struggle to characterize HLA sequence data, including efforts to seek disease associations, quantify gene expression changes, and examine population histories. After all, the diversity of HLA genes is not only a technical challenge, but also a mark of their profound importance to immune system function and human survival.

Genotype mismatches between the 1000 Genomes (next-generation sequencing) and PAG2014 (Sanger sequencing) datasets. Results per polymorphic site (“Position”) and per individual. Dark squares indicate mismatches between genotypes in the two datasets. From Brandt et al.

CITATION:

Brandt, D.Y.C, Aguiar, V.R.C., Bitarello, B.D., Nunes, K., Goudet, J., & Meyer, D. (2015). Mapping Bias Overestimates Reference Allele Frequencies at the HLA Genes in the 1000 Genomes Project Phase I Data
G3: Genes|Genomes|Genetics, 5(5):931-941 doi: 10.1534/g3.114.015784
http://www.g3journal.org/content/5/5/931.full

Dramatic global declines in amphibians have been linked to the fungal pathogen Batrachochytrium dendrobatidis, but some species are more vulnerable than others. In the latest issue of G3: Genes|Genomes|Genetics, as part of the GSA Journals’ Genetics of Immunity collection, Ellison et al. examined the transcriptome of the highly susceptible Panama Golden Frog after exposure to the fungus.

The results showed rigorous innate and acquired immune gene expression, but they also showed indications of immunosuppression. Compared to naïve-infected individuals, previously-infected frogs showed significant increases in expression of fungal-killing genes like chitinase. The authors conclude that susceptibility is not necessarily due to a lack of immune response but a failure of those responses to be effective.

Read the article.

Read summaries of the latest group of Genetics of Immunity articles.

Browse the complete Genetics of Immunity collection.

CITATION:
Ellison A.R., G. V. DiRenzo, P. Langhammer, K. R. Lips & K. R. Zamudio (2014). Fighting a Losing Battle: Vigorous Immune Response Countered by Pathogen Suppression of Host Defenses in the Chytridiomycosis-Susceptible Frog Atelopus zeteki, G3: Genes|Genomes|Genetics, 4 (7) 1275-1289. DOI: http://dx.doi.org/10.1534/g3.114.010744

When Scott Alper and colleagues looked for candidate regulators of mammalian innate immunity based on gene expression data, the hit rate was only around 2%. By using a comparative genomics approach — starting with an RNAi screen in Caenorhabditis elegans — their hit rate rose to nearly 30%. In a new GENETICS article, published as part of the GSA Journals’ Genetics of Immunity collection, the team demonstrates the power of their method by identifying a factor that controls alternative splicing of Myd88, a critical signaling adaptor in many innate immunity signaling pathways in mammals.

Public Domain via Wikimedia Commons.

To protect us from disease, the innate immune system must be delicately regulated. A defective response leaves us vulnerable to infection, while over zealous activation can provoke or aggravate inflammatory diseases like atherosclerosis, asthma and cancer. To find new candidates for innate immune regulators, De Arras et al. screened over half of the C. elegans genome for genes that affect expression of a GFP-fused lectin regulated by several innate immune signaling pathways.

The team identified 20 mammalian orthologues of the C. elegans candidates, and then tested whether they also have regulatory function in a mouse macrophage cell line. They inhibited each gene with RNAi, stimulated the innate immune system and measured production of the pro-inflammatory cytokine IL-6 as an indicator of the innate immune response. Around half the candidates were implicated in immune regulation by this assay, and the effects of two of these genes were validated by demonstrating that their overexpression induced an effect opposite to that induced by RNAi-mediated inhibition.

One of these candidates, Eftud2, is associated with the U5 snRP and is involved in controlling mRNA splicing. The authors investigated whether Eftud2, like other known splicing factors, regulates alternative splicing of the signaling adapter MyD88. MyD88 acts downstream of most toll-like receptors (TLRs), key defense proteins that sense pathogens and danger signals. In contrast to its full-length counterpart which is a positive effector in TLR signaling pathways, a shorter splice variant of MyD88 negatively regulates TLR signaling. Eftud2 inhibition decreased the level of the long (activating) form of MyD88 and increased the level of the short (inhibitory) variant. Crucially, depleting the short splice form of MyD88 was able to substantially rescue the effect of Eftud2 inhibition, indicating that much of Eftud2’s effect is mediated by altered splicing of MyD88.

Genes with functions conserved all the way from nematode to mouse will likely play similar roles in humans, the authors argue, so the candidates emerging from this project may prove relevant to human infectious and inflammatory disease. Despite the millions of years of evolution that separate us from C. elegans, we can still glean valuable clues to human biology by turning to the worm.
Read the article.

Coral reefs around the world are “bleaching”, a threat caused by breakdown of the symbiosis between the coral animals (cnidaria) and the dinoflagellate algae that live within their cells. Yet this crucial symbiotic partnership is poorly understood. To address this knowledge gap, biologists are developing a suite of genomic tools for the sea anemone Aiptasia. This fast-growing cousin of corals maintains a similar symbiotic relationship with dinoflagellates, but it can also survive without its symbiotic friends.

In the February issue of G3: Genes|Genomes|Genetics, Lehnert et al. exploit this ability of Aiptasia to live without dinoflagellate symbionts to study gene expression patterns associated with the symbiotic state. They first had to assemble annotated transcriptomes for both Aiptasia and its dinoflagellate partner using RNA-Seq. They then compared transcript abundances between symbiotic and aposymbiotic (dinoflagellate-free) anemones. The authors identified nearly 1000 differentially expressed genes, in contrast to previous studies using less sensitive methods. These results provide important clues to how the symbionts regulate their relationship and will serve as a crucial genomic resource for future studies.

Read the article.

Lehnert E.M., M. S. Burriesci, N. D. Gallo, J. A. Schwarz & J. R. Pringle (2013). Extensive Differences in Gene Expression Between Symbiotic and Aposymbiotic Cnidarians, G3, 4 (2) 277-295. DOI: 10.1534/g3.113.009084http://www.g3journal.org/content/4/2/277.full