Chromosome 19 has the highest gene density of all human chromosomes, more than double the genome-wide average. The large clustered gene families, corresponding high G1C content, CpG islands and density of repetitive DNA indicate a chromosome rich in biological and evolutionary significance. Here we describe 55.8 million base pairs of highly accurate finished sequence representing 99.9% of the euchromatin portion of the chromosome. Manual curation of gene loci reveals 1,461 protein-coding genes and 321 pseudogenes. Among these are genes directly implicated in mendelian disorders, including familial hypercholesterolaemia and insulin-resistant diabetes. Nearly one-quarter of these genes belong to tandemly arranged families, encompassing more than 25% of the chromosome. Comparative analyses show a fascinating picture of conservation and divergence, revealing large blocks of gene orthology with rodents, scattered regions with more recent gene family expansions and deletions, and segments of coding and non-coding conservation with the distant fish species Takifugu.

DPBII, a gene that suppresses mutations in two essential subunits of Saccharomyces cerevisiae DNA polymerase II(E) encoded by POL2 and DPB2, was isolated on a multicopy plasmid. The nucleotide sequence of the DPBII gene revealed an open reading frame predicting an 87-kDa protein. This protein is homologous to the Schizosaccharomyces pombe rad4+/cut5+ gene product that has a cell cycle checkpoint function. Disruption ofDPBII is lethal, indicating that DPBII is essential for cell proliferation. In thermosensitive dpbll-1 mutant cells, S-phase progression is defective at the nonpermissive temperature, followed by cell division with unequal chromosomal segregation accompanied by loss of viability. dpbll-1 is synthetic lethal with any one of the dpb2-1, pol2-11, and pol2-18 mutations at all temperatures. Moreover, dpbll cells are sensitive to hydroxyurea, methyl methanesulfonate, and UV irradiation. These results strongly suggest that Dpbll is a part of the DNA polymerase II complex during chromosomal DNA replication and also acts in a checkpoint pathwaj during the S phase of the cell cycle to sense stalled DNA replication.Chromosomal DNA is accurately replicated only once in S phase of the cell cycle. In Saccharomyces cerevisiae, if DNA replication is blocked or DNA is damaged by reagents or aberrant DNA synthesis, a checkpoint system arrests the cell cycle (for review, see refs. 1 and 2). The DNA damage checkpoint requires the RAD9, RAD17, RAD24, MEC1, MEC2/ RAD53/SADl/SPKI, and MEC3 genes (3-6). The S-phase checkpoint that finds the replication block partially overlaps the DNA damage checkpoint, as MEC1 and MEC2 are required for both checkpoints (5,6).A checkpoint at the S-phase onset has been also described in the fission yeast Schizosaccharomycespombe (for review, see ref. 7). cdc18, rad4/cut5, and cdtl mutations prevent initiation of DNA synthesis and allow cells to enter mitosis before completion of DNA replication (8-11). In Saccharomyces cerevisiae, Cdc6, the probable homolog of Schizosaccharomyces pombe Cdc18, was reported to have a checkpoint function, acting positively at the initiation of DNA replication and negatively at the entry to mitosis (12). However, homologs of fission yeast Cut5 and Cdtl had not been found in Saccharomyces cerevisiae.Though several checkpoint mutations and the corresponding genes have been isolated, the mechanism sensing the actual DNA damage or replication block remains obscure. Recently, Navas et al. (13) reported that Pol2, the catalytic subunit of Saccharomyces cerevisiae DNA polymerase II, is important for the S-phase checkpoint and they proposed that DNA polymerase II acts as a sensor of DNA replication. In Saccharomyces cerevisiae, DNA polymerase II(e) and two other DNA polymerases, I(a) and III(8), are essential for chromosomalThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.DNA replication (for review, se...

Acute myelogenous leukemia (AML) is a heterogeneous disorder characterized by clonal proliferation of stem cell-like blasts in bone marrow (BM); however, their unique cellular interaction within the BM microenvironment and its functional significance remain unclear. Here, we assessed the BM microenvironment of AML patients and demonstrate that the leukemia stem cells induce a change in the transcriptional programming of the normal mesenchymal stromal cells (MSC). The modified leukemic niche alters the expressions of cross-talk molecules (i.e., CXCL12 and JAG1) in MSCs to provide a distinct cross-talk between normal and leukemia cells, selectively suppressing normal primitive hematopoietic cells while supporting leukemogenesis and chemoresistance. Of note, AML patients exhibited distinct heterogeneity in the alteration of mesenchymal stroma in BM. The distinct pattern of stromal changes in leukemic BM at initial diagnosis was associated with a heterogeneous posttreatment clinical course with respect to the maintenance of complete remission for 5 to 8 years and early or late relapse. Thus, remodeling of mesenchymal niche by leukemia cells is an intrinsic self-reinforcing process of leukemogenesis that can be a parameter for the heterogeneity in the clinical course of leukemia and hence serve as a potential prognostic factor. Cancer Res; 75(11); 2222-31. Ó2015 AACR.

In vivo expression of human telomerase is significantly different from that of mouse telomerase. To assess the basis for this difference, a bacterial artificial chromosome clone containing the entire hTERT (human telomerase reverse transcriptase) gene was introduced in mice. In these transgenic mice, expression of the hTERT transgene was similar to that of endogenous hTERT in humans, rather than endogenous mTERT (mouse telomerase reverse transcriptase). In tissues and cells showing a striking difference in expression levels between hTERT in humans and mTERT in mice (i.e., liver, kidney, lung, uterus, and fibroblasts), expression of the hTERT transgene in transgenic mice was repressed, mimicking hTERT in humans. The transcriptional activity of the hTERT promoter was much lower than that of the mTERT promoter in mouse embryonic fibroblasts or human fibroblasts. Mutational analysis of the hTERT and mTERT promoters revealed that a nonconserved GC-box within the hTERT promoter was responsible for the humanspecific repression. These results reveal that a difference in cisregulation of transcription, rather than transacting transcription factors, is critical to species differences in tissue-specific TERT expression. Our data also suggest that the GC-box-mediated, human-specific mechanism for TERT repression is impaired in human cancers. This study represents a detailed characterization of the functional difference in a gene promoter of mice versus humans and provides not only important insight into speciesspecific regulation of telomerase and telomeres but also an experimental basis for generating mice humanized for telomerase enzyme and its pattern of expression.

With contrasting observations on the effects of b-catenin on hematopoietic stem cells (HSCs), the precise role of Wnt/b-catenin signals on HSC regulation remains unclear. Here, we show a distinct mode of Wnt/b-catenin signal that can regulate HSCs in a stroma-dependent manner. Stabilization of b-catenin in the bone marrow stromal cells promoted maintenance and self-renewal of HSCs in a contactdependent manner, whereas direct stabilization in hematopoietic cells caused loss of HSCs. Interestingly, canonical Wnt receptors and b-catenin accumulation were predominantly enriched in the stromal rather than the hematopoi-

Background: Although various molecular subtypes of bladder cancer (BC) have been investigated, most of these studies have focused on muscle-invasive BC (MIBC). A few studies have investigated non-muscle-invasive BC (NMIBC) or NMIBC and MIBC together, but none has classified progressive NMIBC or immune checkpoint inhibitor (ICI)-based therapeutic responses in early-stage BC patients. Methods: A total of 1,934 samples from seven patient cohorts were used. We performed unsupervised hierarchical clustering to stratify patients into distinct subgroups and constructed a classifier by applying SAM/ PAM algorithms. We then investigated the association between molecular subtypes and immunotherapy responsiveness using various statistical methods. Findings: We explored large-scale genomic datasets encompassing NMIBC and MIBC, redefining four distinct molecular subtypes, including a subgroup containing progressive NMIBC and MIBC with poor prognosis that would benefit from ICI treatment. This subgroup showed poor progression-free survival with the distinct features of high mutation load, activated cell cycle, and inhibited TGFb signalling. Importantly, we verified that BC patients with this subtype were significantly responsive to an anti-PD-L1 agent in the IMvigor210 cohort. Interpretation: Our results reveal an immunotherapeutic option for ICI treatment of highly progressive NMIBC and MIBC with poor prognosis.

Lipocalin-2 (Lcn2) is preferentially expressed in hepatocellular carcinoma (HCC). However, the functional role of Lcn2 in HCC progression is still poorly understood, particularly with respect to its involvement in invasion and metastasis. The purpose of this study was to investigate whether Lcn2 is associated with the epithelial-mesenchymal transition (EMT) in HCC and to elucidate the underlying signaling pathway(s). Lcn2 was preferentially expressed in well-differentiated HCC versus liver cirrhosis tissues, and its expression was positively correlated with the stage of HCC. The characteristics of EMT were reversed by adenoviral transduction of Lcn2 into SH-J1 cells, including the down-regulation of N-cadherin, vimentin, alpha-smooth muscle actin, and fibronectin, and the concomitant up-regulation of CK8, CK18, and desmoplakin I/II. Knockdown of Lcn2 by short hairpin RNA (shRNA) in HKK-2 cells expressing high levels of Lcn2 was associated with EMT. Epidermal growth factor (EGF) or transforming growth factor beta1 (TGF-b1) treatment resulted in down-regulation of Lcn2, accompanied by an increase in Twist1 expression and EMT in HCC cells. Stable Lcn2 expression in SH-J1 cells reduced Twist1 expression, inhibited cell proliferation and invasion in vitro, and suppressed tumor growth and metastasis in a mouse model. Furthermore, EGF or TGFb1 treatment barely changed EMT marker expression in SH-J1 cells ectopically expressing Lcn2. Ectopic expression of Twist1 induced EMT marker expression even in cells expressing Lcn2, indicating that Lcn2 functions downstream of growth factors and upstream of Twist1. Conclusion: Together, our findings indicate that Lcn2 can negatively modulate the EMT in HCC cells through an EGF (or TGF-b1)/Lcn2/Twist1 pathway. Thus, Lcn2 may be a candidate metastasis suppressor and a potential therapeutic target in HCC. (HEPATOLOGY 2013;58:1349-1361 L ipocalin-2 (Lcn2), also known as NGAL, belongs to the lipocalin protein family and was first purified from human neutrophils because of its association with gelatinase. 1 Lcn2 can exist as a 25-kDa monomer, 46-kDa disulfide-linked homodimer, and/or 135-kDa disulfide-linked heterodimer with neutrophil gelatinase. 2 Elevated Lcn2 expression has been observed in multiple human cancers including

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