User:Svarberb/PTPN22

Protein tyrosine phosphatase non-receptor type 22 (PTPN22) is a cytoplasmatic protein encoded by gene PTPN22 and a member of PEST family of protein tyrosine phosphatases. This protein is also called "PEST-domain Enriched Phosphatase" ("PEP") or "Lymphoid phosphatase" ("LYP"). The name LYP is used strictly for the human protein encoded by PTPN22, but the name PEP is used only for its mouse homolog. However, both proteins have similar biological function and show 70% identity in amino acid sequence. PTPN22 function as a negative regulator of T cell receptor signaling, which maintains homeostasis of T cell compartment.

Gene
Gene PTPN22 is located on the p arm of the human chromosome 1. It is nearly 58 000 base pairs long and contains 21 exones. In case of mouse genome It is located on the q arm of the chromosome 3. It is nearly 55 700 base pairs long and contains 23 exons.

Structure
PTPN22 is composed from 807 amino acids and It weights 91,705 kDa. On its N terminus It possess catalytic domain, which shows the highest level of conservation between human and mouse proteins. Other parts of PTPN22 are less conserved. After catalytic domain PTPN22 has approximately 300 amino acids long domain called "interdomain". On the C terminus of PTPN22 there are 4 prolin-rich motifs (P1 - P4), which can mediate interactions with other proteins. P1 motif is the most important among them, because it is crucial for binding of CSK kinase and allele encoding PTPN22 with mutated P1 motif is associated with increased risk of numerous autoimmune diseases.

Regulation of TCR signaling
Activation of T cell receptor by cognate peptide trigger signaling pathway activating T cell. First events of this pathway is activation of SRC family kinase LCK by dephosphorylation of its C terminal inhibition tyrosine (Y505) and by phosphorylation of its activation tyrosine (Y394). LCK then phosphorylate tyrosines in the CD3 complex creating a docking site for SH2 domain of SYK family kinase ZAP70, which is there phosphorylated too. Phosphorylated ZAP70 then propagate signal from TCR, phosphorylating other proteins and creating multi-protein complex, which activates downstream signaling pathways. PTPN22 possess the ability to dephosphorylate proteins included in proximal events of TCR signaling and serves as an important negative regulator of T cell activation. PTPN22 is able to bind LCK with phosphorylated Y394, phosphorylated ZAP70 and phosphorylated ζ chain of CD3 complex. Thus, it binds molecules of proximal TCR signaling only after their activation. PTPN22 can dephosphorylate those proteins and decrease the activating signal obtained by T cell. Dephosphorylation of LCK and ZAP70 by PTPN22 is specific concerning the phosphorylated tyrosine in those proteins - only Y394 of LCK and Y493 of ZAP70 are dephosphorylated. In the absence of PTPN22 activated T cell receive stronger activation signal, which is reflected by greater influx of Ca2+ cations into cytosol, bigger phosphorylation of LCK, ZAP70 and ERK and bigger expansion of those cells. Inhibitory effect on TCR signaling was also verified with the usage of PTPN22 inhibitor on Jurkat T cell line and on human primary T cells, and also with the experiments of PTPN22 overexpression in vitro. Expression of PTPN22 is upregulated after the activation of T cells and antigen-experienced T cell have higher expression of PTPN22 than naive T cell.

The regulatory function of PTPN22 is particularly important during activation by low affinity peptides. In the absence of PTPN22 T cell cannot discriminate between strong and weak antigen sufficiently and those T cells become more responsive, which can be detected like increased upregulation of transcription factors, CD69, increased ERK phosphorylation, increased ability to expand in vivo and to produce cytokines. Increased responsiveness can also break tolerance against low affinity self-antigens and is well visible when PTPN22-deficient T cells get into a lymphopenic environment.

Regulation of regulatory T cells
One particular population of T cells, which is influenced by PTPN22 deficiency is the population of regulatory T cells (Treg cells). PTPN22-deficient mice contain bigger amount of Treg cell in lymph nodes and spleen and this difference is more visible with increasing age of mice. There is also a change of the effector Treg cells : central Treg cells ratio in favor of effector Treg cells. PTPN22 deficiency increases the abilities of Treg cells to survive, differentiation of Treg cell from naive T cells, but not the ability to proliferate in vivo, and It also supports transition of central Treg cells to effector Treg cells. One of the reason of increase survival of PTPN22-deficient Treg cells is that those cells have upregulated expresion of GITR, which increases their expansion in vivo. Treatment of PTPN22-deficient mice with anti-GITR-L blocking antibody suppress the expansion of Treg cells. PTPN22 deficiency does not impair suppressive function of Treg cells. Actually there are some articles suggesting that PTPN22-deficient Treg cells possess enhanced suppressive function or have bigger ability to obtain effector phenotype.

PTPN22-deficient naive T cell diferentiate into Treg cell more effectively in vitro, when they are stimulated by low concentrations of anti-CD3 and anti-CD28 antibodies. With increasing amount of antibodies the ratio of Treg diferentitation becomes lower than in case of Wild Type naive cells. (find the citation)

Regulation of Adhesiveness and motility
Next to the TCR signaling PTPN22 regulates an adhesiveness and motility of T cells. PTPN22-deficient T cells have prolonged interval of contact with antigen presenting cell, which present low affinity peptide. With high affinity peptide the difference is not detectable. Part of the reason of increased adhesiveness of those T cells is that enhanced TCR signaling results in higher activation of RAP1 and boosted inside-out signaling to activate the adhesive molecule LFA-1. In the migrating T cells we can see polarized localization of PTPN22 at the leading edge of migrating T cell, where it colocalizes with its substrates LCK and ZAP70. Downregulation of PTPN22 increased motility, adhesivity and levels of phosphorylated LCK and phosphorylated ZAP70 in those cells. On the contrary, overexpression of PTPN22, but not catalytically inactive PTPN22, increased motility of migrating T cells. Association of PTPN22, but not its disease associated mutant form, with LFA-1 results in decreased LFA-1 clustering and adhesion. Role of PTPN22 in regulation of LFA-1-mediated adhesion and motility is also supported by the observation of increased LFA-1 expression on PTPN22-/- Treg cells.

Among other putative proteins related to TCR signaling which can be dephosphorylated by PTPN22 belong C-CBL, VAV-GEF, BCR-ABL, SKAP-HOM and VCP. (ověřit)

Interaction partners
C-terminal part of PTPN22 bare prolin-rich motifs providing binding sites for putative interaction partners. One of those interaction partners is cytoplasmatic tyrosine kinase CSK, which is a negative regulator of SRC family kinases and TCR signaling as well as PTPN22. CSK binds two prolin-rich motifs (P1 and P2) in the structure of PTPN22 throught its SH3 domain and P1 motif is more important in this interaction. Deletion of P1 motif greatly diminish the inhibitory effect of PTPN22 on TCR signaling. Interaction of those enzymes is needed for their optimal function and the inhibition of TCR signaling. It was also proposed that the interaction of PTPN22 and CSK regulate localization of PTPN22 and dissociation of this complex enable translocation of PTPN22 to lipid rafts of plasma membrane, where it can inhibit TCR signaling. mutant PTPN22, which is unable to bind CSK, is effectively recruited to the plasma membrane.

Another interaction partner of PTPN22 is TRAF3. This protein bind PTPN22 and regulate its translocalization to plasma membrane, in the absence of TRAF3 there is bigger amount of PTPN22 localized at plasma membrane.

Regulation of PTPN22
It was revealed that PTPN22 is regulated by phosphorylation. PTPN22 is phosphorylated on serine in the position 751 by protein PKC (most probably isoform PKCα) after an activation of T cell. This phosphorylation negatively regulate TCR-suppressing function of PTPN22. It also suppress polyubiquitination of PTPN22, which target this protein for degradation, and by this mean It prolong half-life of PTPN22. Phosphorylared PTPN22 interacts better with CSK which hold PTPN22 away from plasma membrane, where it can dephosphorylate proteins of TCR signaling pathway. PTPN22 with mutated serine 751 has shorter half-life, enhanced recruitment to plasma membrane and reduced interaction with CSK.

PTPN22-deficient mice
Young PTPN22-deficient mice do not display any abnormality in peripheral lymphoid organs, but older PTPN22-deficient mice (older than 6 months) develop splenomegaly and lymphadenopathy. In these older mice we can see increased number of T cell with phenotype of effector/memory T cells (CD44hi, CD62Llo), which have higher expression of PTPN22 than naive T cells in Wild Type mice. Expansion of those T cells is supported by PTPN22 deficiency. Compartment of Treg cell is also influenced by PTPN22 deficiency in vivo. Same as with effector/memory T cells, PTPN22-deficient mice contain bigger amount of Treg cells in lymph nodes and spleen and this difference is more visible with increasing age of mice. There is also a change of the effector Treg cells : central Treg cells ratio in favor of effector Treg cells. Influence of PTPN22 deficiency on Treg cells number is consistent with higher expression of PTPN22 in Treg cells than in naive T cells. Another abnormality of PTPN22-deficient mice is spontaneous formation of large germinal centers in spleen and peyer's patches. This formation of germinal centers is dependent on the costimulation molecule CD40L and it is another consequence of T cell dysregulation. PTPN22-deficient mice have increased levels of antibodies. However, there is no increase in levels of autoantibodies. Despite of those effects of PTPN22 deficiency on T cell compartment and antibody production, PTPN22-deficient mice do not show signs of any autoimmune disease.

Disease associated variant of PTPN22
In 2004 Bottini et al. discovered the single nucleotide polymorfism in the PTPN22 gene at nucleotide 1858. In this variant of gene normally occurring cytosin is substitued by thymin (C1858T). This cytosin encodes codon for amino acid arginine in the position 620 of linear protein structure, but mutation to thymin cause change of arginine to tryptophan (R620W). Amino acid 620 is placed in the P1 motif, which is involved in the association with CSK and mutation to tryptophan diminish the ability of PTPN22 to bind CSK. The article reporting existence of this variant also discovered that it is more frequent in Diabetes mellitus type 1 patients. The association of C1858T allele with type 1 diabetes was then confirmed by other studies. In addition, C1858T allele of PTPN22 is associated with other autoimmune disease including Rheumatoid arthritis, systemic lupus erythematosus, juvenile idiopathic arthritis , anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis, Graves’ disease, myasthenia gravis, Addison's disease. Contribution of C1858T PTPN22 allele to those was confirmed by more robust meta-analysis. On the other hand this allele is not linked to autoimmune diseases like multiple sclerosis, Ulcerative colitis, pephigus vulgaris and others. The exact way how the function of PTPN22 is influenced by this mutation is still unknown. Throughout past years there were appearing evidences supporting that C1858T mutation is loss-of-function mutation, but also evidences supporting that It is gain-of-function mutation.