Researchers at the University of New Mexico and Duke University have discovered that, in contrast to prevailing assumptions, the immune system of an invertebrate animal - in their particular study a freshwater snail - has the ability to generate a diverse repertoire of defense molecules, despite lacking the lymphocytes that produce diverse antibodies and T-cell receptors found in vertebrates.
UNM researchers Si-Ming Zhang, Coen Adema and Eric Loker, and Duke’s Thomas Kepler recently announced the discovery based on several years of research. Their findings have been published in the latest edition of Science.
“It’s an exciting time for us, and in general for the field of comparative immunology because similar kinds of discoveries are coming to light in other organisms. Just this week the lead article in Nature was about a unique mechanism of immune diversification in lamprey eels” said Loker, chair of the Biology department and principal investigator for the project funded through a five-year, $1 million grant from the National Institutes of Health. “It was a coming together of minds - parasitologists, comparative immunologists and computational biologists - that led us to realize that snails produce defense molecules with a remarkably high level of diversification.”
The active diversification of defense genes has always been regarded as a characteristic separating vertebrate – including human - immune systems from invertebrates, explained Loker. Vertebrate immune systems are complex in part because of the active diversification of antibodies and T-cell receptors, but invertebrates lack those components, and their immune systems were thought to be correspondingly primitive.
Loker’s group studies the freshwater snail Biomphalaria glabrata, because of the role this snail plays in transmitting parasites that cause schistosomiasis in tropical regions, especially Brazil. Schistosome worms still infect 200 million people in underdeveloped countries. The cycle of infection begins when an infected human, often a child, excretes schistosome eggs into a freshwater pond or stream. The eggs hatch and release a larval stage that then infects B. glabrata snails. After about a month, thousands of cercariae begin to emerge from the infected snail and will penetrate the skin of a person bathing in the water. The parasites then mature into adult worms, with infection eventually damaging many vital organs in the body.
“Our original motivation was to learn more about the snail and how it defends itself from schistosome infection and how to stop the spread of the parasites,” said Loker. “No one fully understands how snails protect themselves from any pathogen or parasite.”
The researchers found that the snails possess in their blood a diverse family of proteins known as fibrinogen-related proteins, or FREPs. FREPs are unusual molecules in having at one end stretches of amino acids distantly related to those found in antibodies and at the other end, a region related to fibrinogen, the protein involved in blood clotting. FREPs increase in abundance following infection of snails and can bind to the surface of schistosome parasites. Molecules comparable to FREPs have not yet been found in other animals.
Further study of FREPs, even when retrieved from one snail, kept revealing more and more different sequences, something that was unexpected and not observed in other non-FREP control genes. The FREPs found in one snail were also found to be quite different from those of another. The variant FREP sequences were inferred to be derived from a small set of nine source sequences by point mutation and recombinatorial processes.
“It took about three years to figure out that the FREP sequences were being diversified in this unusual way,” said Adema, who has played an important role in revealing the original structure of FREPs and their puzzling variability. “We found patterns in our data sets indicating that a relatively simple organism like the snail was able to diversify molecules in a way likely to benefit its immune system. We turned to bio-informatics to gain a deeper understanding of the mechanisms of diversification.”
The project has generated more than 1,000 DNA sequences, which is where the computational aspect of the research has been vitally important to the findings.
Kepler’s contribution to the study was to use his expertise in DNA sequence analysis to recognize patterns in the mass of sequence data that had been generated. “It was clear that the genes in the dataset, though different, were closely related,” said Kepler. He devised a statistical technique for inferring the relationship among the genes and it produced a set of “very likely” reconstructions of their history. All of the reconstructed histories that were at all plausible showed a pattern: genes were swapping pieces of themselves back and forth.
To further test the implications of their findings, the group is now using a technique known as RNAi or RNA interference to knockout FREP genes.
“Our findings with RNAi are still preliminary, but we have indications that the technique is working,” said Zhang, the lead author on the Science paper. Zhang, who has been supported by UNM’s Center for Evolutionary and Theoretical Immunology (CETI), which is funded through the NIH COBRE program, notes, “We know diversification is occurring, but understand little about the mechanism or the importance of diversification to the snail’s survival. We’re using RNAi to begin to understand more about the functions of diversified FREPs.”
In the wake of this discovery made in snails, research by other groups that is now coming to light indicates that snails aren’t the only invertebrates with this ability. Sea urchins, shrimp and other species are showing signs of diversification, but using apparently different mechanisms.
“Invertebrates have much more capability for diversification than we gave them credit for,” said Adema. “Their innate defenses are more sophisticated and complicated than originally thought.”
“Our findings are changing our view of both innate and invertebrate immunity,” added Loker. “Invertebrates can do much more than anyone originally thought. We are grateful to the NIH for giving us the time to bring our research to fruition.”
Contact: Steve Carr, (505) 277-1821