Selective expansion of marginal zone B cells in Emicro-API2-MALT1 mice is linked to enhanced IkappaB kinase gamma polyubiquitination

The translocation t(11;18)(q21;q21) that generates an API2-MALT1 fusion protein is the most common structural abnormality among the genetic defects reported in mucosa-associated lymphoid tissue (MALT)-type lymphomas, and its presence correlates with the apparent lack of further genetic instability or chromosomal imbalances. Hence, constitutive nuclear factor-kappaB (NF-kappaB) activation induced by the API2-MALT1 fusion protein is considered essential for B-cell transformation. To examine its role in B-cell development and lymphomagenesis, Emu-API2-MALT1 transgenic mice were produced. Our data show that expression of the API2-MALT1 fusion protein alone is not sufficient for the development of lymphoma masses within 50 weeks. Nevertheless, API2-MALT1 expression affected B-cell maturation in the bone marrow and triggered the specific expansion of splenic marginal zone B cells. Polyubiquitination of IkappaB kinase gamma (IKKgamma), indicative for enhanced NF-kappaB activation, was increased in splenic lymphocytes and promoted the survival of B cells ex vivo. In addition, we show that the API2-MALT1 fusion resided in the cholesterol- and sphingolipid-enriched membrane microdomains, termed lipid rafts. We provide evidence that association of the MALT1 COOH terminal with the lipid rafts, which is mediated by the API2 portion, is sufficient to trigger NF-kappaB activation via enhanced polyubiquitination of IKKgamma. Taken together, these data support the hypothesis that the API2-MALT1 fusion protein can contribute to MALT lymphoma formation via increased NF-kappaB activation.     

Molecular pathogenesis of MALT lymphoma: two signaling pathways underlying the antiapoptotic effect of API2-MALT1 fusion protein.

At least three recurrent chromosomal translocations, t(11;18)(q21;q21), t(1;14)(p22;q32), t(14;18)(q32;q21), involving the API2-MALT1 fusion protein, BCL10 and MALT1, have been implicated in the pathogenesis of mucosa-associated lymphoid tissue (MALT) lymphoma. Several lines of evidence indicated that both BCL10 and MALT1 are required for nuclear factor kappa B (NF-kappaB) activation by antigen receptor stimulation in lymphocytes, and API2-MALT1 can bypass this BCL10/MALT1 signaling pathway. Nuclear factor kappa B activation may contribute to antiapoptotic effect through NF-kappaB-mediated upregulation of apoptotic inhibitor genes. We recently demonstrated that API2-MALT1 can induce transactivation of the API2 gene through NF-kappaB activation, thus highlighting a positive feedback-loop mechanism of self-activation by upregulating its own expression in t(11;18) MALT lymphomas. We also demonstrated that API2-MALT1 possesses an antiapoptotic effect, in part, through its direct interaction with apoptotic regulators. These findings therefore led us to hypothesize that the antiapoptotic effect by API2-MALT1 may be mediated by its interaction with apoptotic regulators, on the one hand, and by NF-kappaB-mediated upregulation of apoptotic inhibitor genes on the other. We also found that BCL10 and MALT1 are shuttling between nucleus and cytoplasm, and that MALT1 can regulate the subcellular location of BCL10.     

Stability and subcellular localization of API2-MALT1 chimeric protein involved in t(11;18) (q21;q21) MALT lymphoma

t(11; 18) (q21; q21) is a chromosomal aberration specific to low-grade mucosa-associated lymphoid tissue (MALT) lymphoma, and generates the chimeric product apoptosis inhibitor 2 (API2)-MALT1, which has been suggested to play an important role in MALT lymphomagenesis. However, little is known about the characteristics of API2, MALT1, and API2-MALT1 proteins. We therefore investigated the subcellular localization and stability of these products. API2 was localized in the nucleus and the cytoplasm, and MALT1 and API2-MALT1 in the cytoplasm only. Western blot analysis showed that the products of API2 and MALT1 were unstable, while the API2-MALT1 product was stable. The API2 deletion mutants at the end of the C-terminal and the MALT1 deletion mutants at the end of the N-terminal were stable compared with the full-length products. These results indicate that the domains responsible for protein instability are located in the end of the C-terminal of API2 and in that of the N-terminal of MALT1, and also that API2-MALT1 became stable because it lacks these domains. It has been suggested that NF-kappaB activation plays an important role in the tumorigenesis of MALT lymphoma. Our findings further suggest that the stabilized expression of API2-MALT1 products may continuously stimulate the NF-kappaB activating pathway, thus leading to MALT lymphomagenesis.     

Primary pulmonary MALT lymphomas show frequent and heterogeneous cytogenetic abnormalities, including aneuploidy and translocations involving API2 and MALT1 and IGH and MALT1.

API2-MALT1 fusion and aneuploidy are common chromosomal abnormalities in MALT lymphoma. In studying their incidence and relationship in primary pulmonary MALT lymphomas, a translocation involving MALT1 and IGH was also identified. In all, 28 primary pulmonary MALT lymphomas were studied by fluorescence in situ hybridization using an API2-MALT1 probe and multiple centromeric probes, as well as IGH-BCL2, IGH-MALT1, and MALT1 breakapart probes in selected cases. Seven (25%) had API2-MALT1 fusion; all seven lacked aneuploidy except for two with trisomy 3 in a small clone. Three (11%) had IGH-MALT1 fusion; two also showed trisomy 3 and 12. A total of 11 (39%) had aneuploidy only, with trisomy 3 and 18 being the most common. Ectopic nuclear bcl-10 expression, which has been previously associated with API2-MALT1, was seen by immunohistochemistry in 86% of API2-MALT1 fusion-positive cases, one IGH-MALT1 fusion-positive case, two aneuploidy-only cases, and two normal cases. In primary pulmonary MALT lymphomas, cytogenetic abnormalities are common (75%) and heterogeneous, encompassing API2-MALT1 and IGH-MALT1, which are mutually exclusive, as well as aneuploidy, which may be present in the latter but is rare in the former. Ectopic nuclear bcl-10 expression is associated with API2-MALT1 but may also be seen in IGH-MALT1 fusion-positive, aneuploidy-only, and normal cases.     

黏膜相关淋巴组织结外边缘区B细胞淋巴瘤BCL-10蛋白表达及其与API2-MALT1融合基因的关系

目的 探讨MALT淋巴瘤中BCL-10蛋白核表达与API2-MALT1融合基因的关系。方法 对86例MALT淋巴瘤进行回顾性研究,采用SP法检测BCL-10的表达,RT-PCR检测API2-MALT1融合基因。结果 10例桥本甲状腺炎均表达BCL-10于胞浆,86例MALT淋巴瘤患者中,42例（48．8％）同时表达BCL-10于胞核和胞浆,35例（40．7％）检测出API2-MALT1融合基因。BCL-10核表达的检出与API2-MALT1融合基因具有相关性（r=0．374,P=0．000）。结论 MALT淋巴瘤中BCL-10蛋白的核表达与API2-MALT1融合基因相关。     

A dual role for the API2 moiety in API2-MALT1-dependent NF-kappaB activation: heterotypic oligomerization and TRAF2 recruitment

Mucosa-associated lymphoid tissue (MALT) lymphoma is the most common extranodal lymphoid neoplasm. Chromosomal translocation t(11;18)(q21,q21) is found in 30% of gastric MALT lymphomas and is associated with a failure to respond to standard treatment and a tendency to disseminate. This translocation generates a chimeric protein composed of N-terminal sequences of Inhibitor of Apoptosis 2 (API2, also known as BIRC3 and cIAP2) fused to C-terminal sequences of MALT1. API2-MALT1 promotes cell survival and proliferation via activation of nuclear factor-kappaB (NF-kappaB). Here, we investigate the mechanism by which the API2 moiety contributes to NF-kappaB stimulation. We find that the API2 moiety mediates oligomerization of API2-MALT1 as well as interaction with tumor necrosis factor receptor-associated factor 2 (TRAF2). Surprisingly, oligomerization does not occur via homotypic interaction; rather, the API2 moiety of one monomer interacts with the MALT1 moiety of another monomer. Further, the specific region of the API2 moiety responsible for mediating oligomerization is distinct from that mediating TRAF2 binding. Although deletion or mutation of the TRAF2 binding site does not inhibit oligomerization, it does lead to dramatically decreased NF-kappaB activation. Deletion of both TRAF2 binding and oligomerization regions results in near-complete loss of NF-kappaB activation. Thus, API2 moiety-mediated heterotypic oligomerization and TRAF2 binding both contribute to maximal API2-MALT1-dependent NF-kappaB stimulation.     

Genetic and epigenetic factors involved in B-cell lymphomagenesis

Malignant lymphomas have been classified by the WHO into disease categories based not only on histological features, but also on cell surface markers, cytogenetic and clinical features. It is known that chromosome translocation plays an important role in lymphoma development, but it is not entirely clear yet why a given type of chromosome translocation is associated with a specific type of lymphoma. This review deals with molecular mechanisms of B-cell lymphoma development in association with chromosome translocations. The outcome of chromosome translocations can be categorized into three factors: enhancement of proliferation, inhibition of differentiation and anti-apoptotic activity. It is well known that chromosome translocation by itself cannot cause cells to become malignant because it is only one of the growth advantages leading to malignancy, while additional genetic and epigenetic alterations are required for cells to become fully malignant. Mucosa-associated lymphoid tissue (MALT) lymphomas of the stomach are unique in that a majority can be cured by Helicobacter pylori eradication, although 20 to 30% remain resistant. Others as well as we have demonstrated that the presence of the API2-MALT1 chimeric gene correlates well with resistance to H. pylori eradication treatment. These characteristics have led to the speculation that the classification of MALT lymphoma falls somewhere between tumor and inflammation. Although MALT lymphoma seems to have unique features in comparison with other types of B-cell lymphomas, it shares common molecular mechanisms with B-cell lymphoma development.     

Constitutive NF-kappaB activation by the t(11;18)(q21;q21) product in MALT lymphoma is linked to deregulated ubiquitin ligase activity

Mucosa-associated lymphoid tissue (MALT) lymphoma is a common type of lymphoma in extranodal sites. The most frequent chromosome translocation associated with MALT lymphoma is t(11;18)(q21;q21), which generates a chimeric protein of c-IAP2 and MALT1/paracaspase. The c-IAP2/MALT1 fusion protein activates the NF-kappaB pathway, which is considered critical to malignant B cell transformation and lymphoma progression. The mechanism by which this fusion protein activates NF-kappaB, however, remains unclear. Here we show that self-oligomerization of the c-IAP2/MALT1 protein causes deregulated ubiquitin ligase activity of MALT1/paracaspase. The chimeric protein targets NEMO for polyubiquitination and thereby activates NF-kappaB. Consistent with this finding, NEMO ubiquitination is increased in t(11;18)(q21;q21)-positive MALT lymphoma samples. Thus, t(11;18)(q21;q21) deregulates MALT1/paracaspase ubiquitin ligase activity, causing constitutive NF-kappaB activation and promoting tumorigenesis.     

黏膜相关淋巴组织淋巴瘤病因学及分子遗传学研究进展

 黏膜相关淋巴组织（MALT）淋巴瘤是起源于淋巴结外MALT的低度恶性B细胞性淋巴瘤。除早期发现胃MALT淋巴瘤与幽门螺杆菌（HP）感染有关外,最近又发现一些与其他部位MALT淋巴瘤有关的病原体。 t(11;18)(q21;q21)、t(1;14)(p22;q32)、t(14;18)(q32;q21)是MALT淋巴瘤特征性的染色体异常,在不同部位的MALT淋巴瘤中发生率也不同。这三种染色体易位产物激活共同的分子通路NF-κB通路。另外还发现了一些新的基因异常。这些病因学及分子遗传学的研究有助于了解MALT淋巴瘤的发病机制,也有助于早期诊断和合理治疗。     

Helicobacter pylori-negative / API2-MALT1 translocation-negative low-grade MALT lymphoma

A 71-year-old man with a Helicobacter pylori infection-negative and API2-MALT1 translocation-negative extranodal marginal-zone B-cell lymphoma of mucosa-associated lymphoid tissue (MALT) type of the stomach has been followed conservatively for over 5 years. The lesion has shown no major morphological changes or malignant progression into a diffuse large-cell type during the time course. The absence of genetic translocation of API2-MALT1 was confirmed with fluorescence in situ hybridization (FISH). The prognosis of H. pylori-negative and API2-MALT1 translocation-negative low-grade MALT lymphoma is unknown, and a standard treatment for such lymphoma has yet to be defined. The case of MALT lymphoma negative for both of the above factors that we report has shown no obvious rapid progression or malignant change over the long-term course. Although curative operation and/or chemoradiotherapy should still be discussed as the treatment of choice, the treatment of this type of lymphoma must be carefully determined on a case-by-case basis, according to its biological status and prognosis.     

Molecular pathogenesis of mucosa-associated lymphoid tissue lymphoma.

Extranodal marginal zone B-cell lymphomas of mucosa-associated lymphoid tissue (MALT) type occur in a number of anatomic sites, but share overlapping morphologic and immunophenotypic features. Helicobacter pylori infection has been identified as an etiologic factor in gastric MALT lymphoma, and a growing list of other infectious organisms have recently been shown to be associated with MALT lymphomas at other anatomic sites. Although cause and effect has not been established for most of these infectious agents, our understanding of the biology has significantly improved, in part through the application of standard cytogenetic analyses. The common karyotypic alterations that characterize MALT lymphomas include the trisomies 3 and 18, the translocations t(11;18)(q21;q21), t(1;14)(p22;q32), t(14;18)(q32;q21), t(3;14)(q27;q32), and the recently described t(3;14)(p14.1;q32). This apparent complexity of cytogenetic alterations that have now been implicated in the pathogenesis of extranodal MALT lymphoma serves as a paradigm for molecular cross talk in neoplastic disease. Recent data have shown that at least three of the disparate translocations affect a common signaling mechanism, and thus unify all three under a common pathogenesis, resulting in the constitutive activation of the nuclear factor kappa B (NF-kappaB) pathway. It may be that the new MALT-related translocation involving the FOXP1 gene and other as yet undiscovered translocations may all have in common increased NF-kappaB signaling.