SH2B Adapter Protein 3 (SH2B3)
SH2B3 belongs to the Src homology 2B (SH2B) adapter protein family described in the late 1990s. The SH2B family contains three members (SH2B1, SH2B2, and SH2B3) in mammals. SH2B1, SH2B2, and SH2B3 were originally named as SH2-B [also known as SH2 domain-containing signaling mediator (PSM)], adapter protein with PH and SH2 domains (APS) and Lnk, respectively. The SH2B family is evolutionarily conserved from insects through humans. Unlike mammals, insects have only one SH2B gene (also called Lnk or dSH2B). SH2B3 cDNA was initially cloned and characterized in rat and then in mice in 1997 by Takaki and colleagues who described the first transgenic mice for Lnk/SH2B3 and established SH2B3 as a regulator of signaling in T and B cells (Takaki et al. 1997). Subsequent studies revealed a more widespread role for SH2B3 in myeloid development and hematopoiesis. The deduced amino acid sequences of mouse and rat SH2B3 share 96% identity and 98% similarity overall, with complete identity within the SH2 domain. The human SH2B3 cDNA (GenBank accession no. NM_005475.2) was released in 2000 and shows high homology to both rat and mouse SH2B3 with 73% amino acid sequence identity. Human SH2B3 gene encodes a 575-amino acid protein with a predicted molecular mass of 63 kDa. Initially reported to be preferentially and most strongly expressed in hematopoietic stem cells (HSCs), SH2B3 displays a wide expression in all hematopoietic cells. Importantly and consistent with common cell precursors during embryogenesis, SH2B3 is also expressed in endothelial progenitor cells (EPCs) as well as in mature differentiated endothelial cells (ECs). The description of SH2B3 in activated human vascular ECs was concomitant to the description of SH2B3 in hematopoietic cells (Boulday et al. 2002).
The generation of SH2B3 −/− knock-out mice in 2002 provided the first in-depth description of the regulatory inhibitory functions of SH2B3 in lymphopoiesis, erythropoiesis, and megakaryocytopoiesis (Velazquez et al. 2002). Although it was originally reported that SH2B3 functions as a negative mediator of the T cell receptor signaling, development and activation of T cells are normal in SH2B3-deficient mice. In contrast, these mice exhibit a significant increase in the number of pre-B cells in the spleen and pre- and pro-B cells in the bone marrow, indicating an important role for SH2B3 in regulating B cell development. Subsequently, Takaki et al. demonstrated that SH2B3-deficient mice display a significant increase in hematopoietic progenitor cells (HSC) in the adult bone marrow (Takaki et al. 2003).
SH2B3 has no enzymatic activity; its function is totally dependent on its binding partners. SH2B3 acts as an adapter protein modulating the direction, amplitude, and duration of signal mediated by its binding partner. Functionally, data from several groups established SH2B3 as a negative regulator of signaling mediated by cytokines and growth factors and highlighted the importance of SH2B3 in the control of JAK2/STAT3 signaling and its key role in B cell differentiation and maturation. Typically, wild-type or activated mutant tyrosine kinases phosphorylate SH2B3 at a tyrosine residue which becomes the scaffold for other SH2 domain-containing proteins to attenuate the activated tyrosine kinase. This represents a negative regulatory feedback mechanism. Consequently, impairment of SH2B3 has been reported in relation with myelolymphoproliferative disorders. SH2B3 also plays a role in cytoskeleton, focal adhesion, cell adhesion, and migration via regulator function on integrin signaling in ECs and platelets. During the past decade, and supportive of a major role of SH2B3 in a broader set of pathologies, numerous genetic linkage analysis and genome-wide association studies (GWAS) of single nucleotide polymorphisms (SNP) revealed SH2B3 as a critical genetic determinant for myeloproliferative neoplasms (MPNs), acute lymphoblastic leukemia but also for cardiovascular diseases, blood pressure, type 1 diabetes, autoimmune diseases, lifespan and thus support a role for SH2B3 gene polymorphism and genetic variants. Functional genomic studies to decipher the functional impact of SH2B3 mutations in human pathologies are currently emerging.
SH2B in Drosophila
In Drosophila, Lnk gene encodes a single SH2B protein (dSH2B or Lnk) which acts as a positive regulator of insulin-like signaling. Disruption of the dSH2B gene results in reduced body size and weight, growth retardation, reduced female fertility, increased longevity, increased resistance to starvation and oxidative stress, increased lipid and triglyceride content in fat bodies, and increased hemolymph carbohydrate level (Slack et al. 2010). These changes are accompanied by an impaired PI3K/AKT and insulin signaling pathway as established by a decrease in membrane-associated phosphatidylinositol 3-phosphate (PtdIns3P) and AKT activation in tissues. Consistent with this phenotype, silencing of dSH2B in insect cells inhibits insulin-induced tyrosine phosphorylation of Chico (the Drosophila homolog of Insulin Receptor Substrates, IRS), AKT and ERK activation, as well as Drosophila FoxO phosphorylation and nucleo-cytoplasmic translocation. Drosophila SH2B acts as a direct binding partner of both InR (insulin receptor) and Chico in Drosophila tissues. SH2B acts upstream of Chico. Localization of Chico at the plasma membrane is ensured by both its PH domain and the interaction with dSH2B (Almudi et al. 2013). Furthermore, dSH2B is able to recruit an intracellular InR fragment to the membrane. Interestingly, the Chico-dSH2B and Chico-InR interactions are insulin dependent, whereas dSH2B and InR are able to interact in the absence of insulin stimulation. Systemic overexpression of dSH2B in Drosophila results in a phenotype opposite to that of dSH2B depletion. Similar to dSH2B-deficient flies, SH2B1 null mice accumulate abnormally high levels of lipids in their fat bodies. Of interest the loss of dSH2B increases resistance to oxidative stress as well as lifespan in flies, suggesting that dSH2B may regulate aging and longevity. However, SH2B1-null mice have a shorter lifespan compared with wild-type littermates. These data may reflect the fact that in mammals the functions of adapters segregate among the SH2B proteins. The reduced lifespan may alternatively results from the higher incidence of obesity and obesity-associated diseases that may contribute to early death of SH2B1-deficient mice. However, the role of mammalian SH2B family members in aging still remains unclear.
Structure of the SH2B3 Adapter Protein
The SH2B N-terminal dimerization domain is a conserved domain of approximately 60 amino acids, referred to as a phenylalanine zipper, mediates the homo- and heterodimerization of SH2B1/B2 adapters. It consists of a compact U-shaped four-helix bundle with a stack of interdigitated phenylalanine side chains. Multimerization of SH2B3 proteins has also been reported and occurs through homophilic interaction of the N-terminal domains. Interestingly, noninhibitory SH2B3 mutants with combined deletions in the PH domain and the C-terminal tail act as dominant negative mutants by forming an inactive multimer complex with wild-type protein (Takizawa et al. 2006).
The SH2B3 PH domain is involved in driving protein localization into the cell. PH domain is able to bind phosphoinositide phosphates (PIPs) with high affinity and specificity and are distinguished from other PIP-binding domains by their specific high-affinity binding to PIPs with two vicinal phosphate groups: PtdIns(3,4)P2, PtdIns(4,5)P2, or PtdIns(3,4,5)P3. This interaction results in targeting SH2B3 PH domain proteins to the plasma membrane. Several SH2B3 genetic variants have been reported in GWAS; most of them were found within exon 3 encoding the PH domain and could results in mislocalization and altered function (Gery et al. 2007). The most common SH2B3 nonsynonymous SNP (rs3184504) corresponds to position 262 in the PH domain.
The SH2 domain interacts with phosphotyrosine-containing peptide sequences and leads to protein/protein interactions, and this domain is present in many signal transducing molecules. The consensus of the SH2B3 SH2 domain binding motif is xxpY(V/I/E)xL (Cheng et al. 2016). Of note, the SH2B3 SH2 has some preference for a Pro at pTyr-2 and an Asp at pTyr-1 positions. The preference for an Asp at pTyr-1 is reinforced by the almost total absence of Arg or Lys at the same position. The JAK2 (Tyr813) and JAK3 motifs (Tyr785) are PDpYELL and SDpYELL, respectively. SH2B3 interacts with phosphorylated JAK3 but not kinase-inactive mutant Y785F and, importantly, SH2B3 itself was tyrosine phosphorylated by JAK3 (Cheng et al. 2016). JAK1 lacks the SH2B3 consensus motif and is not able to phosphorylate SH2B3. In mice, the tyrosine 526 residue is subjected to phosphorylation upon SCF stimulation (Takaki et al. 2002).
SH2B3 Is an Inhibitory Regulator of JAK-STAT Signaling
SH2B3 forms a negative feedback loop by binding to MPL and JAK2 and inhibits downstream STAT activation. Although SH2B3 binds to MPL constitutively, it displays only weak binding to JAK2 in basal condition (Bersenev et al. 2008). Upon stimulation with TPO, however, JAK2 becomes phosphorylated and binds to SH2B3 with a higher affinity, thus initiating a critical negative feedback loop. A kinase-inactive mutant of JAK2 results in decreased binding to SH2B3, suggesting that JAK2 phosphorylates SH2B3, which may lead to its enhanced binding. SH2B3 binds to MPL and colocalizes with MPL at the plasma membrane (Gery et al. 2007). The SH2 domain of SH2B3 is essential for this interaction, as well as for the inhibition of downstream signaling. In addition, the PH domain is required for membrane localization. Murine models indicated that loss of SH2B3 function can promote the development of a MPN phenotype (Gery et al. 2007; Oh 2011).
The activating JAK2 V617F mutation is the most frequent mutation associated with MPNs, while other JAK2 mutations (e.g., in exon 12) or activating mutations of the TPO receptor MPL, such as MPL W515 L, occur with less prevalence. Nevertheless, aberrant activation of JAK-STAT signaling in MPN patients without JAK2 or MPL mutations revealed the role of inhibitory regulators of JAK-STAT signaling in MPN pathogenesis. SH2B3 gene mutations have been described in patients with MPN, acute lymphoblastic leukemia (ALL), and acute T-cell leukemia, supporting a role for SH2B3 deficiency in leukemia of both myeloid and lymphoid origin (Oh et al. 2010). These findings provide a novel genetic paradigm of loss of negative feedback regulation of JAK-STAT activation in MPNs and have implications for the future development of targeted therapies in MPNs.
SH2B3 Is a Negative Regulator of IL-7 and IL-11 Signaling
SH2B3 plays a direct role in B cell progenitors. It controls pro-B/pre-B homeostasis and aging by regulating IL-7–mediated JAK/STAT signaling. IL-7 is the primary cytokine regulating pro-B cell expansion. SH2B3 deletion mutations associated with activating mutations in IL7R (which signals via JAK1 and JAK3) were recently reported in high-risk acute lymphoblastic leukemia (ALL) (Roberts et al. 2014). SH2B3 is a negative regulator of IL-7R-mediated JAK/STAT signaling in B progenitors through a direct interaction between SH2B3 and JAK3. The concomitant loss of SH2B3 and p53 has a synergistic effect and results in enhanced IL-7/JAK/STAT signaling and B progenitor cell transformation (Cheng et al. 2016).
SH2B3 is a negative regulator of IL-11 signaling and suppresses radiation resistance and radiation-induced B cell malignancies. IL-11 is critical for the ability of SH2B3 −/− HSPCs to recover from irradiation and become leukemic after a long latency. In IL-11 signaling, SH2B3 suppresses tyrosine phosphorylation of the SH2 domain-containing phosphatase-2/protein tyrosine phosphatase nonreceptor type 11 (Shp2/PTPN11) and its association with the growth factor receptor-bound protein 2 (Grb2), as well as the subsequent activation of the ERK MAP kinase pathway (Louria-Hayon et al. 2013). Indeed, Shp2 has a binding motif for the SH2B3 SH2 domain that is phosphorylated in response to IL-11 stimulation. SH2B3 usually inhibits Shp2 signaling downstream of the IL-11–gp130 complex, and enhanced Shp2/Grb2–pERK signaling causes the development of radiation-related malignancies in SH2B3 −/− mice. The effects of mutations in human SH2B3 on responses to radiotherapy remain to be explored. The finding that SH2B3 −/− mice do not usually develop acute malignancy despite their hyperproliferative disorder suggests that additional genetic mutations, which might be caused by irradiation, could be required for disease progression.
SH2B3 in Solid Tumors Concurs to Tumor Progression
Only few studies have examined activities of SH2B3 in solid tumors. The levels of SH2B3 expression were found elevated in high-grade ovarian cancer and in several other tumors (bladder and kidney cancer, melanoma, sarcoma). Overexpression of SH2B3 in several ovarian cancer cell lines rendered the cells resistant to death induced by either serum or nutriment starvation and generated larger tumors in a murine xenograft model. Resistance to cell death in cancer cells overexpressing SH2B3 associated with enhanced and sustained activation of PI3K and MAP kinases and proliferation. In contrast, silencing of SH2B3 reduces PI3K and MAPK activity and decreases ovarian cancer cell growth in vitro and in vivo (Ding et al. 2015). These results suggest that in solid tumors and in contrast with hematologic malignancies, overexpression of SH2B3 could be a positive contributor of tumor progression. In this study, 14-3-3 proteins, a family of phosphoserine-binding proteins, were identified as one of the SH2B3-binding partners in ovarian cancer cells.
In contrast to MPN that associate with JAK/MPL mutations, ovarian cancers mostly contain mutations associated with alterations in the retinoblastoma, PI3K/RAS, Notch, BRCA1/2 signaling pathways, and the FOXM1 transcription factor. The phosphorylation levels of AKT (upstream of mTOR) and p70S6 (downstream of mTOR) were both found increased upon SH2B3 overexpression, suggesting that the mTOR pathway is upregulated by SH2B3 in ovarian cancer cells. SH2B3 also affects cell adhesion and ECM interaction and inhibits cell migration in ovarian cancer cells possibly by affecting the integrin pathway. These data further support a role for SH2B3 in various cell type beside hematopoietic cells, such as epithelial cells, and also highlight possible cell type and pathway-specific effects.
SH2B3 Is a Negative Regulator of Inflammation and Motility in Endothelial Cells
SH2B3 adapter is basally expressed in resting human vascular ECs (Boulday et al. 2002). Nobuhisa et al. reported that in the mouse embryo, SH2B3 was expressed in the AGM (aorta–gonad–mesonephros) region at E11.5 and was present in the endothelial cells lining the dorsal aorta (Nobuhisa et al. 2003). As expression pattern of SH2B3 overlaps with that of CD34 in the dorsal aorta at a stage of embryonic hematopoiesis, this suggests that SH2B3 might be involved in hematopoietic cell development from endothelial precursors. Moreover, these findings indicated that SH2B3 expression is not only restricted to hematopoietic cells but also extend to nonhematopoietic cells including vascular ECs.
In EC cultures, SH2B3 is rapidly phosphorylated and subsequently upregulated at mRNA and protein level by the proinflammatory cytokine TNF supporting a role for SH2B3 in the TNF signaling. Fitau and colleagues demonstrated that in vitro overexpression of SH2B3 in ECs significantly prevents the induction of VCAM-1 and E-selectin expression in response to TNF signaling (Fitau et al. 2006). Functionally, the reduced adhesion molecule associates with a decreased ability of monocytes to bind activated endothelial monolayer that highlights a new function for SH2B3 as a negative regulator of inflammation and leukocyte trafficking via the endothelium. Mechanistically, overexpression of SH2B3 was not found associated with changes in NFκB/p65 phosphorylation and translocation, nor IκBα phosphorylation and degradation mediated by TNF, suggesting that SH2B3 does not modulate NFκB activity. However, SH2B3 activates the PI3K through AKT phosphorylation. Endothelial NOS (eNOS) was identified as a downstream target of SH2B3-mediated activation of the PI3K/AKT pathway and HO-1 as a new substrate of AKT. SH2B3-mediated activation of PI3K in TNF-activated ECs correlates with the inhibition of ERK1/2 phosphorylation, while phosphorylation of p38 and JNK MAPKs was unchanged. Together these data indicate that SH2B3 negatively regulates TNF signaling and vascular inflammation via activation of AKT and ERK inhibition.
Consistent with a role for SH2B3 in integrin-mediated signaling and cytoskeleton organization reported in platelets, SH2B3 is also a key signaling mediator of cell adhesion and migration in ECs through regulation of β1 integrin signaling. Devallière and colleagues showed that an interaction between SH2B3 and focal adhesion (FA) complexes occurs in adhering, quiescent, and migrating ECs which is temporally regulated during the early stage of the adhesive process (Devalliere et al. 2012). SH2B3 controls FA turnover and sustained SH2B3 expression blocks FA disassembly resulting in a gain in cell attachment that impedes cell migration. SH2B3 takes part in integrin signaling through its direct interaction with (integrin-linked kinase) ILK and is required for efficient phosphorylation of AKT and GSK3β during integrin activation. SH2B3 upregulation was found associated with a functional decrease in α-parvin expression. SH2B3 promotes the formation of ILK-SH2B3 complex and concomitantly inhibits the formation of ILK-α-parvin complex (Devalliere et al. 2012). This correlates with an accumulation of FA and delayed cell migration. Interestingly, in platelets SH2B3 interacts with Fyn. Fyn is functionally distinct from its Src family members in that it interacts with FAK and paxillin in the regulation of cell morphology and motility (Takizawa et al. 2010).
SH2B3 Controls Megakaryocytes Growth and Maturation and Stabilizes Developing Thrombi
SH2B3 is expressed by megakaryocytes, where it plays different regulatory, integrin-dependent and independent roles to control their growth and maturation. Inhibition of TPO-mediated signaling results in elevated megakaryocyte and circulating platelet counts in SH2B3 −/− mice (Velazquez et al. 2002). The absence of SH2B3 causes enhanced and prolonged TPO induction of STAT3, STAT5, AKT, and MAPK signaling pathways in CD41+ megakaryocytes. The SH2 domain of SH2B3 is essential for this inhibitory function. In contrast, the conserved tyrosine near the C-terminus is dispensable and the PH domain of SH2B3 contributes to, but is not essential for, inhibiting TPO-dependent 32D cell growth or megakaryocyte development. In addition, in megakaryocytes, SH2B3 also acts through integrin signaling to regulate their growth and maturation. SH2B3 contribute to αIIbβ3- and actin-dependent morphological responses induced by platelet adhesion to fibrinogen (Takizawa et al. 2010). This effect on cytoskeleton occurs independently of its negative impact on proliferation. SH2B3 is required for recruitment of Fyn (a membrane-associated tyrosine kinase of the Src-family) to its binding site, residues 721–725 (IHDRK) in the β3-cytoplasmic domain, and the subsequent efficient tyrosine phosphorylation of Tyr747 (P-Tyr747) of the β3 integrin subunit, leading to actin cytoskeletal reorganization. Thus, SH2B3 contributes to the stabilization of developing thrombi via Tyr747 phosphorylation of β3 integrin. SH2B3 deficiency in platelets impairs stabilization of the developing thrombus under flow conditions, which may lead to an increase in re-bleeding events in SH2B3 −/−. mice.
SH2B3 Loss of Function Promotes Thrombosis and Atherosclerosis
In studies using human cord blood, the common risk allele (rs3184504*T) of SH2B3 was found associated with expansion of HSC and enhanced megakaryocytopoiesis, demonstrating reduced SH2B3 function and increased MPL signaling (Wang et al. 2016). In mice, hematopoietic SH2B3 deficiency leads to accelerated arterial thrombosis and atherosclerosis, but only in the setting of hypercholesterolemia. Hypercholesterolemia acts synergistically with SH2B3 deficiency to increase IL-3/granulocyte-macrophage colony-stimulating factor (GMCF) receptor signaling in bone marrow myeloid progenitors, whereas in platelets cholesterol loading combines with SH2B3 deficiency to increase activation. Platelet SH2B3 deficiency increases MPL signaling and AKT activation, whereas cholesterol loading decreases SHIP-1 phosphorylation, acting in concert to increase AKT and platelet activation. Together with increased myelopoiesis, platelet activation promotes prothrombotic and proatherogenic platelet/leukocyte aggregate formation.
Hematopoietic and Endothelial Stem Cells Self-Renewal and Commitment
In SH2B3−/− mice, long-term marrow repopulating activity is markedly elevated because of increases in both absolute number and self-renewal activity of HSCs. These results suggest that SH2B3 negatively regulates the key signaling pathways of HSC self-renewal. SH2B3 negatively regulates HSC expansion and self-renewal in part by restricting the TPO/MPL/JAK2 signaling pathway. SH2B3 also interacts via its SH2 domain with Kit and downregulates SCF signaling in BM-derived cells (Velazquez et al. 2002). Overexpression experiments with a series of SH2B3 mutants in AGM cultures showed that the SH2 domain of SH2B3 was a prerequisite for inhibition of the generation of CD45+ nonadherent cells (Nobuhisa et al. 2003). Stem cell factor-induced tyrosine phosphorylation of c-Kit receptors generates the binding site for signal-transducing proteins, such as Grb2, the p85 subunits of PI3K, phospholipase C-γ1, and Src kinase, and consequently leads to proliferation, survival, calcium mobilization, cell migration, and differentiation. SH2B3 suppressed SCF-induced ERK activation. These observations raise the possibility that SH2B3 blocks MAP kinase and PI3-kinase pathways by associating with c-Kit receptors. Importantly, SH2B3, but not APS or SH2-B, regulates AGM hematopoiesis.
The role of SH2B3 in ischemic vasculogenesis was also explored by examining SH2B3 mRNA levels in various populations of bone marrow cells and several organs of WT mice in the presence or absence of limb ischemia. Kwon et al. provided in vitro and in vivo evidence that SH2B3 plays a pivotal role in specific modulation of endothelial progenitor cells (EPCs) in terms of cell growth, commitment into endothelial lineage cell types, mobilization from BM into peripheral blood, and recruitment to ischemic sites for neovascularization (Kwon et al. 2009). From a therapeutic point of view, it has been proposed that SH2B3 may provide a suitable molecular target for enhancement of self-renewal capacity of HSCs in patients. Alternatively, SH2B3 inhibitors could be used for ex vivo expansion of HSCs or EPCs. Takizawa et al. have reported that transient inhibition of endogenous SH2B3 activity by introduction of a dominant-negative form of SH2B3 can increase engraftment rates of HSCs (Takizawa et al. 2006).
SH2B3 Mutations Associate with a Wide Sprectrum of Disease Susceptibilities
The SH2B3 gene maps on chromosome 12 at the band 12q24 and contains eight exons. The chromosome 12q24.12 region contains one of the largest blocks of linkage disequilibrium in the human genome across a 987 kb for a set of genes including SH2B3, ATXN2, BRAP, TRAFD1, ACAD10, ALDH2, and MAPKAPK5. The most common genetic variant (rs3184504) at SH2B3 gene is located in exon 3 and displays a minor allele frequency of approximately 0.147 (1000Genome study). This SNP, which causes a missense mutation at position 262 (protein R262W, codon 784T>C) within the PH domain, was associated in GWAS with blood pressure, coronary heart disease, hypothyroidism, rheumatoid arthritis, and type 1 diabetes. The risk allele rs3184504*T associates with lower SH2B3 expression and function (Wang et al. 2016).
Myeloproliferative Neoplasms – Mutations in SH2B3 were described in approximately 6% of patients with chronic phase MPN (Oh et al. 2010) and up to 10% of blast phase MPN patients (Pardanani et al. 2010). Both somatic and germline mutations in SH2B3 have been reported in patients with MPNs (Oh 2011). Additive mutations in SH2B3 have been reported in patients (McMullin and Cario 2016; Oh et al. 2010). In exon 2, a 5-bp deletion and missense mutation (NM_005475.2: codon [603_607delGCGCT; 613C>G]) leading to a premature stop codon invalidates both the PH and SH2 domains. Missense mutations (NM_005475.2: codon 622G>C) leading to a glutamic acid to glutamine substitution (E208Q) and A215V in the PH domain were also identified. In most cases, SH2B3 PH domain mutations identified in MPN patients are heterozygous, suggesting that SH2B3 haploinsufficiency is sufficient to contribute to MPN pathogenesis. It has been proposed that these mutations, which retain the SH2B3 N-terminal dimerization domain, could act by binding and sequestering wild-type SH2B3, resulting in a dominant negative effect. Interestingly, although activating mutations in the JAK2/STAT pathway (e.g., JAK2, MPL mutations) are considered mutually exclusive, SH2B3 mutations can occur concurrently with JAK2 V617F (Pardanani et al. 2010), suggesting that, functionally, these mutations may cooperate. Interestingly, SH2B3 can bind and inhibit JAK2 V617F as well as MPL W515L (Bersenev et al. 2008) (Gery et al. 2009), suggesting that even in the presence of these activating mutations, additional mechanisms may need to be recruited in order to evade inhibition by SH2B3.
Hypertension and renal disease – SH2B3 gene has been linked to human hypertension and renal disease by GWAS via SNP rs3184504 in the coding region of the PH domain (Levy et al. 2009). Recently, the SH2 domain was functionally deleted in a rat model of hypertension in order to explore further the role of SH2B3. This study showed that abrogation of the phosphotyrosine binding ability of its SH2 domain augments lymphocyte development and ultimately attenuates hypertension. Infiltration of leukocytes into the kidneys, a key mediator of hypertension in this experimental model, was significantly blunted in mutant rats (Dale and Madhur 2016). Similarly in mice, SH2B3 deficiency in hematopoietic cells is primarily responsible for the aggravated hypertensive response (Saleh et al. 2015). These data support a role for both the immune system and the inflammatory mechanisms as modulators of disease severity in the pathogenesis of hypertension and provide insight into SH2B3 at play in human hypertension and renal disease.
Celiac disease – In several GWAS datasets, rs3184504 at SH2B3 was reported to belong to the strongest signatures of positive selection for celiac disease together with rs17810546*T from the IL12A locus and rs917997 from the IL18RAP locus in the European populations (Zhernakova et al. 2010). Functional investigation of the SH2B3 genotype in response to lipopolysaccharide and muramyl dipeptide showed that carriers of the SH2B3 rs3184504*T risk allele showed stronger activation of the NOD2 recognition pathway, suggesting that SH2B3 plays a role in protection against bacterial infection. SH2B3 is strongly expressed in the small intestine, and higher expression in inflamed celiac biopsies was observed that may reflect leukocyte recruitment and activation (Hunt et al. 2008).
Blood pressure – The rs3184504*T allele was found associated with an increment in blood pressure. A large expression quantitative trait locus (eQTL) meta-analysis to identify the downstream effects of disease-associated SNPs was performed in peripheral blood samples. In this study, rs3184504*T decreases expression levels of nine genes, most of which are involved in toll-like receptor signaling (C12orf75, FOS, IDS, IL8, LOC338758, NALP12, PPP1R15A, S100A10, and TAGAP) and increases expression of five genes involved in interferon-γ response (GBP2, GBP4, STAT1, UBE2L6, and UPP1) (Westra et al. 2013). The SNP rs3184504 is a trans-eQTL for 6 of our 34 blood pressure signature genes from the meta-analysis (FOS, MYADM, PP1R15A, TAGAP, S100A10, and FGBP2). These six genes are highly expressed in neutrophils and are coexpressed.
Longevity – Recently, lead SNPs for exceptional longevity have been identified in eight loci and SH2B3 has been reported as an attractive candidate gene, among few others, for extreme longevity (Fortney et al. 2015). In this GWAS analysis, the centenarian allele of the lead SNP (rs3184504*C) is protective for lung and pancreatic cancer, coronary artery disease, rheumatoid arthritis, diastolic blood pressure, and bone mineral density. The protective allele (*C, R262) in human SH2B3 is associated with lower expression of SH2B3 in peripheral blood, consistent with the loss-of-function results for lifespan extension in Drosophila dSH2B, considering the association of the SH2B3/ATXN2 locus with diastolic blood pressure and rheumatoid arthritis. It has been proposed that a panel of SNPs, including the rs3184504, are not specific to any one disease, but instead are associated with a general mechanism such as reduced rate of aging that acts across multiple diseases.
SH2B3 belongs with SH2B1 and SH2B2 proteins to a family of adapter molecules that share structural similarities including a dimerization and proline-rich domain in N-terminus, a central pleckstrin homology, and SH2 domains at C-terminus. SH2B3 expression is found in almost all hematopoietic cells but also extent to endothelial progenitor and mature cells and to some epithelial cancer cells. SH2B3 has no enzymatic activity but mediates protein-protein interaction via its SH2 and PH domains. In most cases, SH2B3 plays regulatory roles in several signaling pathways by providing a negative feedback. SH2B3 canonical inhibitory function is mediated by its SH2 domain. SH2B3 is involved in the negative control of signals induced by IL-3, IL-7, IL-11, SCF, TPO, MPL, EPO, and TNF and its action is exemplified by the JAK/STAT inhibition in hematopoietic cells. SH2B3 is also a negative regulator of integrin outside-in signal in endothelial cells and platelets and controls cytoskeleton organization, cell adhesion, and migration. Deficiency in SH2B3 associates with uncontrolled expansion and self-renewal of stem or progenitor cell, B cell proliferation, endothelial commitment and promotes thrombosis and atherosclerosis. A major genetic variant has been identified at SH2B3 gene (rs3184504) and causes a missense mutation within the PH domain (R262W). In genome-wide association studies, the risk allele rs3184504*T associates with myeloproliferative neoplasms, elevated blood pressure, heart coronary disease, celiac disease, type1 diabetes while the protective allele seems to associate with longevity. Together genetic and functional studies propose SH2B3 as both a genetic marker and a molecular target for drug development and to manipulate hematopoietic cells for immune or cell therapy purpose.
This work was realized in the context of the IHU-Cesti, LabEx IGO, and LabEx Transplantex projects which received French government financial support managed by the National Research Agency (ANR) via the “Investment Into The Future” programs ANR-10-IBHU-005, ANR-11-LABX-0016-01, and ANR-11-LABX-0070. The IHU-Cesti project is also supported by Nantes Metropole and the Pays de la Loire Region. This study is also supported by the Region Pays de la Loire, the “Fondation Centaure” (RTRS) which supports a French transplantation research network and by The Agence de la Biomédecine. I apologize that all references could not be cited due to space restrictions.
- Hunt KA, Zhernakova A, Turner G, Heap GA, Franke L, Bruinenberg M, Romanos J, Dinesen LC, Ryan AW, Panesar D, Gwilliam R, Takeuchi F, McLaren WM, Holmes GK, Howdle PD, Walters JR, Sanders DS, Playford RJ, Trynka G, Mulder CJ, Mearin ML, Verbeek WH, Trimble V, Stevens FM, O’Morain C, Kennedy NP, Kelleher D, Pennington DJ, Strachan DP, McArdle WL, Mein CA, Wapenaar MC, Deloukas P, McGinnis R, McManus R, Wijmenga C, van Heel DA. Newly identified genetic risk variants for celiac disease related to the immune response. Nat Genet. 2008;40:395–402.PubMedPubMedCentralCrossRefGoogle Scholar
- Kubo-Akashi C, Iseki M, Kwon SM, Takizawa H, Takatsu K, Takaki S. Roles of a conserved family of adaptor proteins, Lnk, SH2-B, and APS, for mast cell development, growth, and functions: APS-deficiency causes augmented degranulation and reduced actin assembly. Biochem Biophys Res Commun. 2004;315:356–62.PubMedCrossRefGoogle Scholar
- Kwon SM, Suzuki T, Kawamoto A, Ii M, Eguchi M, Akimaru H, Wada M, Matsumoto T, Masuda H, Nakagawa Y, Nishimura H, Kawai K, Takaki S, Asahara T. Pivotal role of lnk adaptor protein in endothelial progenitor cell biology for vascular regeneration. Circ Res. 2009;104:969–77.PubMedCrossRefGoogle Scholar
- Levy D, Ehret GB, Rice K, Verwoert GC, Launer LJ, Dehghan A, Glazer NL, Morrison AC, Johnson AD, Aspelund T, Aulchenko Y, Lumley T, Kottgen A, Vasan RS, Rivadeneira F, Eiriksdottir G, Guo X, Arking DE, Mitchell GF, Mattace-Raso FU, Smith AV, Taylor K, Scharpf RB, Hwang SJ, Sijbrands EJ, Bis J, Harris TB, Ganesh SK, O’Donnell CJ, Hofman A, Rotter JI, Coresh J, Benjamin EJ, Uitterlinden AG, Heiss G, Fox CS, Witteman JC, Boerwinkle E, Wang TJ, Gudnason V, Larson MG, Chakravarti A, Psaty BM, van Duijn CM. Genome-wide association study of blood pressure and hypertension. Nat Genet. 2009;41:677–87.PubMedPubMedCentralCrossRefGoogle Scholar
- Louria-Hayon I, Frelin C, Ruston J, Gish G, Jin J, Kofler MM, Lambert JP, Adissu HA, Milyavsky M, Herrington R, Minden MD, Dick JE, Gingras AC, Iscove NN, Pawson T. Lnk adaptor suppresses radiation resistance and radiation-induced B-cell malignancies by inhibiting IL-11 signaling. Proc Natl Acad Sci USA. 2013;110:20599–604.PubMedPubMedCentralCrossRefGoogle Scholar
- Roberts KG, Li Y, Payne-Turner D, Harvey RC, Yang YL, Pei D, McCastlain K, Ding L, Lu C, Song G, Ma J, Becksfort J, Rusch M, Chen SC, Easton J, Cheng J, Boggs K, Santiago-Morales N, Iacobucci I, Fulton RS, Wen J, Valentine M, Cheng C, Paugh SW, Devidas M, Chen IM, Reshmi S, Smith A, Hedlund E, Gupta P, Nagahawatte P, Wu G, Chen X, Yergeau D, Vadodaria B, Mulder H, Winick NJ, Larsen EC, Carroll WL, Heerema NA, Carroll AJ, Grayson G, Tasian SK, Moore AS, Keller F, Frei-Jones M, Whitlock JA, Raetz EA, White DL, Hughes TP, Guidry Auvil JM, Smith MA, Marcucci G, Bloomfield CD, Mrozek K, Kohlschmidt J, Stock W, Kornblau SM, Konopleva M, Paietta E, Pui CH, Jeha S, Relling MV, Evans WE, Gerhard DS, Gastier-Foster JM, Mardis E, Wilson RK, Loh ML, Downing JR, Hunger SP, Willman CL, Zhang J, Mullighan CG. Targetable kinase-activating lesions in Ph-like acute lymphoblastic leukemia. N Engl J Med. 2014;371:1005–15.PubMedPubMedCentralCrossRefGoogle Scholar
- Saleh MA, McMaster WG, Wu J, Norlander AE, Funt SA, Thabet SR, Kirabo A, Xiao L, Chen W, Itani HA, Michell D, Huan T, Zhang Y, Takaki S, Titze J, Levy D, Harrison DG, Madhur MS. Lymphocyte adaptor protein LNK deficiency exacerbates hypertension and end-organ inflammation. J Clin Invest. 2015;125:1189–202.PubMedPubMedCentralCrossRefGoogle Scholar
- Takizawa H, Nishimura S, Takayama N, Oda A, Nishikii H, Morita Y, Kakinuma S, Yamazaki S, Okamura S, Tamura N, Goto S, Sawaguchi A, Manabe I, Takatsu K, Nakauchi H, Takaki S, Eto K. Lnk regulates integrin alphaIIbbeta3 outside-in signaling in mouse platelets, leading to stabilization of thrombus development in vivo. J Clin Invest. 2010;120:179–90.PubMedCrossRefGoogle Scholar
- Westra HJ, Peters MJ, Esko T, Yaghootkar H, Schurmann C, Kettunen J, Christiansen MW, Fairfax BP, Schramm K, Powell JE, Zhernakova A, Zhernakova DV, Veldink JH, Van den Berg LH, Karjalainen J, Withoff S, Uitterlinden AG, Hofman A, Rivadeneira F, Hoen PA’t, Reinmaa E, Fischer K, Nelis M, Milani L, Melzer D, Ferrucci L, Singleton AB, Hernandez DG, Nalls MA, Homuth G, Nauck M, Radke D, Volker U, Perola M, Salomaa V, Brody J, Suchy-Dicey A, Gharib SA, Enquobahrie DA, Lumley T, Montgomery GW, Makino S, Prokisch H, Herder C, Roden M, Grallert H, Meitinger T, Strauch K, Li Y, Jansen RC, Visscher PM, Knight JC, Psaty BM, Ripatti S, Teumer A, Frayling TM, Metspalu A, van Meurs JB, Franke L. Systematic identification of trans eQTLs as putative drivers of known disease associations. Nat Genet. 2013;45:1238–43.PubMedPubMedCentralCrossRefGoogle Scholar
- Zhernakova A, Elbers CC, Ferwerda B, Romanos J, Trynka G, Dubois PC, de Kovel CG, Franke L, Oosting M, Barisani D, Bardella MT, Joosten LA, Saavalainen P, van Heel DA, Catassi C, Netea MG, Wijmenga C. Evolutionary and functional analysis of celiac risk loci reveals SH2B3 as a protective factor against bacterial infection. Am J Hum Genet. 2010;86:970–7.PubMedPubMedCentralCrossRefGoogle Scholar