virology blog

About viruses and viral disease

retrovirus

A retrovirus makes chicken eggshells blue

by

Araucana eggWhen you purchase chicken eggs at the market, they usually have white or brown shells. But some breeds of chicken produce blue or green eggs. The blue color is caused by insertion of a retrovirus into the chicken genome, which activates a gene involved in the production of blue eggs.

The Araucana, a chicken breed from Chile, and Dongxiang and Lushi chickens in China lay blue eggs. Blue eggshell color is controlled by an autosomal dominant gene: eggs produced by homozygote chickens are darker blue than those from heterozygotes. The gene causing blue eggshell color is called oocyan (O) and was previously mapped to the short arm of chromosome 1.

To further refine the location of the O gene, genetic crosses were performed using molecular markers on chromosome 1. The O gene was then located in a ~120 kb region which contained four genes. Only the SLCO1B3 was expressed in the uterus of Dongxiang chickens that produce blue eggs; it was not expressed in chickens that produce brown eggs.

Sequence analysis of the SLCO1B3 revealed that an endogenous avian retrovirus called EAV-HP has inserted just upstream of the gene. This insertion places a promoter sequence in front of the SLCO1B3 gene. As a consequence, the SLCO1B3 gene is transcribed. In chickens that produce brown eggs, no retrovirus is inserted before the SLCO1B3 gene, and no mRNA encoding the protein is produced.

The retrovirus insertion has occurred at different positions in the Chilean and Chinese chicken genomes. This observation indicates that the insertion arose independently during breeding of chicken strains several hundred years ago to produce blue egg layers. The chicken genome contains multiple copies of endogenous retroviruses, which can duplicate and move to other locations. We can assume that a random insertion upstream of the SLCO1B3 gene was selected for by breeding procedures that were aimed at producing blue egg-laying chickens.

The SLCO1B3 gene encodes a membrane transporter protein that mediates the uptake of a wide range of organic compounds into the cell. The blue eggshell color is produced by deposition of biliverdin on the eggshell as it develops in the uterus. Biliverdin is one component of bile salts, which are transported by SLCO1B3, providing a plausible hypothesis for the role of the protein in making blue eggshells.

Blue eggshell color is another example of the important roles that retroviruses have played in animal development. One other is the help provided by retroviruses in producing the placenta of mammals. Not all retroviral insertions are beneficial – integration next to an oncogene can lead to transformation and oncogenesis.

TWiV 242: I want my MMTV

by

On episode #242 of the science show This Week in Virology, the complete TWiV team talks about how two different viruses shape the evolution of an essential housekeeping protein.

You can find TWiV #242 at www.microbe.tv/twiv.

Dual virus-receptor duel

by

transferrin receptorViruses are obligate intracellular parasites: they must enter a cell to reproduce. To gain access to the cell interior, a virus must first bind to one or more specific receptor molecules on the cell surface. Cell receptors for viruses do not exist only to serve viruses: they also have cellular functions. An example is the transferrin receptor, which regulates iron uptake and assists in the entry of viruses from three different families. It might appear that such dual-use proteins cannot evolve to block virus entry because their cellular function would then be compromised. A study of two viruses that bind to the same cell surface receptor protein reveals how a cellular protein can change to prevent infection without affecting its role in the cell.

The virus-cell receptor interaction is one of the many arenas where the evolution of host-virus conflict can be studied. Because the virus-receptor interaction is essential for viral replication, host cells with a mutation in the receptor gene that prevents virus infection survive and eventually dominate the population. A virus could overcome this block with an amino acid change allowing binding to the altered receptor. Mutations that alter the interaction to favor the virus or the host are called ‘positively selected’ mutations. Such back-and-forth evolution between viruses and their host cells has been called host-virus arms races. Most have been identified by studying antiviral genes. This study is unusual in that it involves a housekeeping gene that has been usurped for viral attachment.

Evidence for positive selection of host genes can be detected by comparing gene sequences of phylogenetically related species. Nonsynonymous mutations lead to a change in the amino acid sequence, while synonymous mutations do not. The rate at which nonsynonymous mutations occur in the genome is typically much slower than synonymous mutations. The reason for this difference is that most mutations that change the amino acid sequence of a protein are lethal to the host. When genes have been subjected to positive selection by a virus, the ratio of nonsynonymous to synonymous mutations is higher, typically in host amino acids that interact with viral proteins. Computer programs have been designed to scan gene sequences and identify codons which are under positive selection by virtue of a high ratio of nonsynonymous to synonymous mutations.

To determine if the transferrin receptor (TfR1) has evolved to prevent virus attachment, sequences of the protein from seven different rodent species were compared. The analysis revealed that much of the protein is highly conserved, but a small part, comprising six amino acids, is evolving rapidly. Three of these amino acids  are located on the part of TfR1 that binds arenaviruses, and three are at the binding site for the retrovirus mouse mammary tumor virus (MMTV) (see illustration). Changing these three amino acids of TfR1 of the house mouse, which is susceptible to MMTV, to the sequence found in TfR1 of the MMTV-resistant vesper mouse, blocked entry of the virus into cells. In turn, changing these three amino acids of TfR1 of the MMTV-resistant short-tailed zygodont to the sequence of the house mouse enabled virus entry into cells. None of these changes had an effect on ferritin binding by TfR1.

Evidence for positive selection can also be detected in viral genes encoding proteins that interact with the host. The arenavirus glycoprotein, GP, is known to bind to TfR1. Ten GP amino acids were identified that are under positive selection, and four of these directly contact TfR1.

These findings demonstrate that there has been an arms race between TfR1 and both an arenavirus and retrovirus. An interesting question is whether human TfR1 will enter into an arms race with arenaviruses. As these viruses emerge into the human population, it is expected that humans with mutations that make them less susceptible to infection or severe disease will be positively selected. Amino acid 212 of human TfR1, which is near the positively selected resides in murine TfR1, varies in the human population. When this amino acid change (leucine to valine) is introduced into TfR1, it confers some protection against arenavirus entry. Curiously, this polymorphism has only been found in Asian populations, where arenaviruses that bind TfR1 are not found. The polymorphism is probably neutral with respect to TfR1 function, and if TfR1-binding arenaviruses are introduced into Asia, this change could be positively selected.

Because all viruses depend on many host proteins for replication, it will be interesting to use this approach to see how other highly conserved cell proteins balance cell function with the ability to resist virus infections. There are like to be many cell proteins that cannot change to evade viral use without destroying their cell function. Fortunately for cells there are exceptions.

Museum pelts help date the koala retrovirus

by

friendly-male-koalaThe genomes of most higher organisms contain sequences from retroviral genomes called endogenous retroviruses (ERVs). These are DNA copies of retroviral RNAs that are integrated into the germ line DNA of the host, and passed from parent to offspring. In most species the infections that lead to germ line ERVs appear to have occurred millions of years ago. The Koala retrovirus, KoRV, is the only retrovirus that we know of that is currently invading the germ line of its host species. A study of Koala pelts preserved in museums suggests how recently the virus infected this animal.

The koala is native to Australia, and all koalas in northern Australia are infected with KoRV. However not all animals in the southeast or on southern islands are infected. It is believed that KoRV crossed into koalas from another species (possibly the Asian mouse Mus caroli) some time within the past two hundred years. To test this hypothesis, DNA was extracted from 28 koala skins that were held in museums and which had been collected from the late 1800s to the 1980s. Polymerase chain reaction was used to detect KoRV DNA in the koala genome. The results show that KoRV was already widespread in Northern Australian koalas by the late 1800s. It has since spread slowly because the virus is not ubiquitous in southern koalas. The slow dispersal may due to the sedentary and solitary nature of koalas. Examination of mitochondrial DNA from the koala skins confirmed that there has been limited movement of the animals with Australia.

The sequence of the KoRV gene encoding the viral glycoprotein, env, was also determined. The results reveal that env sequences from museum specimens are remarkably similar to those of KoRV found in contemporary koalas. At first glance this result might not seem surprising: the endogenous KoRV genomes are evolving at the same slow rate as the host DNA into which they are integrated. However, there appear to have been multiple transmissions and germ line invasions by KoRV, leading the authors to suggest that in all cases very similar retroviruses were involved.

Infection with KoRV in captive animals is believed to cause immunosuppression, leading to fatal lymphomas or Chlamydia infection. A Chlamydia epidemic is believed to have killed many koalas in 1887-1889, consistent with the PCR results indicating that KoRV was widely present at that time.

Update: I had meant to discuss the possibility of dating the invasion of Koalas by KoRV by using older samples, but neglected to include this in the original article. Several days after it was published, Professor Paul Young sent me a note expressing exactly this sentiment:

What would be even better would be to have access to fossilised material that predates European settlement, that we could examine. We collaborated with an “Ancient DNA” specialist and tried this several years ago but we weren’t able to recover usable template DNA. Still worth some future effort though.

Avila-Arcos MC et al (2012). 120 years of koala retrovirus evolution determined from museum skins. Mol Biol Evol. 2012 Sep 14.

TWiV Special: A paradigm for pathogen de-discovery

by

On this special episode of the science show This Week in Virology, Vincent and Ian review a multicenter blinded analysis which finds no association between chronic fatigue syndrome/myalgic encephalomyelitis and XMRV or polytropic murine leukemia virus.

You can find this TWiV Special at www.microbe.tv/twiv.