While a number of growth factors have been described that are highly specific for particular cell lineages, neither a factor nor a receptor uniquely specific to the skeletal muscle lineage has previously been described. Here we identify a receptor tyrosine kinase (RTK) specific to skeletal muscle, which we term "MuSK" for muscle-specific kinase. MuSK is expressed at low levels in proliferating myoblasts and is induced upon differentiation and fusion. In the embryo, it is specifically expressed in early myotomes and developing muscle. MuSK is then dramatically down-regulated in mature muscle, where it remains prominent only at the neuromuscular junction; MuSK is thus the only known RTK that localizes to the neuromuscular junction. Strikingly, MuSK expression is dramatically induced throughout the adult myofiber after denervation, block of electrical activity, or physical immobilization. In humans, MuSK maps to chromosome 9q31.3-32, which overlaps with the region reported to contain the Fukuyama muscular dystrophy mutation. Identification of MuSK introduces a novel receptor-factor system that seems sure to play an important and selective role in many aspects of skeletal muscle development and function.

Loss of a whole chromosome 5 or a deletion of the long arm, del(5q), is a recurring abnormality in myelodysplastic syndromes (MDSs) and acute myeloid leukemia (AML). To identify a leukemiarelated gene on chromosome 5, we previously delineated a 970-kb segment of 5q31 that is deleted in all patients examined, and prepared a transcript map of this region.

One of the most serious consequences of cytotoxic cancer therapy is the development of therapy-related acute myeloid leukemia (t-AML), a neoplastic disorder arising from a multipotential hematopoietic stem cell. To gain insights into the molecular basis of this disease, we performed gene expression profiling of CD34 ؉ hematopoietic progenitor cells from t-AML patients. Our analysis revealed that there are distinct subtypes of t-AML that have a characteristic gene expression pattern. T herapy-related myelodysplastic syndrome (t-MDS) and acute myeloid leukemia (t-AML) are late complications of cytotoxic therapy (radiation and͞or chemotherapy) used in the treatment of both malignant and nonmalignant diseases (1-3). These neoplasms are thought to be the direct consequence of mutational events induced by cytotoxic therapy. Several distinct cytogenetic and clinical subtypes of t-MDS͞t-AML are recognized that are closely associated with the nature of the preceding treatment. Survival times of t-AML patients are short, because current forms of therapy are largely ineffective, and new therapeutic approaches are needed (1-4).The most common subtype of t-AML (Ϸ75% of cases) develops after exposure to alkylating agents and is characterized by loss of a whole chromosome 5 and͞or 7 or a deletion of the long arms of these chromosomes [Ϫ5͞del(5q), Ϫ7͞del(7q)] (4). Patients who develop t-MDS͞t-AML in this setting typically show a latency of 3-7 years from alkylating agent exposure and present with cytopenias and myelodysplasia. The median time until progression to t-AML is 6 mo. Frequently, all three hematopoietic cell lineages (erythroid, myeloid, and megakaryocytic) are involved in the dysplastic process, suggesting that this subtype of t-AML arises in a hematopoietic stem͞progenitor cell. The prognosis is poor, with a median survival of 5 mo. In the University of Chicago's series of 306 patients with t-MDS͞t-AML, 64 (21%) patients had abnormalities of chromosome 5; 85 (28%) patients had abnormalities of chromosome 7; and 65 (21%) patients had abnormalities of both chromosomes 5 and 7 (3).A second, and distinctly different, form of t-AML arises after therapy with topoisomerase II inhibitors (1-4). These patients are younger, rarely present with MDS, and have a more favorable response to remission induction therapy (1, 2). Balanced translocations involving MLL at 11q23 or RUNX1͞AML1 at 21q22 are common in this subgroup (4). t-AML represents an important model for cancer for several reasons. First, the incidence of t-AML is rising, as a result of the increasing number of cancer survivors at risk of developing this disorder and the changes in therapeutic trends. Second, t-AML provides a unique opportunity to examine the effects of mutagens on carcinogenesis in humans, as well as the role of genetic susceptibility to cancer. Third, the mechanisms of leukemogenesis that are uncovered in t-AML will likely apply to those subtypes of AML de novo, which share the same cytogenetic abnormalities, e.g., AML de novo with abnormalities of ...

Loss of a whole chromosome 5 or a deletion of the long arm of chromosome 5, ؊5/del(5q), is a recurring abnormality in myeloid neoplasms. The APC gene is located at chromosome band 5q23, and is deleted in more than 95% of patients with a ؊5/del(5q), raising the question of whether haploinsufficiency of APC contributes to the development of myeloid neoplasms with loss of 5q. We show that conditional inactivation of a single allele IntroductionMyelodysplastic syndromes (MDSs) are a group of heterogeneous disorders of hematopoietic stem cells (HSCs) characterized by blood cytopenias due to ineffective hematopoiesis. 1 Approximately 30% to 40% of primary MDS transforms to acute myeloid leukemia (AML). 1 MDS can arise de novo or as a result of previous cytotoxic therapy (t-MDS), including chemotherapy, radiation therapy, and immunosuppressive therapy. Recurring karyotypic abnormalities, including Ϫ5/ del(5q), Ϫ7/del(7q), ϩ8, and del(20q), have been identified in approximately 50% of patients with a primary MDS. 2,3 Loss of a whole chromosome 5 or a del(5q) are among the most common recurring cytogenetic abnormalities, and are noted in approximately 10% to 15% of patients with primary MDS or AML de novo, and in more than 40% of patients with t-MDS/t-AML. 2,4 MDS with an isolated del(5q) (formerly termed the 5qϪ syndrome) is a distinct subtype of MDS, characterized by macrocytosis, anemia, and a low rate of leukemic transformation. 5 In contrast, the advanced stages of MDS, AML de novo, or t-MDS/t-AML with Ϫ5/del(5q) are characterized by complex karyotypes, a poor prognosis, and relative resistance to conventional therapies. 4,6,7 These features suggest that additional acquired genetic alterations occur in the advanced stages of MDS or t-MDS/t-AML, accelerating the progression of the disease.Two commonly deleted segments (CDSs) within 5q33.1 and 5q31 have been identified by cytogenetic analysis of MDS with an isolated del(5q) (5q33.1), or MDS, AML de novo, and t-MDS/t-AML with a del(5q) (5q31). 8,9 Thus far, biallelic deletions or inactivating mutations have been not reported in genes located in the 5q CDSs [8][9][10] The existing data support a haploinsufficiency model, in which loss of a single allele of 1 or more genes on 5q contributes to the pathogenesis of MDS or t-MDS/t-AML with a Ϫ5/del(5q). 10,11 A number of genes located on 5q, including RPS14, 12 EGR1, 13 NPM1, 14 and CTNNA1, 15 have been implicated in the development of myeloid disorders due to a gene dosage effect.The APC tumor suppressor gene is located at chromosome band 5q23, and loss of a single allele of APC occurs in more than 95% of patients with myeloid neoplasms and Ϫ5/del(5q). Loss of function of APC is responsible for the initiation and progression of colorectal cancer. 16 APC, a multifunctional protein, is involved in the regulation of the Wnt signaling pathway via its ability to control the degradation of ␤-catenin. 16 Other cellular processes in which APC plays a role include cell migration, cell adhesion, spindle assembly, and chromosome seg...

Mapping origins of replication has been challenging in higher eukaryotes. We have developed a rapid, genome-wide method to map origins of replication in asynchronous human cells by combining the nascent strand abundance assay with a highly tiled microarray platform, and we validated the technique by two independent assays. We applied this method to analyse the enrichment of nascent DNA in three 50-kb regions containing known origins of replication in the MYC, lamin B2 (LMNB2 ) and haemoglobin b (HBB ) genes, a 200-kb region containing the rare fragile site, FRAXA, and a 1,075-kb region on chromosome 22; we detected most of the known origins and also 28 new origins. Surprisingly, the 28 new origins were small in size and located predominantly within genes. Our study also showed a strong correlation between origin replication timing and chromatin acetylation.

The adenomatous polyposis coli (Apc) tumor suppressor is involved in the initiation and progression of colorectal cancer via regulation of the Wnt signaling cascade. In addition, Apc plays an important role in multiple cellular functions, including cell migration and adhesion, spindle assembly, and chromosome segregation. However, its role during adult hematopoiesis is unknown. We show that conditional inactivation of Apc in vivo dramatically increases apoptosis and enhances cell cycle entry of hematopoietic stem cells (HSCs)/ hematopoietic progenitor cells (HPCs), leading to their rapid disappearance and bone marrow failure. The defect in HSCs/HPCs caused by Apc ablation is cell autonomous. In addition, we found that loss of Apc leads to exhaustion of the myeloid progenitor pool (common myeloid progenitor, granulocytemonocyte progenitor, and megakaryocyte-erythroid progenitor), as well as the lymphoidprimed multipotent progenitor pool. Down-regulation of the genes encoding Cdkn1a, Cdkn1b, and Mcl1 occurs after acute Apc excision in candidate HSC populations. Together, our data demonstrate that Apc is essential for HSC and HPC maintenance and survival.

Mammalian chromosome ends contain long arrays of TTAGGG repeats that are complexed to a telomere specific protein, the TTAGGG repeat binding factor, TRF1. Here we describe the characterization of genes encoding the human and mouse TRF1 proteins, hTRF1 and mTRF1. The mTRF1 cDNA was isolated based on sequence similarity to the hTRF1 cDNA and the mTRF1 mRNA was shown to be ubiquitously expressed as a single 1.9 kb polyadenylated transcript in mouse somatic tissues. High levels of a 2.1 kb transcript were found in testes. In vitro translation of the mTRF1 cDNA resulted in a 56 kDa protein that binds to TTAGGG repeat arrays. mTRF1 displayed the same sequence specificity as hTRF1, preferring arrays of TTAGGG repeats as a binding substrate over TTAGGC and TTGGGG repeats. Expression of an epitope-tagged version of mTRF1 showed that the protein is located at the ends of murine metaphase chromosomes. In agreement, conceptual translation indicated that mTRF1 and hTRF1 are similarly-sized proteins with nearly identical C-terminal Myb-related DNA binding motifs. In addition, comparison of the predicted mTRF1 and hTRF1 amino acid sequences showed that the acidic nature of the N-terminus of TRF1 is conserved and revealed a highly conserved novel domain of approximately 200 amino acids in the middle of the proteins. However, other regions of the proteins are poorly conserved (<35% identity) and the overall level of identity of the mTRF1 and hTRF1 amino acid sequences is only 67%. The TRF1 genes are not syntenic; the hTRF1 gene localized to human chromosome 8 band q13 while the mTRF1 gene localized to mouse chromosome 17 band E3. The data indicate that the genes for mammalian telomeric proteins evolve rapidly.

• Egr1 haploinsufficiency in cooperation with reduced Tp53 activity accelerates the development of hematologic disease in mice.• Loss of 1 copy of Egr1 and Apc in hematopoietic stem cells, in cooperation with Tp53 loss, results in myeloid neoplasms.An interstitial deletion of chromosome 5, del(5q), is the most common structural abnormality in primary myelodysplastic syndromes (MDS) and therapy-related myeloid neoplasms (t-MNs) after cytotoxic therapy. Loss of TP53 activity, through mutation or deletion, is highly associated with t-MNs with a del(5q). We previously demonstrated that haploinsufficiency of Egr1 and Apc, 2 genes lost in the 5q deletion, are key players in the progression of MDS with a del(5q). Using genetically engineered mice, we now show that reduction or loss of Tp53 expression, in combination with Egr1 haploinsufficiency, increased the rate of development of hematologic neoplasms and influenced the disease spectrum, but did not lead to overt myeloid leukemia, suggesting that altered function of additional gene(s) on 5q are likely required for myeloid leukemia development. Next, we demonstrated that cell intrinsic loss of Tp53 in hematopoietic stem and progenitor cells haploinsufficient for both Egr1 and Apc led to the development of acute myeloid leukemia (AML) in 17% of mice. The long latency (234-299 days) and clonal chromosomal abnormalities in the AMLs suggest that additional genetic changes may be required for full transformation. Thus, loss of Tp53 activity in cooperation with Egr1 and Apc haploinsufficiency creates an environment that is permissive for malignant transformation and the development of AML. (Blood. 2014;123(7):1069-1078)