David N. Leff
Editor's note: Science Scan is a roundup of recently published, biotechnology-relevant research.
Antibiotics, if delivered promptly, can usually cut short the agonizing, whole-body symptoms of Lyme disease, perpetrated by its bacterial spirochete, Borrelia burgdorferi. But now and again, a victim who congratulates himself and his physician for aborting the infection is brought to his knees by the Lyme backlash. The patient's knees swell up and hurt, weeks or months after that antibiotic treatment apparently did its job. Those painful joints may look for all the world like rheumatoid arthritis (RA), an autoimmune disease unrelated to Lyme borreliosis.
Autoimmunity is the immune system's treacherous attack on its own cells and tissues - in the case of RA the antigenic synovial fluid that lubricates joints. Spearheading that onslaught are cell-slaying T cells, programmed to target antigens, foreign or domestic, propelled to their attention by the MHC - major histocompatibility complex. For Lyme disease research, the preferred B. burgdorferi antigenic target is the pathogen's outer surface peptide - Osp.
However, arthritis in the wake of Lyme infection is thought to arise when B. burgdorferi develops autoimmunity, reminiscent of the RA mechanism. A paper in the Proceedings of the National Academy of Sciences (PNAS), dated Oct. 10, 2000, implies that autoimmunity may be implicated. Its title: "Direct enumeration of Borrelia-reactive CD4 T cells ex vivo by using MHC class II tetramers." Its senior author is research immunologist and clinical neurologist David Hafler, at Harvard-affiliated Brigham and Women's Hospital in Boston.
"It has nothing to do with antibiotic resistance," Hafler told BioWorld Today. It implies an autoimmune-mediated tissue destruction in the joints. What causes Lyme arthritis," he went on, "is a major issue. But there's a larger issue in the PNAS paper, which is: How does one measure antigen-reactive T cells? It's one of the most fundamental questions in clinical immunology. The problem," he explained, "is that if you take one MHC complex, displaying an antigen - in this context, OspA - that is recognized by the T-cell receptor, the affinity is too low to bind to that T cell. So we made that monomeric MHC complex into a tetramer - a cluster of four MHC molecules linked together - presenting a highly antigenic, 12-base-pair fragment of that OspA target, which greatly increased the affinity. For the first time, we could get recognition of antigens, or binding of this complex to T cells, in a very specific fashion.
"To detect that binding in a patient with arthritis," Hafler recounted, "we took cells from the synovial fluid of six patients who had antibiotic-resistant arthritis secondary to Lyme disease, and showed that our MHC tetramer complex could directly identify the Lyme-reactive T cells in the synovial fluid.
"The frequency of T cells that reacted to the OspA protein in the fluid was at a pretty high level - about 3.5 percent. Particularly in infectious and autoimmune diseases," Hafler pointed out, "it's important to be able to quickly measure what the T cells are recognizing. Rather than taking two or three weeks to be able to do that, our tetramers can do it in 30 minutes - and much more accurately."
Hafler foresees extending this tetrameric method to diagnosis and monitoring of other autoimmune diseases. "For multiple sclerosis," he said, "you'd want tetramers loaded with myelin antigens. For Type II diabetes, with pancreatic islet antigens. The main thing we're now pursuing," he concluded, "is testing these tetramers for immune tolerance in clinical situations."
300,000 SNPshots Of Gene Base Variants Will Complement Complete Human Genome Sequence
When finally fully sequenced, the human genome is guesstimated at containing 60,000 to 100,000 genes. That nearly completed scorecard of humankind's genetic endowment needs to be able to detect the individual variations in genes. These either/or alleles reflect the genomic differences between an individual's two parents - notably complex traits such as disease susceptibility and response to drugs. Most sequence variants are single nucleotide polymorphisms - sites on DNA where two alternate bases (A, C, T and G) occur at one position.
The SNPs tell their story.
The SNP Consortium, an international collaboration of academic centers, pharmaceutical companies and a private foundation, aims to discover and release at least 300,000 human SNPs. So far, it has identified 148,459. Of that number, 47,172 employed a novel technology called RRS.
The Sept. 28, 2000, Nature describes it in a paper titled, "An SNP map of the human genome generated by reduced representation shotgun [RRS] sequencing." A companion paper in the same issue of Nature, by participants in the consortium, reports: "An SNP map of human chromosome 22." It contains 2,730 SNPs.
Chromosome 22's long arm alone carries genes responsible for a long list of familial disorders - notably, congenital heart disease, chronic myeloid leukemia, schizophrenia, as well as some breast, ovarian and colon cancers.
Structure of Bacterium That Feeds On CarbonMonoxide Elucidates Gene On/Of Switching
A freshwater bacterium has disclosed for the first time how transcription factor proteins change shape in order to bind with DNA and turn genes "on" and "off." The bacterium, Rhodospirillum rubrum, inhabits lakes and ponds, and turns sunlight into energy the way photosynthetic green plants and algae do.
When it can find no sunlight, R. rubrum senses carbon monoxide (CO) - its nutrient source - and triggers a set of genes that allows it to consume this energy supplier. The protein catalyst that jump-starts this process is a transcription factor called CooA. When it senses CO, CooA turns on by changing its molecular structure, which allows it to bind DNA.
CooA wears two hats. It belongs to the family of catabolic activator proteins, best known as CAP. It's also a heme protein, the oxygen-carrying component of hemoglobin. Its CO connection accounts for that gaseous compound's oxygen-robbing toxicity.
Both hats figure in an article in Nature Structural Biology for October 2000. Its title: "Structure of the CO sensing transcription activator CooA." A News & Views commentary is headed, "CooA, CAP and allostery." The paper reports how X-ray crystallography exposed "the first look at what the structural changes from off to on involve."