In 2012, two independent studies found that the FLCN-interacting protein FNIP1 is required for proper β-cell maturation, and were the first studies to suggest a role for FLCN and its partners in the immune system. Following their 2012 discovery that FNIP1 is necessary for β-cell development (Park et al., 2012), the same group decided to further elucidate the role of FNIP1 in the immune system. Park et al., (2014) analysed the phenotype of multiple immune cells in FNIP1-null mice, and found that FNIP1 is also required for the correct development and maturation of a specific subset of T-cells called invariant Natural Killer T cells (iNKT cells) (reviewed by Cianferoni, 2013).
iNKT cells are a subset of T cells that express an invariant T cell receptor and can recognise lipid bound antigens, such as glycolipids, on the surface of other cells. When activated, these cells produce cytokines to illicit an immune response. iNKT cells can recognise bacterial infections, viruses, and tumours. Their development is highly regulated, alternating between TCR rearrangement, proliferative and maturation phases, starting at stage 0, where DP thymocytes become committed to the iNKT cell lineage, and fully mature stage 3 iNKT cells.
Park et al. found that iNKT cells became arrested during stage 1 and 2, with few mature stage 3 cells found in FNIP1-null mice. Additionally, PLZF – which regulates iNKT cell development – was overexpressed in FNIP1-null iNKT cells. Bone marrow transplants into nude mice using a 1:1 mixture of wild-type and FNIP1-null cells showed that β-cells and iNKT cells were derived from wild-type mice only, whereas CD4 and CD8 thymocytes were derived from both types of donor cell. This suggests that FNIP1-null iNKT cells arrest during development due to cell autonomous effects, rather than due to a defective environment.
BrdU pulse experiments showed that FNIP1-null cells over-proliferated and subsequently died in early stage 3, as shown by an increase in Caspase 3-positive cells. Furthermore, FNIP1-null iNKT cells showed reduced mitochondrial mass in stage 1, decreased levels of ATP, larger cell size and increased mTOR signalling. Together, this suggests that dysregulated mTOR signalling leads to higher energy consumption, meaning that cells do not have the required energy reserves for proliferation and maturation, and die between stage 2 and 3.
However, mTOR dysregulation is not fully responsible for this phenotype, as in vivo treatment of pups, beginning in utero, did not rescue the iNKT cell phenotype. AMPK, which FNIP1 binds, activates the proautophagy gene Vps34, which is also required for iNKT cell development in mice (Parekh et al., 2013). Thus, concomitant loss of autophagy and mTOR dysregulation might explain the loss of metabolic homeostasis in FNIP1-null iNKT cells.
These results are similar to those reported in FNIP1-null β-cells, which were reported to be due a loss of metabolic homeostasis (Park et al., 2012) and increased apoptosis (Baba et al., 2012). They also correspond well with recent data showing that loss of FLCN leads to metabolic transformation in cells. However, it is currently unclear why loss of FNIP1 leads to such specific phenotypes in the immune system, only affecting a subset of lineages. It would be interesting to determine whether FLCN and its other binding proteins – FNIP2, PKP4, RPT4 and the Rag proteins – control the development of other immune cell lineages. If so, this would provide compelling evidence to support previous observations that FLCN function changes in different cell types (Hudon et al., 2010).
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- Cianferoni A (2014). Invariant Natural Killer T Cells. Antibodies, 3 (1), 16-36 doi: 10.3390/antib3010016
- Hudon V, Sabourin S, Dydensborg AB, Kottis V, Ghazi A, Paquet M, Crosby K, Pomerleau V, Uetani N, & Pause A (2010). Renal tumour suppressor function of the Birt-Hogg-Dubé syndrome gene product folliculin. Journal of medical genetics, 47 (3), 182-9 PMID: 19843504
- Parekh VV, Wu L, Boyd KL, Williams JA, Gaddy JA, Olivares-Villagómez D, Cover TL, Zong WX, Zhang J, & Van Kaer L (2013). Impaired autophagy, defective T cell homeostasis, and a wasting syndrome in mice with a T cell-specific deletion of Vps34. Journal of immunology (Baltimore, Md. : 1950), 190 (10), 5086-101 PMID: 23596309
- Park H, Staehling K, Tsang M, Appleby MW, Brunkow ME, Margineantu D, Hockenbery DM, Habib T, Liggitt HD, Carlson G, & Iritani BM (2012). Disruption of Fnip1 reveals a metabolic checkpoint controlling B lymphocyte development. Immunity, 36 (5), 769-81 PMID: 22608497
- Park H, Tsang M, Iritani BM, & Bevan MJ (2014). Metabolic regulator Fnip1 is crucial for iNKT lymphocyte development. Proceedings of the National Academy of Sciences of the United States of America, 111 (19), 7066-71 PMID: 24785297
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