Inhibition of MALT1 Decreases Neuroinflammation and Pathogenicity of Virulent Rabies Virus in Mice.

Rabies virus is a neurovirulent RNA virus, which causes about 59,000 human deaths each year. Treatment for rabies does not exist due to incomplete understanding of the pathogenesis. MALT1 mediates activation of several immune cell types and is involved in the proliferation and survival of cancer cells. MALT1 acts as a scaffold protein for NF-κB signaling and a cysteine protease that cleaves substrates, leading to the expression of immunoregulatory genes. Here, we examined the impact of genetic or pharmacological MALT1 inhibition in mice on disease development after infection with the virulent rabies virus strain CVS-11. Morbidity and mortality were significantly delayed in Malt1-/- compared to Malt1+/+ mice, and this effect was associated with lower viral load, proinflammatory gene expression, and infiltration and activation of immune cells in the brain. Specific deletion of Malt1 in T cells also delayed disease development, while deletion in myeloid cells, neuronal cells, or NK cells had no effect. Disease development was also delayed in mice treated with the MALT1 protease inhibitor mepazine and in knock-in mice expressing a catalytically inactive MALT1 mutant protein, showing an important role of MALT1 proteolytic activity. The described protective effect of MALT1 inhibition against infection with a virulent rabies virus is the precise opposite of the sensitizing effect of MALT1 inhibition that we previously observed in the case of infection with an attenuated rabies virus strain. Together, these data demonstrate that the role of immunoregulatory responses in rabies pathogenicity is dependent on virus virulence and reveal the potential of MALT1 inhibition for therapeutic intervention.IMPORTANCE Rabies virus is a neurotropic RNA virus that causes encephalitis and still poses an enormous challenge to animal and public health. Efforts to establish reliable therapeutic strategies have been unsuccessful and are hampered by gaps in the understanding of virus pathogenicity. MALT1 is an intracellular protease that mediates the activation of several innate and adaptive immune cells in response to multiple receptors, and therapeutic MALT1 targeting is believed to be a valid approach for autoimmunity and MALT1-addicted cancers. Here, we study the impact of MALT1 deficiency on brain inflammation and disease development in response to infection of mice with the highly virulent CVS-11 rabies virus. We demonstrate that pharmacological or genetic MALT1 inhibition decreases neuroinflammation and extends the survival of CVS-11-infected mice, providing new insights in the biology of MALT1 and rabies virus infection.

The API2-MALT1 fusion exploits TNFR pathway-associated RIP1 ubiquitination to promote oncogenic NF-κB signaling.

The API2-MALT1 fusion oncoprotein is created by the recurrent t(11;18)(q21;q21) chromosomal translocation in mucosa-associated lymphoid tissue (MALT) lymphoma. We identified receptor interacting protein-1 (RIP1) as a novel API2-MALT1-associated protein, and demonstrate that RIP1 is required for API2-MALT1 to stimulate canonical nuclear factor kappa B (NF-κB). API2-MALT1 promotes ubiquitination of RIP1 at lysine (K) 377, which is necessary for full NF-κB activation. Furthermore, we found that TNF receptor-associated factor 2 (TRAF2) recruitment is required for API2-MALT1 to induce RIP1 ubiquitination, NF-κB activation and cellular transformation. Although both TRAF2 and RIP1 interact with the API2 moiety of API2-MALT1, this moiety alone is insufficient to induce RIP1 ubiquitination or activate NF-κB, indicating that API2-MALT1-dependent RIP1 ubiquitination represents a gain of function requiring the concerted actions of both the API2 and MALT1 moieties of the fusion. Intriguingly, constitutive RIP1 ubiquitination was recently demonstrated in several solid tumors, and now our study implicates RIP1 ubiquitination as a critical component of API2-MALT1-dependent lymphomagenesis.

DUSP16 is an epigenetically regulated determinant of JNK signalling in Burkitt's lymphoma.

BACKGROUND: The mitogen-activated protein kinase (MAPK) phosphatases or dual specificity phosphatases (DUSPs) are a family of proteins that catalyse the inactivation of MAPK in eukaryotic cells. Little is known of the expression, regulation or function of the DUSPs in human neoplasia. METHODS: We used RT-PCR and quantitative PCR (qPCR) to examine the expression of DUSP16 mRNA. The methylation in the DUSP16 CpG island was analysed using bisulphite sequencing and methylation-specific PCR. The activation of MAPK was determined using western blotting with phospho-specific antibodies for extra-cellular signal-related kinase (ERK), p38 and c-Jun N-terminal kinase (JNK). The proliferation of cell lines was assessed using the CellTiter 96 Aqueous One assay. RESULTS: The expression of DUSP16, which inactivates MAPK, is subject to methylation-dependent transcriptional silencing in Burkitt's Lymphoma (BL) cell lines and in primary BL. The silencing is associated with aberrant methylation in the CpG island in the 5' regulatory sequences of the gene blocking its constitutive expression. In contrast to BL, the CpG island of DUSP16 is unmethylated in other non-Hodgkin's lymphomas (NHLs) and epithelial malignancies. In BL cell lines, neither constitutive nor inducible ERK or p38 activity varied significantly with DUSP16 status. However, activation of JNK was increased in lines with DUSP16 methylation. Furthermore, methylation in the DUSP16 CpG island blocked transcriptional induction of DUSP16, thereby abrogating a normal physiological negative feedback loop that limits JNK activity, and conferred increased cellular sensitivity to agents, such as sorbitol and anthracycline chemotherapeutic agents that activate JNK. CONCLUSION: DUSP16 is a new epigenetically regulated determinant of JNK activation in BL.

The pathogenesis of MALT lymphomas: where do we stand?

Mucosa-associated lymphoid tissue (MALT) lymphoma is a heterogeneous form of a B-cell non-Hodgkin's lymphoma with extranodal location. The gastrointestinal tract is the most common site of disease, but involvement of multiple other organ systems has been documented. Four translocations, t(11;18)(q21;q21), t(1;14)(p22;q32), t(14;18)(q32;q21) and t(3;14)(p13;q32), are specifically associated with MALT lymphoma. Remarkably, the genes targeted by at least three of these translocations are involved in one and the same pathway, leading to the activation of nuclear factor-kappaB (NF-kappaB). This review presents MALT lymphoma as a model of how sustained inflammation increases the risk of genotoxic insults and how these genetic events initiate oncogenesis.

A novel N-ras mutation in malignant melanoma is associated with excellent prognosis.

Mutations in the ras gene are key events in the process of carcinogenesis; in particular, point mutations in codon 61 of exon 2 of the N-ras gene occur frequently in malignant melanoma (MM). We searched for point mutations in the N-ras gene in a large series of primary and metastatic MM from 81 different retrospectively selected patients using the very sensitive denaturing gradient gel electrophoresis technique, followed by sequencing. The classical codon 12 and codon 61 mutations were found in 21 and 17% of the cases, respectively. No codon 13 mutation was found. A novel mutation at codon 18 of exon 1, consisting of a substitution of alanine (GCA) by threonine (ACA), was found in 15% of the primary MMs but in none of the metastatic MMs. All of the other cases were free of mutations. Using microdissected cells from distinctive MM growth phases as source of DNA for mutation analysis, this particular N-ras exon 1 mutation at codon 18 was already present in the radial growth phase and preserved throughout the successive growth phases; it was also found in a dysplastic nevi in continuity with a MM, indicating a clonal relationship between both lesions. Our findings also illustrate the clonal relationship between the distinctive growth phases in MM and suggest the codon 18 mutation to occur early in MM development. The MM in patients with this mutation were significantly thinner than those without a codon 18 mutation (P = 0.0257). Statistical analysis, comparing the group of codon 18 patients with the group of patients with the classical mutations and without mutations, revealed a highly significant difference in overall outcome. The cumulative probability of developing metastasis was significantly lower for the group patients with a codon 18 mutation (P = 0.0130). We can thus conclude that this codon 18 mutation identifies a group of patients with better prognosis than patients with melanoma that harbor wild-type sequence or classical activating point mutations in codon 12 or 61. Preliminary nucleotide binding measurements could not detect a difference between wild-type Ras protein and the mutant Ras(A18T) protein. However, for a precise elucidation of the role of the N-Ras(A18T) mutant in melanoma, additional studies aimed to measure the affinity to guanine nucleotide exchange factors and GTPase-activating proteins are needed.

Structure of the MLT gene and molecular characterization of the genomic breakpoint junctions in the t(11;18)(q21;q21) of marginal zone B-cell lymphomas of MALT type.

The t(11;18)(q21;q21) between the inhibitor of apoptosis API2 and the MLT gene is a distinct feature of marginal zone B-cell lymphomas of MALT-type. Hitherto the chimeric API2-MLT transcripts are all "in-frame" and predominantly fuse exon 7 of API2 to different MLT exons. Recurrent chromosomal translocations are common in lymphoid neoplasms and might represent by-products of the rearrangement processes generating antigen receptor diversity. The genomic structure of the MLT gene was determined to facilitate amplification of the genomic breakpoint junctions from 5 MALT-type lymphomas with t(11;18). Their sequence analysis showed scattering of the chromosome 11 breakpoints in intron 7 of API2 whereas rearrangements in MLT occurred in intron 2, 4, 7, or 8, respectively. Sequences around the junctions did not display recognition signal sequences mediating lymphocytic V(D)J recombination or other sequence motifs associated with recombination. The breakpoints occurred in a copy of an AluSx repeat in three cases, but interchromosomal Alu-mediated homologous recombination could be ruled out as the repeat resided only on one of the participating chromosomes. The t(11;18) was associated with a deletion in 4 out of 5 cases, ranging in size from 53 bp up to more than 200 kb. These deletions were observed on one or sometimes both derivative chromosomes that might indicate the susceptibility of these regions for breakage. Our data suggest that the API2-MLT fusion might result from a non-homologous end joining event after multiple double-strand breaks. The clustering of breaks in intron 7 of API2 and the consistent "in frame" API2-MLT fusions could therefore reflect certain functional constraints crucial for clonal outgrowth.

Detection of t(11;18)(q21;q21) by interphase fluorescence in situ hybridization using API2 and MLT specific probes.

The translocation of chromosome 11, long arm, region 2, band 1, to chromosome 18, long arm, region 2, band 1 (t(11;18)(q21;q21)) represents a recurrent chromosomal abnormality in extranodal marginal zone B-cell lymphoma (MZBCL) of mucosa-associated lymphoid tissue (MALT) type and leads to a fusion of the apoptosis inhibitor-2 (API2) gene on chromosome 11 and the MALT lymphoma-associated translocation (MLT) gene on chromosome 18. A 2-color fluorescence in situ hybridization (FISH) assay, which can be used for the detection of t(11;18) in interphase nuclei and metaphase chromosomes on fresh and archival tumor tissue, was developed. The P1 artificial chromosome (PAC) clone located immediately telomeric to the MLT gene and the PAC clone spanning the API2 gene were differentially labeled and used to visualize the derivative chromosome 11 resulting from t(11;18), as evident by the overlapping or juxtaposed red and green fluorescent signals. The assay was applied to interphase nuclei of 20 cases with nonmalignant conditions and 122 B-cell non-Hodgkin's lymphomas (NHLs). The latter group comprised 20 cases of nodal follicle center cell lymphoma and diffuse large B-cell NHL, 10 cases of gastric diffuse large B-cell lymphoma, 10 cases of hairy cell leukemia, and 82 cases of MZBCL (41 extranodal from various locations, 19 nodal, and 22 splenic MZBCL) including 35 cases with an abnormal karyotype, 2 of which revealed t(11;18). By interphase FISH, t(11;18) was detected in 8 gastrointestinal low-grade MALT-type lymphomas including the 2 cytogenetically t(11;18)(+) cases. In the 8 t(11;18)(+) cases, the FISH results were confirmed by reverse transcriptase-polymerase chain reaction (RT-PCR) using API2 and MLT specific primers. Our results indicate that t(11;18)(q21;q21) specifically characterizes a subgroup of low-grade MZBCL of the MALT-type and that the FISH assay described here is a highly specific and rapid test for the detection of this translocation.

The product of the t(11;18), an API2-MLT fusion, is an almost exclusive finding in marginal zone cell lymphoma of extranodal MALT-type.

BACKGROUND: Extranodal marginal zone cell lymphoma (MZCL) of MALT-type share similar features with nodal and splenic MZCL regarding morphology and immunophenotype. At the genetic level, recent cytogenetic studies have shown that t(11;18) is a recurring abnormality in extranodal MALT-type MZCL but has hitherto never been reported in nodal or splenic MZCL. The aim of the present study was to determine the prevalence of t(11;18) in a large series of nodal, splenic and extranodal MALT-type MZCL, using a sensitive real-time RT-PCR method. MATERIALS AND METHODS: Ninety-three MZCL cases were divided on clinical grounds into 61 extranodal MALT-type, 19 splenic and 12 nodal MZCL. One case that presented with a massive splenomegaly but for which also gastro-intestinal localisations were found, was left unclassified. A real-time RT-PCR method for the detection of the API2-MLT fusion resulting from t(11;18) was performed on RNA extracted from frozen tissue sections. RESULTS: The API2-MLT fusion was detected in 12 cases, which were all extranodal MALT-type lymphomas of the stomach, except for one case. The remaining positive case was the unclassified case, for which the translocation was detected in the spleen and in hilar lymph node tissue. CONCLUSIONS: While similarities between MZCL from different anatomic sites have lend us to propose that all MZCL have a common normal counterpart, the almost exclusive detection of t(11;18) in gastric MALT-type lymphoma favours its recognition as a separate lymphoma entity. The absence of the translocation in nodal and splenic MZCL challenges the idea of these lymphomas being secondary to MALT-type lymphomas of the gut. The unclassified case illustrates the inadequate approaches available at present to identify and define the various MZCL.

The product of the t(11;18), an API2-MLT fusion, marks nearly half of gastric MALT type lymphomas without large cell proliferation.

Recently we demonstrated that the t(11;18)(q21;q21) associated with extranodal marginal zone B cell lymphomas of MALT type results in the expression of a chimeric transcript fusing 5' API2 on chromosome 11 to 3' MLT on chromosome 18. Here we report the development of an RT-PCR approach for the detection of the API2-MLT fusion transcript and its application for the analysis of 58 cases of gastric lymphoma. Initially nested PCR amplification was combined with Southern analysis using internal API2 and MLT probes. A genuine API2-MLT fusion transcript of variable length was demonstrated in 11 out of 58 cases. Sequence analysis revealed that in all cases the breakpoint on chromosome 11 occurred between exons 7 and 8 of the API2 gene. In contrast, the breakpoints on chromosome 18 appeared to be heterogeneous as fusions to bp 814, 1123, and 1150, respectively, of MLT were observed. These observations allowed us to work out a highly sensitive diagnostic test for the API2-MLT fusion on an ABI Prism 7700 sequence detector that confirmed the results of our initial approach. The API2-MLT fusion was found in 48% of gastric marginal zone cell lymphomas of MALT type that did not contain a large cell component and it was lacking in all other lymphomas of the stomach.

Three differentially expressed survivin cDNA variants encode proteins with distinct antiapoptotic functions.

Survivin is a member of the inhibitor of apoptosis protein (IAP) family that is believed to play a role in oncogenesis. To elucidate further its physiologic role(s), we have characterized the murine survivin gene and complementary DNA (cDNA). The structural organization of the survivin gene, located on chromosome 11E2, is similar to that of its human counterpart, both containing 4 exons. Surprisingly, 3 full-length murine survivin cDNA clones were isolated, predicting the existence of 3 distinct survivin proteins. The longest open reading frame, derived from all 4 exons, predicts a 140-amino acid residue protein, survivin(140), similar to human survivin, which contains a single IAP repeat and a COOH-terminal coiled-coil domain that links its function to the cell cycle. A second cDNA, which retains intron 3, predicts the existence of a 121-amino acid protein, survivin(121) that lacks the coiled-coil domain. Removal of exon 2-derived sequences by alternative pre-messenger RNA (mRNA) splicing results in a third 40-amino acid residue protein, survivin(40), lacking the IAP repeat and coiled-coil structure. Predictably, only recombinant survivin(140) and survivin(121) inhibited caspase-3 activity. All 3 mRNA species were variably expressed during development from 7.5 days postcoitum. Of the adult tissues surveyed, thymus and testis accumulated high levels of survivin(140) mRNA, whereas survivin(121)-specific transcripts were detected in all tissues, while those representing survivin(40) were absent. Human counterparts to the 3 survivin mRNA transcripts were identified in a study of human cells and tissues. The presence of distinct isoforms of survivin that are expressed differentially suggests that survivin plays a complex role in regulating apoptosis. (Blood. 2000;95:1435-1442)

Recurrent molecular deletion of the 12p13 region, centromeric to ETV6/TEL, in T-cell prolymphocytic leukemia.

INTRODUCTION: T-cell prolymphocytic leukemia is a rare form of mature leukemia which occurs in adults and in younger patients suffering ataxia telangiectasia. Among others, complex chromosome aberrations of chromosome 12 have been described in this disease. We searched for deletions of the 12p13 region as the result of these chromosome rearrangements. MATERIAL AND METHODS: Paired leukemic and non-leukemic cells were obtained from a series of 21 patients suffering T-cell prolymphocytic leukemia. Loss of heterozygosity was searched for by microsatellite typing using a fluorescent automated laser DNA sequencer to analyze the amplification products. Proteins were analyzed by Western blot. Southern blot analysis of one patient was conducted. RESULTS AND CONCLUSION: Loss of heterozygosity of the 12p13 region, including the ETV6 and CDKN1B genes, was detected in nine of these 21 cases (43%). Western and Southern blot analyses of one case demonstrated a biallelic deletion which did not include ETV6. Taken together, our results defined a minimal region of deletion of less than one Mb flanked by the markers b312C2T7 and D12S320, excluding ETV6 as a candidate gene. Deletion of the 12p13 region is thus a highly recurrent genetic event in T-cell prolymphocytic leukemia.

The apoptosis inhibitor gene API2 and a novel 18q gene, MLT, are recurrently rearranged in the t(11;18)(q21;q21) associated with mucosa-associated lymphoid tissue lymphomas.

Marginal zone cell lymphomas of the mucosa-associated lymphoid tissue (MALT) are the most common subtype of lymphoma arising at extranodal sites. The t(11;18)(q21;q21) appears to be the key genetic lesion and is found in approximately 50% of cytogenetically abnormal low-grade MALT lymphomas. We show that the API2 gene, encoding an inhibitor of apoptosis also known as c-IAP2, HIAP1, and MIHC, and a novel gene on 18q21 characterized by several Ig-like C2-type domains, named MLT, are recurrently rearranged in the t(11;18). In both MALT lymphomas analyzed, the breakpoint in API2 occurred in the intron separating the exons coding respectively for the baculovirus IAP repeat domains and the caspase recruitment domain. The breakpoints within MLT differed but the open reading frame was conserved in both cases. In one case, the translocation was accompanied by a cryptic deletion involving the 3' part of API2. As a result, the reciprocal transcript was not present, strongly suggesting that the API2-MLT fusion is involved in the oncogenesis of MALT lymphoma.

A physical, transcript, and deletion map of chromosome region 12p12.3 flanked by ETV6 and CDKN1B: hypermethylation of the LRP6 CpG island in two leukemia patients with hemizygous del(12p).

FISH analyses and loss of heterozygosity studies have delineated a commonly deleted region in hematological malignancies flanked by ETV6 and CDKN1B on chromosome 12p12.3. The same chromosomal region is also a target for deletions in certain solid tumors. As an initial step toward the cloning of a potential tumor suppressor gene at 12p12.3, we mapped the ETV6-CDKN1B region physically using bacterial artificial chromosome (BAC) and P1-derived clone (PAC) contigs. The 1.2-Mb high-resolution, contiguous map extends from D12S1095 to D12S929 and consists of 19 PACs and 20 BACs. Pulsed-field gel electrophoresis experiments confirmed the integrity of the clone-based map and identified six CpG islands in the region. A transcript map was generated by performing hybridization selection experiments with the genomic clones, by evaluating known 12p ESTs for their presence in the contig, and by sequence analysis of CpG islands in the region. Altogether evidence was gathered for the presence of the recently published LRP6 gene and at least seven other new genes in this chromosomal region. The CLAPS3 gene, mapped between D12S391 and D12S358, was reassigned to chromosome 5 since genomic sequencing demonstrated the chromosome 12p sequence to be a pseudogene. Polymorphic CA repeats were identified approximately every 100 kb, which will support future analysis of loss of heterozygosity in tumors. Fluorescence in situ hybridization analysis of leukemia patients with del(12p) further refined the commonly deleted segment to 600 kb between ETV6 and D12S358, which apparently excludes CDKN1B. Methylation changes of the CpG islands in the ETV6-CDKN1B interval were assessed by Southern analysis for leukemia patients with hemizygous 12p deletions. A "de novo" methylation was detected only at the LRP6 CpG island in 2 of 22 leukemia patients tested and was confirmed by methylation-sensitive PCR and sequencing. The genomic structure of LRP6 was elucidated to allow screening for inactivating mutations, but only intragenic polymorphisms were identified. Hypermethylation of CpG islands associated with gene promoters is reported as a common mechanism for gene silencing and tumor suppressor inactivation. Therefore the consequences of the LRP6 CpG island methylation and its role in the observed phenotype need further investigation.

Rearrangement between the MYH11 gene at 16p13 and D12S158 at 12p13 in a case of acute myeloid leukemia M1 (AML-M1).

A case of acute myeloid leukemia (AML) M1 with bone marrow eosinophilia was characterized by cytogenetics and fluorescence in situ hybridization (FISH). A complex karyotype including a der(12)t(12;17)(p12-13;q11) and a der(16)t(16;20)(p13;p11) was found at diagnosis. FISH studies with probes for chromosome 16 and for the short arm of chromosome 12 showed even more complex rearrangements. Analysis with a panel of probes for 12p showed that D12S158 spanned the breakpoint on the der(12). Unexpectedly, FISH signals were found on the der(12) and on the der(6) at band p13, the site of juxtaposition between the short arm of chromosome 16 and chromosome 20. Moreover, both YAC 854E2, containing the MYH11 gene, and cosmid ZIT133, encompassing the MYH11 breakpoint in inv(16) and t(16;16) of AML-M4 with eosinophilia, demonstrated fluorescent signals on the normal 16, on the der(16), and on the der(12). These data clearly support a reciprocal exchange between D12S158 at 12p13.3 and the MYH11 gene at 16p13. In addition, experiments with two PAC clones for the CBFB gene at 16q22 excluded the presence of a masked inv(16). An interstitial deletion, independent from the translocation and flanked by VWF and KRAS2, was also detected on the der(12).

Fluorescence in situ hybridization characterization of new translocations involving TEL (ETV6) in a wide spectrum of hematologic malignancies.

The ETV6 (also known as TEL) gene on chromosome 12p13 is the target of a number of translocations associated with various hematologic malignancies. The contribution of ETV6 to leukemogenesis occurs through different mechanisms that involve either its helix-loop-helix dimerization domain or its E26 transformation-specific (ETS) DNA-binding domain. Using fluorescence in situ hybridization we characterized seven new ETV6 rearrangements in chronic myeloid leukemia, acute myeloid leukemia, acute lymphoblastic leukemia, and non-Hodgkin's lymphoma. These aberrations, not always discernible at the cytogenetic level, include a t(5;12)(q31;p13), t(6;12;17)(p21;p13;q25), t(7;12)(p15;p13), t(7;12)(p12;p13), t(7;12)(q36;p13), t(12;13)(p13;q12), and a not completely defined t(12;?)(p13;?). Loss or disruption of the second ETV6 allele by a del(12)(p12p13) or by an intragenic ETV6 deletion was detected in two cases. In six cases the 12p13 breakpoint occurred in the 5' end of ETV6, upstream to exons encoding the HLH domain, whereas the remaining case had a breakpoint between the exons coding for the HLH domain and the exons coding for the ETS domain of ETV6. These observations provide further evidence for the multiple contributions of ETV6 in the pathogenesis of a wide range of hematologic malignancies.

Fusion of TEL, the ETS-variant gene 6 (ETV6), to the receptor-associated kinase JAK2 as a result of t(9;12) in a lymphoid and t(9;15;12) in a myeloid leukemia.

Translocations in hematologic disease of myeloid or lymphoid origin with breakpoints at chromosome band 12p13 frequently result in rearrangements of the Ets variant gene 6 (ETV6). As a consequence either the ETS DNA-binding domain or the Helix-Loop-Helix (HLH) oligomerization domain of ETV6 is fused to different partner genes. We show here that a t(9;12)(p24;p13) in a case of early pre-B acute lymphoid leukemia and a t(9;15;12)(p24;q15;p13) in atypical chronic myelogenous leukemia in transformation involve the ETV6 gene at 12p13 and the JAK2 gene at 9p24. In each case different fusion mRNAs were found, with only one resulting in an open reading frame for a chimeric protein consisting of the HLH oligomerization domain of ETV6 and the protein tyrosine kinase (PTK) domain of JAK2. The cloning of the complete human JAK2 coding and genomic sequences and of the genomic junction fragments of the translocations allowed a characterization of the different splice events leading to the various mRNAs. JAK2 plays a central role in non-protein tyrosine kinase receptor signaling pathways, which could explain its involvement in malignancies of different hematologic lineages. Besides hop in Drosophila no member of the JAK family has yet been implicated in tumorigenesis.