The Yin and Yang-Like Clinical Implications of the CDKN2A/ARF/CDKN2B Gene Cluster in Acute Lymphoblastic Leukemia.

Acute lymphoblastic leukemia (ALL) is a malignant clonal expansion of lymphoid hematopoietic precursors that exhibit developmental arrest at varying stages of differentiation. Similar to what occurs in solid cancers, transformation of normal hematopoietic precursors is governed by a multistep oncogenic process that drives initiation, clonal expansion and metastasis. In this process, alterations in genes encoding proteins that govern processes such as cell proliferation, differentiation, and growth provide us with some of the clearest mechanistic insights into how and why cancer arises. In such a scenario, deletions in the 9p21.3 cluster involving CDKN2A/ARF/CDKN2B genes arise as one of the oncogenic hallmarks of ALL. Deletions in this region are the most frequent structural alteration in T-cell acute lymphoblastic leukemia (T-ALL) and account for roughly 30% of copy number alterations found in B-cell-precursor acute lymphoblastic leukemia (BCP-ALL). Here, we review the literature concerning the involvement of the CDKN2A/B genes as a prognosis marker of good or bad response in the two ALL subtypes (BCP-ALL and T-ALL). We compare frequencies observed in studies performed on several ALL cohorts (adult and child), which mainly consider genetic data produced by genomic techniques. We also summarize what we have learned from mouse models designed to evaluate the functional involvement of the gene cluster in ALL development and in relapse/resistance to treatment. Finally, we examine the range of possibilities for targeting the abnormal function of the protein-coding genes of this cluster and their potential to act as anti-leukemic agents in patients.

Multiple genome analyses reveal key genes in Vitamin C and Vitamin D synthesis and transport pathways are shared.

Vitamin C (VC) and vitamin D (VD) have been widely used as the dietary supplements and in treatment of diseases both independently and in combination. Whether there is a connection between their pathways is critical for their therapeutic applications. Using whole-genome expression profiles, we performed multiple measures of associations, networks, eQTL mappings and expressions of key genes of interest in VC and VD functions. Several key genes in their pathways were found to be associated. Gc and Rgn play important roles connecting VC and VD pathways in mice. The r values of expression levels between Gc and Rgn in mouse spleen, liver, lung, and kidney are 0.937, 0.558, 0.901, and 0.617, respectively. The expression QTLs of Gc and Rgn are mapped onto the same locations, i.e., 68-76 MB in chromosome 7 and 26-36 MB in chromosome 9. In humans, there are positive correlations between CYP27B1 and SLC23A1 expression levels in kidney (r = 0.733) and spleen (r = 0.424). SLC23A2 and RXRA are minimally associated in both mouse and human. These data indicate that pathways of VC and VD are not independent but affect each other, and this effect is different between mice and humans during VC and VD synthesis and transportation.

Therapy-related myeloid neoplasm in an 18-year-old boy with B-lymphoblastic leukemia.

BACKGROUND: Acute lymphoblastic leukemia (ALL) is the most common pediatric malignancy. Acute myeloid leukemia or myelodysplastic syndrome during the course of ALL is a rare entity. Some of these cases are therapy-related while the others occur due to lineage switch. The correct diagnosis relies on comparing the immunophenotypes and cytogenetic/molecular alterations of the myeloid neoplasm and the ALL. We present the clinical, pathologic and cytogenetic features of a case of an 18-year-old male with prior treatment for B-lymphoblastic leukemia (B-ALL) who developed therapy-related myeloid neoplasm (t-MN) 4-5years after his initial diagnosis of B-ALL. CASE PRESENTATION: A 13-year-old boy with no significant past medical history presented to Harbor-UCLA Medical Center (HUMC) in November 2012 with night sweats, fevers and chills, nausea, vomiting, diarrhea, fatigue, weakness, and weight loss. Peripheral blood flow cytometric analysis disclosed B-ALL. The blasts expressed CD10, CD19, CD22 (dim), CD34, CD38, HLA-DR, and TdT, and were negative for CD20, CD13, CD33, CD117, and cytoplasmic MPO. Chromosomal analysis and a supplemental fluorescence in situ hybridization (FISH) study performed on the bone marrow aspirate showed an abnormal karyotype (47,XY,+X,del(9)(p21p21)[4]/46,XY[16]). He achieved remission after induction chemotherapy and remained in remission until March 2016 when bilateral testicular masses were noted. Biopsy of the left testicular mass showed relapsed B-ALL. Cerebrospinal fluid (CSF) contained rare TdT-positive blasts, suggestive of minimal/early involvement by B-ALL. However, there was no evidence of acute leukemia in his bone marrow at this time. He was then treated with COG protocol AALL1331 randomized to blinatumomab arm and achieved second remission. In June 2017, the patient's peripheral blood smear showed 11% circulating monoblasts. By flow cytometry, the blasts expressed CD4, CD11b, CD13, CD15, CD33, CD38, CD56, and CD64. In addition, a few TdT-positive blasts were seen in his CSF cytospin smear. Bone marrow biopsy was subsequently performed which was consistent with evolving acute myeloid leukemia. A diagnosis of myeloid neoplasm, consistent with t-MN was made. Chromosomal analysis and FISH studies performed on his bone marrow aspirate showed normal karyotype (46,XY[20]), negative FISH result for deletion 9p21 locus, and positive KMT2A (MLL) rearrangement, respectively. Despite of chemotherapy, the patient died within one month after diagnosis. DISCUSSION AND CONCLUSION: Diagnosis of t-MN should be suspected in patients with a history of receiving cytotoxic agents and/or irradiation. In this case study, we diagnosed t-MN with KMT2A rearrangement in a patient with history of B-ALL with 9p deletion and gain of X chromosome. Unusual features associated with this case are discussed.

Practical Approaches for Whole-Genome Sequence Analysis of Heart- and Blood-Related Traits.

Whole-genome sequencing (WGS) allows for a comprehensive view of the sequence of the human genome. We present and apply integrated methodologic steps for interrogating WGS data to characterize the genetic architecture of 10 heart- and blood-related traits in a sample of 1,860 African Americans. In order to evaluate the contribution of regulatory and non-protein coding regions of the genome, we conducted aggregate tests of rare variation across the entire genomic landscape using a sliding window, complemented by an annotation-based assessment of the genome using predefined regulatory elements and within the first intron of all genes. These tests were performed treating all variants equally as well as with individual variants weighted by a measure of predicted functional consequence. Significant findings were assessed in 1,705 individuals of European ancestry. After these steps, we identified and replicated components of the genomic landscape significantly associated with heart- and blood-related traits. For two traits, lipoprotein(a) levels and neutrophil count, aggregate tests of low-frequency and rare variation were significantly associated across multiple motifs. For a third trait, cardiac troponin T, investigation of regulatory domains identified a locus on chromosome 9. These practical approaches for WGS analysis led to the identification of informative genomic regions and also showed that defined non-coding regions, such as first introns of genes and regulatory domains, are associated with important risk factor phenotypes. This study illustrates the tractable nature of WGS data and outlines an approach for characterizing the genetic architecture of complex traits.

A Functional SNP in BNC2 Is Associated with Adolescent Idiopathic Scoliosis.

Adolescent idiopathic scoliosis (AIS) is the most common spinal deformity. We previously conducted a genome-wide association study (GWAS) and detected two loci associated with AIS. To identify additional loci, we extended our GWAS by increasing the number of cohorts (2,109 affected subjects and 11,140 control subjects in total) and conducting a whole-genome imputation. Through the extended GWAS and replication studies using independent Japanese and Chinese populations, we identified a susceptibility locus on chromosome 9p22.2 (p = 2.46 × 10(-13); odds ratio = 1.21). The most significantly associated SNPs were in intron 3 of BNC2, which encodes a zinc finger transcription factor, basonuclin-2. Expression quantitative trait loci data suggested that the associated SNPs have the potential to regulate the BNC2 transcriptional activity and that the susceptibility alleles increase BNC2 expression. We identified a functional SNP, rs10738445 in BNC2, whose susceptibility allele showed both higher binding to a transcription factor, YY1 (yin and yang 1), and higher BNC2 enhancer activity than the non-susceptibility allele. BNC2 overexpression produced body curvature in developing zebrafish in a gene-dosage-dependent manner. Our results suggest that increased BNC2 expression is implicated in the etiology of AIS.

Omega-3 fatty acids and the genetic risk of early onset acute coronary syndrome.

BACKGROUND AND AIMS: Recent gene-environment interaction studies suggest that diet may influence an individual's genetic predisposition to cardiovascular risk. We evaluated whether omega-3 fatty acid intake may influence the risk for acute coronary syndrome (ACS) conferred by genetic polymorphisms among patients with early onset ACS. METHODS AND RESULTS: Our population consisted of 705 patients of white European descent enrolled in GENESIS-PRAXY, a multicenter cohort study of patients aged 18-55 years and hospitalized with ACS. We used a case-only design to investigate interactions between the omega-3 index (a validated biomarker of omega-3 fatty acid intake) and 30 single nucleotide polymorphisms (SNPs) robustly associated with ACS. We used logistic regression to assess the interaction between each SNP and the omega-3 index. Interaction was also assessed between the omega-3 index and a genetic risk score generated from the 30 SNPs. All models were adjusted for age and sex. An interaction for increased ACS risk was found between carriers of the chromosome 9p21 variant rs4977574 and low omega-3 index (OR 1.57, 95% CI 1.07-2.32, p = 0.02), but this was not significant after correction for multiple testing. Similar results were obtained in the adjusted model (OR 1.55, 95% CI 1.05-2.29, p = 0.03). We did not observe any interaction between the genetic risk score or any of the other SNPs and the omega-3 index. CONCLUSION: Our results suggest that omega-3 fatty acid intake may modify the genetic risk conferred by chromosome 9p21 variation in the development of early onset ACS and requires independent replication.

CKIT mutation in therapy-related acute myeloid leukemia with MLLT3/MLL chimeric transcript from t(9;11)(p22;q23).

Gain-of-function mutations of the CKIT gene have been reported to specifically occur in core-binding factor (CBF) acute myeloid leukemia (AML) with a poor prognostic implication. Here we report a case of therapy-related AML with t(9;11)(p22;q23) who had CKIT mutation. A 48-year-old woman with breast cancer received partial mastectomy followed by 6 cycles of adjuvant chemotherapy and radiation therapy. At 28 months from the diagnosis of breast cancer, she was diagnosed as having AML with blasts 81% in bone marrow. Cytogenetic analysis revealed t(9;11)(p22;q23), and FISH showed 96.5% of MLL break-apart signals. RT-PCR study revealed MLL(11q23)/MLLT3(9p22) chimeric transcript. FLT3-ITD and NPM1 mutations were both negative. Unexpectedly, mutation analyses for CKIT identified D816Y mutation. The patient received induction chemotherapy and achieved complete remission at 1 month. To the best of our knowledge, this is the first report on CKIT mutation in therapy-related AML with MLL rearrangement.

Quantitative real time-polymerase chain reaction method in Bcr-Abl translocation diagnostics.

PURPOSE: The quantitative real-time polymerase chain reaction (qRT-PCR) is used in the detection of molecular events involved in leukemogenesis, such as the Bcr-Abl gene translocation, the most important factor in the pathogenesis of chronic myeloid leukaemia (CML). The main aim of our study was to test the reproducibility, specificity and sensitivity of the qRT-PCR in the detection of Bcr-Abl gene translocation. METHODS: In complementary (c)DNA, isolated from K562 Bcr-Abl positive cell line, we performed qRT-PCR analysis with Bcr-Abl specific primers. For qRT-PCR analysis, we used serial dilutions of the newly synthesized cDNA in order to establish the detection threshold of this method. RESULTS: Using the specific primers for the Bcr-Abl translocation, we obtained the specific translocation product in cDNA sample of K562 human erythroid leukemia cell line. qRT- PCR showed significant sensitivity with the detection threshold for the Bcr-Abl fluorescent signal, which enabled the precise detection that was accurate within a 10-fold dilution range, and a dynamic range of 5 orders of magnitude. CONCLUSION: The results of our study showed that the application of the qRT-PCR is the optimal method for the detection of Bcr-Abl gene translocation, characterized by high reproducibility, specificity and sensitivity.

Impact of adding a single allele in the 9p21 locus to traditional risk factors on reclassification of coronary heart disease risk and implications for lipid-modifying therapy in the Atherosclerosis Risk in Communities study.

BACKGROUND: A single-nucleotide polymorphism on chromosome 9p21, rs10757274 (9p21 allele), has been shown to predict coronary heart disease (CHD) in whites. We evaluated whether adding the 9p21 allele to traditional risk factors (RFs) improved CHD risk prediction in whites from the Atherosclerosis Risk in Communities study and whether changes in risk prediction would modify lipid therapy recommendations. METHODS AND RESULTS: Whites (n=9998) in the Atherosclerosis Risk in Communities study for whom the 9p21 genotype and traditional RF information was available were included. Using Cox proportional hazards models, the Atherosclerosis Risk in Communities Cardiovascular Risk Score, which is based on traditional RFs, was determined. A total of 1349 individuals (13.5%) developed incident CHD events during a period of 14.6 years. Adding the 9p21 allele to traditional RFs was associated with a hazard ratio of incident CHD of 1.2 per allele (P<0.000003) and a significant increase in the area under the curve of the receiver operating characteristic from 0.782 to 0.786 (95% CI, 0.001, 0.007). The 9p21 allele's greatest influence to the Atherosclerosis Risk in Communities Cardiovascular Risk Score was observed in the intermediate-low (>5% to 10% to 100 mg/dL. CONCLUSIONS: Adding the 9p21 allele to traditional RFs in whites in the Atherosclerosis Risk in Communities study modestly improved CHD risk prediction in the intermediate categories.

Susceptibility to coronary artery disease and diabetes is encoded by distinct, tightly linked SNPs in the ANRIL locus on chromosome 9p.

Genome-wide association studies have identified a region on chromosome 9p that is associated with coronary artery disease (CAD). The region is also associated with type 2 diabetes (T2D), a risk factor for CAD, although different SNPs were reported to be associated to each disease in separate studies. We have undertaken a case-control study in 4251 CAD cases and 4443 controls in four European populations using previously reported ('literature') and tagging SNPs. We replicated the literature SNPs (P = 8x10(-13); OR = 1.29; 95% CI: 1.20-1.38) and showed that the strong consistent association detected by these SNPs is a consequence of a 'yin-yang' haplotype pattern spanning 53 kb. There was no evidence of additional CAD susceptibility alleles over the major risk haplotype. CAD patients without myocardial infarction (MI) showed a trend towards stronger association than MI patients. The CAD susceptibility conferred by this locus did not differ by sex, age, smoking, obesity, hypertension or diabetes. A simultaneous test of CAD and diabetes susceptibility with CAD and T2D-associated SNPs indicated that these associations were independent of each other. Moreover, this region was not associated with differences in plasma levels of low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, fibrinogen, albumin, uric acid, bilirubin or homocysteine, although the CAD-high-risk allele was paradoxically associated with lower triglyceride levels. A large antisense non-coding RNA gene (ANRIL) collocates with the high-risk haplotype, is expressed in tissues and cell types that are affected by atherosclerosis and is a prime candidate gene for the chromosome 9p CAD locus.

JAK2 V617F tyrosine kinase mutation in cell lines derived from myeloproliferative disorders.

A mutation in the JH2 pseudokinase domain of the Janus kinase 2 gene (JAK2 V617F) has been described in chronic myeloproliferative disorders (MPD). We screened 79 acute myeloid leukemia (AML) cell lines and found five positive for JAK2 V617F (HEL, MB-02, MUTZ-8, SET-2, UKE-1), 4/5 with histories of MPD/MDS. While SET-2 expressed both mutant (mu) and wild-type (wt) JAK2, remaining positives carried homo-/hemizygous JAK2 mutations. Microsatellite analysis confirmed losses of heterozygosity (LOH) affecting the JAK2 region on chromosome 9p in MB-02, MUTZ-8 and UKE-1, but also in HEL, the only JAK2mu cell line lacking any reported MPD/MDS history. All five JAK2mu cell lines displayed cytogenetic hallmarks of MDS, namely losses of 5q or 7q, remarkably in 4/5 cases affecting both chromosomes. Our combined FISH and microsatellite analysis uncovered a novel mechanism to supplement mitotic recombination previously proposed to explain JAK2 LOH, namely chromosome deletion with/without selective JAK2mu amplification. Confirming the importance of the mutated JAK2 protein for growth and prevention of apoptosis, JAK2mu cell lines displayed higher sensitivities to JAK2 inhibition than JAK2wt cell lines. In summary, JAK2 V617F cell lines, derived from patients with history of MPD/MDS, represent novel research tools for elucidating the pathobiology of this JAK2 mutation.

Cellular drug sensitivity in MLL-rearranged childhood acute leukaemia is correlated to partner genes and cell lineage.

Rearrangements in the 11q23 region, the site of the mixed lineage leukaemia (MLL) gene, are found in both childhood acute myeloid (AML) and lymphoblastic (ALL) leukaemia. We studied the in vitro drug resistance by the fluorometric microculture cytotoxicity assay (FMCA) in 132 children with AML and 178 children with ALL (aged 0-17 years). In AML, children with t(9;11) (n = 10) were significantly more sensitive to cytarabine (P < 0.001) and doxorubicin (P = 0.005) than non-11q23 rearranged patients (n = 108). Children with other 11q23 rearrangements (n = 14) differed less from non-rearranged children. The 'AML-profile' common to all three groups included relative resistance to glucocorticoids and vincristine. In ALL, children with 11q23 rearrangement (n = 22) were significantly more sensitive to cytarabine (P = 0.026) than children without 11q23 rearrangement (n = 156), also after stratification for white blood cell count. In conclusion, the findings indicate that the cellular drug resistance is correlated to both the cell lineage and the type of 11q23 rearrangement. High cellular sensitivity to cytarabine and doxorubicin might explain the excellent treatment results in children with AML and t(9;11). The present study supports the strategy of contemporary protocols to include high-dose cytarabine in the treatment of 11q23-positive patients both in AML and ALL.

Coenzyme Q deficiency and cerebellar ataxia associated with an aprataxin mutation.

Primary muscle coenzyme Q10 (CoQ10) deficiency is an apparently autosomal recessive condition with heterogeneous clinical presentations. Patients with these disorders improve with CoQ10 supplementation. In a family with ataxia and CoQ10 deficiency, analysis of genome-wide microsatellite markers suggested linkage of the disease to chromosome 9p13 and led to identification of an aprataxin gene (APTX) mutation that causes ataxia oculomotor apraxia (AOA1 [MIM606350]). The authors' observations indicate that CoQ10 deficiency may contribute to the pathogenesis of AOA1.

VE-statin, an endothelial repressor of smooth muscle cell migration.

The recruitment and proliferation of smooth muscle cells and pericytes are two key events for the stabilization of newly formed capillaries during angiogenesis and, when out of control in the adult, are the main causes of arteriosclerosis. We have identified a novel gene, named VE-statin for vascular endothelial-statin, which is expressed specifically by endothelial cells of the developing mouse embryo and in the adult, and in early endothelial progenitors. The mouse and human VE-statin genes have been located on chromosome 2 and 9, respectively, they span >10 kbp and are transcribed in two major variants arising from independent initiation sites. The VE-statin transcripts code for a unique protein of 30 kDa that contains a signal peptide and two epidermal growth factor (EGF)-like modules. VE-statin is found in the cellular endoplasmic reticulum and secreted in the cell supernatant. Secreted VE-statin inhibits platelet-derived growth factor (PDGF)-BB-induced smooth muscle cell migration, but has no effects on endothelial cell migration. VE-statin is the first identified inhibitor of mural cell migration specifically produced by endothelial cells.

Lysophosphatidic acid acyltransferase-beta: a novel target for induction of tumour cell apoptosis.

Phosphatidic acid (PA) is a component of cellular membranes that is also a mediator of certain cell signalling functions associated with oncogenesis. These include ras/raf/Erk and Akt/mTor [1-3]. The authors have investigated whether it would be possible to interrupt these known oncogenic pathways through the inhibition of lysophosphatidic acid acyltransferase (LPAAT), an enzyme that catalyses the biosynthesis of PA. The expression and activity of the LPAAT-beta isoform are elevated in human tumours, and the respective gene displays transforming capacity when overexpressed in vitro. Inhibition by either genetic means or by isoform-specific small molecules results in a block to cell signalling pathways and apoptosis. Furthermore, the small-molecule inhibitors of LPAAT-beta are not cytotoxic to a number of normal cell types, including primary bone marrow progenitors, indicating a differential dependence of tumour cells on LPAAT-beta function. These discoveries indicate that LPAAT-beta represents a potential novel cancer therapy target.

Dystonia update.

PURPOSE OF REVIEW: Dystonia is a movement disorder with a complex and not fully understood pathophysiology. Its better understanding would enable more focused treatment for the disorder. In this review, we provide an overview of recent studies of the pathophysiology of primary and secondary dystonia, with an emphasis on functional brain imaging. Potential mechanisms underlying the beneficial effects of deep brain stimulation for dystonia are also summarized. RECENT FINDINGS: The recognition of dysfunction at different levels of the nervous system has extended the classical notions of localized striatal abnormalities in primary dystonia. Recent biochemical studies have revealed evidence of abnormal torsion activity in DYT1 dystonia. Abnormal patterns of brain metabolism have also been identified using functional brain imaging in different dystonia genotypes. These findings, in conjunction with new electrophysiological techniques, can be utilized to help define a common mechanism for the neural dysfunction in dystonia. SUMMARY: New insights into the pathophysiology of dystonia have been provided by recent studies using electrophysiology, biochemistry and human genetics, as well as functional brain imaging studies. These advances together may create the basis for new therapies for this disorder.

Serum selenium levels in relation to markers of neoplastic progression among persons with Barrett's esophagus.

BACKGROUND: Persons with Barrett's esophagus have a substantially greater risk of esophageal adenocarcinoma than the general population. Higher serum selenium levels have been associated with a reduced risk of several cancers; however, their association with the risk of esophageal adenocarcinoma is unknown. We used a cross-sectional study to investigate the relationship between serum selenium levels and markers of neoplastic progression among persons with Barrett's esophagus. METHODS: Medical history, blood, and esophageal tissue specimens were collected from 399 members of a cohort study of Barrett's esophagus patients undergoing endoscopic surveillance. Serum selenium levels were measured by flameless atomic absorption spectrophotometry. DNA content of tissue samples was measured by flow cytometry. Loss of heterozygosity (LOH) at 9p and 17p, chromosomal regions which include the p16 and p53 tumor suppressors, respectively, was detected by automated fluorescent genotyping. Logistic regression was used to calculate odds ratios (ORs) and 95% confidence intervals (CIs). All statistical tests were two-sided. RESULTS: Persons with serum selenium levels in the upper three quartiles (i.e., >1.5 micro M) were less likely to have high-grade dysplasia (OR = 0.5, 95% CI = 0.3 to 0.9) or aneuploidy (OR = 0.4, 95% CI = 0.2 to 0.8) than those with levels in the lowest quartile. Serum selenium levels in the upper three quartiles were associated with similar reductions in risk of 17p (p53) LOH (OR = 0.5, 95% CI = 0.2 to 0.9) and increased 4N fraction (OR = 0.6, 95% CI = 0.3 to 1.2). By contrast, serum selenium levels were not associated with 9p (p16) LOH (OR = 1.0, 95% CI = 0.5 to 1.7), a marker that appears early in neoplastic progression. CONCLUSION: Our preliminary results, from a cross-sectional analysis with biologic markers, suggest that higher serum selenium levels may be associated with a reduced risk of esophageal adenocarcinoma among persons with Barrett's esophagus. Because serum selenium was not associated with 9p (p16) LOH, we speculate that selenium may act primarily at later stages of progression toward adenocarcinoma.

Human neuronal calcium sensor-1 shows the highest expression level in cerebral cortex.

Neuronal calcium sensors (NCS) are important constituents in the intracellular signaling pathways. A novel human gene, NCS-1, was identified in the present study. Among the 16 human tissues examined, NCS-1 is expressed most abundantly in the brain. Among the brain regions, the expression level of NCS-1 in cerebral cortex is the highest, which is about six times higher than the average level of the whole brain and a hundred times higher than the spinal cord. In the 12 different anatomical regions of human brain, the expression level of NCS-1 is very high in the temporal lobe, occipital pole, frontal lobe, thalamus, amygdala and hippocampus; moderate in cerebellum, putamen, caudate nucleus; low in the medulla, substantia nigra and the lowest in corpus callosum. Our results suggest that NCS-1 in human brain might be involved in a variety of brain functions such as sensory processing, motor control, emotional control, learning and memory.

The human formin-binding protein 17 (FBP17) interacts with sorting nexin, SNX2, and is an MLL-fusion partner in acute myelogeneous leukemia.

We have cloned a fusion partner of the MLL gene at 11q23 and identified it as the gene encoding the human formin-binding protein 17, FBP17. It maps to chromosome 9q34 centromeric to ABL. The gene fusion results from a complex chromosome rearrangement that was resolved by fluorescence in situ hybridization with various probes on chromosomes 9 and 11 as an ins(11;9)(q23;q34)inv(11)(q13q23). The rearrangement resulted in a 5'-MLL/FBP17-3' fusion mRNA. We retrovirally transduced murine-myeloid progenitor cells with MLL/FBP17 to test its transforming ability. In contrast to MLL/ENL, MLL/ELL and other MLL-fusion genes, MLL/FBP17 did not give a positive readout in a serial replating assay. Therefore, we assume that additional cooperating genetic abnormalities might be needed to establish a full malignant phenotype. FBP17 consists of a C-terminal Src homology 3 domain and an N-terminal region that is homologous to the cell division cycle protein, cdc15, a regulator of the actin cytoskeleton in Schizosaccharomyces pombe. Both domains are separated by a consensus Rho-binding motif that has been identified in different Rho-interaction partners such as Rhotekin and Rhophilin. We evaluated whether FBP17 and members of the Rho family interact in vivo with a yeast two-hybrid assay. None of the various Rho proteins tested, however, interacted with FBP17. We screened a human kidney library and identified a sorting nexin, SNX2, as a protein interaction partner of FBP17. These data provide a link between the epidermal growth factor receptor pathway and an MLL fusion protein.

Cloning and characterization of LUN, a novel ring finger protein that is highly expressed in lung and specifically binds to a palindromic sequence.

We isolated cDNAs encoding a novel RING finger protein (LUN), the mRNAs of which were expressed at high levels in the lung. In situ hybridization revealed that LUN mRNAs were expressed in the alveolar epithelium of the lung. The LUN gene locus was assigned to chromosome 9p21, which contains candidate tumor suppressor genes associated with loss of heterozygosity in more than 86% of small cell lung cancers. We clarified that LUN is localized to the nucleus and reveals Zn(2+)-dependent DNA binding activity. The region from amino acids 51 to 374 of LUN is responsible for DNA binding. Furthermore, we identified a novel palindromic binding consensus (5'-TCCCAGCACTTTGGGA-3') for the LUN binding. Interestingly, this LUN binding palindromic sequence is found in the upstream transcriptional regulatory region of the E-cadherin gene and two intervening regions of the talin gene. Our results suggested that LUN might be an important trans-acting transcriptional regulator for lung cancer-associated genes including E-cadherin and talin genes.