A research study from the Massachusetts General Hospital (MGH) has identified a gene malfunction that appears to be central to the development of type 1 diabetes. The study in a classic animal model of type 1 diabetes found that a gene required to help teach the immune system to recognize so-called "self" proteins is somehow inactivated, even though its sequence is not mutated. The scientists also found that the inactivation of this gene called Lmp2 has a profound impact on another key protein called nuclear factor kappa-B (NF-kB), a major controller of several immune system activities. Finding ways to correct or circumvent the malfunction identified in this study may lead to ways of preventing or stopping the autoimmune reaction at the heart of type 1 diabetes.
"This is a true genetic abnormality we can associate with type 1 diabetes," says Denise Faustman, MD, PhD, of the MGH Diabetes Unit, who led the study appearing in the December issue of Molecular and Cellular Biology. "While Lmp2 is located in a portion of the genome that had been linked to the disease, it hadn't been considered a candidate because its sequence is normal. But studying the gene's function has showed us a previously unsuspected problem: In the immune system cells we looked at, the Lmp2 protein virtually disappears by the time these mice reach a certain age, therefore most likely initiating their diabetes."
Type 1 diabetes (also called juvenile or insulin-dependent diabetes) is an autoimmune disease in which the immune system mistakenly attacks the insulin-producing beta cells of the pancreas. Normally, immune system components called T cells learn not to attack the body's own tissues through the action of MHC Class I molecules, which sit on the surface of B cells and macrophages. There the molecules display tiny bits of "self" proteins to the T cells, the immune system's killers, essentially teaching them to ignore the body's own tissues. Another kind of MHC molecule, called Class II, displays proteins from invading microorganisms.
In 1991, Faustman and her colleagues discovered that macrophages from children at high risk for developing diabetes had significantly reduced levels of correctly assembled MHC Class I molecules on their surfaces. Instead of containing a peptide - a protein fragment - to display to T cells, many of the molecules were empty. This observation helped to establish Class I's role in developing immune tolerance to self proteins and was confirmed by later studies.
But Faustman's work has been controversial because decades of genetic studies suggested that type 1 diabetes was associated with a single abnormality in the portion of the genome responsible for encoding MHC Class II molecules. Her team believes that the actual problems arise with proteins that play key roles in the processing of self-peptides for display in MHC Class I molecules - proteins whose genes lie in the middle of the Class II genes. Two of the proteins the researchers have been focusing on, called Lmp2 and Lmp7, are part of a cellular structure called the proteasome, which breaks larger proteins down into their peptide components either for disposal or for delivery to the Class I molecules.
In the current paper, Faustman and her postdoctoral fellow Takuma Hayashi, PhD, studied immune system cells from NOD (non-obese diabetic) mice, a classic animal model for type 1 diabetes. They found that, although NOD mouse embryos had normal levels of Lmp2 protein in macrophages and other immune cells, mice that were 6 to 8 weeks old lacked Lmp2 protein in corresponding cells. "This almost complete shutdown of protein production is very rare," Faustman explains. "The finding suggests that the problem is not in the DNA segment that codes for the protein - which appears totally normal - but in other factors that control how DNA first is transcribed into RNA and then translated into a protein."
Lacking the Lmp2 protein, the cells studied have defective proteosomes, which means they cannot properly produce the self peptides to be displayed on MHC Class I moledules. If the T cells are not exposed to important self peptides via the Class I system, they may mistakenly attack those peptides on the body's organs, causing autoimmune disease. In addition, functioning proteasomes - and particularly the Lmp2 portion - are required for production and activation of NF-kB, a major regulator of immune system function. Faustman and Hayashi found that key activities of NF-kB are reduced in the NOD mouse lymphocytes and that, among other abnormalities, the cells are more sensitive to a specific type of cell death usually protected against by NF-kB.
"Finding the connection with NF-kB was unexpected and quite exciting," Faustman says. "It's one of the most important proteins we know about for controlling major immune system activities, but no one suspected it could be involved in diabetes or other autoimmune diseases. These findings are bringing seemingly unrelated areas of basic research together to address one of the most significant human diseases. They almost bring us full circle on the complex genetics of this disease, because the Lmp2 gene, while it is critical to Class I processing, is located in the Class II region of chromosome 6, right where people have been searching for diabetes genes."
Faustman says the next steps in this investigation include confirming whether the same deficit in Lmp2 protein production occurs in human diabetes patients and searching for ways to restore the disrupted cellular processes. The current study was supported by grants from The Iacocca Foundation and the National Institute of Child Health and Human Development.
Journal
Molecular and Cellular Biology