Antibody-producing cell. B-lymphocyte or T-lymphocyte in blood with red blood cells.
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The serine-threonine protein phosphatase 2A (PP2A) enzyme is critical for regulatory T cells to function—without it, they don’t have the ability to suppress effector T cells and can’t protect against autoimmunity, according to new research published in Nature Immunology.
Researchers found that conditional knockout mice—in which PP2A expression is knocked out only in regulatory T cells—developed a severe lymphoproliferative disorder affecting just about every organ. The findings—the latest layers of the onion peeled away in the lab of George Tsokos, MD, chief of the Rheumatology Division at Beth Israel Deaconess Medical Center, Boston—offer a potential new target for the treatment of systemic lupus erythematosus.
Dr. Tsokos says the findings bring critical new insight into how regulatory T cells do the job of regulating the immune system.1
“In this paper, we saw that it is one enzyme, one phosphatase, that is absolutely needed for the regulatory T cell to be functional,” he says. “And we understand the molecular underpinning for the good function of regulatory T cells.”
PP2A is known to regulate key cellular processes, such as cell-cycle progression, apoptosis, cellular metabolism and migration, and is involved in the development of cancer and neurodegenerative diseases, as well as SLE.
The new understanding about the enzyme is another piece of the puzzle in the complex pathogenesis of SLE, Dr. Tsokos says. Genome-wide association studies have identified genetic susceptibility to the disease and identified more and more people in whom a deficiency in a single gene is responsible for expression of SLE. Smoking, sun exposure and other environmental factors have an epigenetic influence. Hormones clearly play a role, as well, he adds—nine out of 10 people with SLE are women—although the role of hormones is not very well understood.
And then there is abnormal immune regulation.
“All these factors contribute to the expression of the disease, and I would say they act either serially or all together, and in each patient, these factors contribute differentially,” Dr. Tsokos said. “One patient may have more genetic, another patient more epigenetic and so on and so on. … All of these factors acting together generate what I call ‘common effectors of the immune response’: autoantibodies, immune complexes, autoreactive T cells and pro-inflammatory cytokines.” Additionally, he says, organ-specific factors determine which organ will be subject to inflammation and damage.
Long-Term Research
Researchers in Dr. Tsokos’ lab are interested in identifying molecular abnormalities in the immune system and in the organs and then trying to specifically target them. They have been studying T cells in SLE for more than three decades.
They began by examining the inability of SLE T cells to produce interleukin 2 (IL‑2), one of the most important cytokines of the immune system. They eventually found that CREB, a transcription factor responsible for the enhancement of IL-2 production, is insufficient in these cells.2
Whey they searched for the causes, they found that SLE T cells have increased PP2A, and that its increase is controlled both genetically and epigenetically and some people are more prone to express more PP2A. They also found that PP2A was central in regulating the activity of other molecules important in regulating T cell function. Researchers then turned to constructing a mouse that overexpresses PP2A only in T cells and found the mouse was highly prone to developing glomerulonephritis.
When they began to look at PP2A in different T cell subsets, they found that regulatory T cells highly express PP2A. They then designed a mouse with regulatory T cells that do not express PP2A.
PP2A in the Crosshairs
Sokratis Apostolidis, MD, the paper’s lead author, who was a research fellow at Beth Israel at the time and is now a resident at the University of Pittsburgh, says the suppression of PP2A had to be handled with great precision.
“PP2A is a highly conserved enzyme from humans all the way to yeast and ubiquitously expressed in the cells of the mammalian body. As a result, and in order to specifically study the role of PP2A in Tregs, we had to generate genetically engineered mice that would lack PP2A activity only in Treg cells. This targeted approach gave us the ability to exclude the effects of PP2A resulting from its deficiency in non-Treg immune cells as well as the rest of the cells of the mice that could confound our findings,” he says.
“The second biggest consideration we needed to take into account was that PP2A is a multi-protein complex and targeting the correct subunit of this complex in Treg cells was crucial. We wanted to cripple the activity of the PP2A enzyme, but at the same time preserve cell viability, because PP2A is involved in the cell cycle as well. That gave us the chance to study live PP2A-deficient Treg cells.”
They found that these mice lacking PP2A in Treg cells developed severe disease.
“This mouse had a lot of inflammation in practically every organ. And continuously it was producing a lot of autoantibodies,” Dr. Tsokos says. These autoantibodies were “practically against everything,” he says, and they included anti-nuclear antibodies, which are common in SLE.
This was occurring, he says, despite the expression of Foxp3, a transcription factor involved in immune suppression.
“Tregs from this mouse did not have the ability to suppress the function of effector cells—they were expressing Foxp3 but could not function,” he says. “In some ways, we saw that PP2A is there to muscle up Treg cells and if you don’t have PP2A, the cells become emasculated.”
PP2A is known to regulate key cellular processes, such as cell-cycle progression, apoptosis, cellular metabolism & migration, & is involved in the development of cancer & neurodegenerative diseases, as well as SLE.
When looking into the ways PP2A is controlled in regulatory T cells, they found that Treg cells express high amounts of the lipid ceramide—a result of Foxp3 suppressing the production of the enzyme SGMS1, or sphingomyelin synthase 1, Dr. Tsokos says.
This further clarifies the chain of events taking place within regulatory T cells, he says. Without SGMS1, you end up with a lot of ceramide, which activates PP2A, which then inactivates the mTOR signaling pathway, which is responsible for the inflammatory response.
“If you don’t have enough ceramide, you don’t make active PP2A, and the mTOR component of the cell takes over, and it deprives it of its ability to suppress effector cells,” Dr. Tsokos says.
mTORC1 Effects
The researchers’ findings on exactly how PP2A affects the mTOR pathway could be particularly important. They found that when PP2A is activated in Treg cells, there is a distinct effect on the mTORC1 complex, but not on mTORC2. The mTORC1 complex has been shown to promote immunity.
The lab of Andras Perl, MD, PhD, chief of Rheumatology at SUNY Upstate Medical Center, has shown that mTORC1 is increased in SLE T cells compared with those in healthy controls.
When PP2A is activated in Treg cells, there is a distinct effect on the mTORC1 complex, but not on mTORC2. The mTORC1 complex has been shown to promote immunity.
Dr. Perl says the findings from Dr. Tsokos’ lab mark significant progress.
“This is a well-designed study [that] is presented with great precision,” he says. “Due to the clarity of the findings, it is a major step forward.”
He was especially struck by the way the findings dovetailed with findings from his own lab. “My laboratory has uncovered the activation of mTORC1 complex 1 in lupus T cells and we also showed that blockade of mTORC1 with rapamycin or N-acetylcysteine (NAC) provides clinical benefit within the context of clinical trials,” he says. “With direct relevance to this paper, we showed that blockade of mTORC1 by both rapamycin and NAC expanded regulatory T cells in SLE patients in vivo. The current paper suggests that PP2A may act through the very same pathway, and it confirms the importance of mTORC1 in controlling regulatory T cell function in SLE.”
Bottom Line
Developing therapies based on these new findings is likely still far down the road, Dr. Tsokos cautions.
“I think it’s pretty early right now, and we would need to find ways to enhance the activity of this enzyme and to do so only in regulatory T cells and not in the rest of the cells,” he says. With PP2A composed of many specific regulatory subunits, a key to any therapy would likely be altering activity at the subunit level, he says.
“By altering the function of a subunit, you can change the function of this enzyme only in this specific cell and not affect its function, its expression, in other cells,” he says. This is a “hot direction” where the lab is now heading, he says. The lab is also developing mouse models that are PP2A deficient in other immune cell subsets.
The finding that PP2A interacts with the mTOR pathway, and in particular decreases the activity of mTORC1, could be particularly useful in clinical application, Dr. Apostolidis says.
“mTOR is a central hub of cellular input integration implicated in metabolism, cell proliferation and cytokine production—it is the focus of intense research in immunology, diabetes, transplant and cancer. By identifying PP2A as an important upstream regulator of mTOR activity, we provided an additional layer of control that can be targeted to modulate mTOR function,” he says. “At the moment, PP2A-targeting drugs are being developed in cancer [research]. Based on our recent work, these drugs can also be used to modify PP2A activity in autoimmune diseases such as SLE, RA, multiple sclerosis and IBD [inflammatory bowel disease].”
He says further studies will be needed to determine whether preferential delivery of PP2A to immune cells is necessary to avoid off-target effects and to maximize the effectiveness on Treg cells.
A more immediate clinical impact of the findings could stem from the observation that ceramides—“the good lipids,” as Dr. Tsokos calls them—are increased in well-functioning Treg cells.
He stopped short of making any specific dietary recommendations, but says, “If you come to think of ways to reconstitute the good lipids, the ceramides, that Tregs need to function well, that will be an advance.”
Thomas R. Collins is a medical writer based in Florida.
References
- Apostolidis SA, Rodriguez-Rodriguez N, Suarez-Fueyo A, et al. Phosphatase PP2A is requisite for the function of regulatory T cells. Nat Immunol. 2016 Mar 14 [Epub ahead of print]
- Sunahori K, Juang YT, Tsokos GC. Methylation status of CpG islands flanking a cAMP response element motif on the protein phosphatase 2Ac alpha promoter determines CREB binding and activity. J Immunol. 2009 Feb 1;182(3):1500–1508.
- Kato H, Perl A. Mechanistic target of rapamycin complex 1 expands Th17 and IL-4+ CD4-CD8- double-negative T cells and contracts regulatory T cells in systemic lupus erythematosus. J Immunol. 2014 May 1;192(9):4134–4144.
