Human uterine leiomyomas (ULMs) are the most common neoplasms of women. Many genes are dysregulated in ULMs and some of this dysregulation may be due to abnormal expression of micro-RNAs (miRNAs). In this study, 55 ULMs and matched myometrium were collected from 41 patients for microarray-based global miRNA expression analysis. Of 206 miRNAs examined, 45 miRNAs were significantly up- or down-regulated in ULMs in comparison to the matched myometrium (P < 0.001). The top five dysregulated miRNAs in ULMs are the let-7 family, miR-21, miR-23b, miR-29b, and miR-197. Four polycistronic clusters of miRNAs were either up- or down-regulated, but not in a mixed pattern, indicative of coordinated regulation of these miRNAs. Significance analysis revealed that subsets of miRNAs were strongly associated with tumor sizes and race. By prediction analysis we identified some important tumorigenic genes previously identified in ULMs that may be targeted by the dysregulated miRNAs. HMGA2 was identified as one of target genes of the let-7 family of miRNAs and has been found to be suppressed by let-7 in vitro. This article contains Supplementary material available at http://www.interscience.wiley.com/jpages/1045-2257/suppmat.

Studies of recombination-dependent replication (RDR) in the T4system have revealed the critical roles played by mediator proteins in the timely and productive loading of specific enzymes onto single-stranded DNA (ssDNA) during phage RDR processes. The T4 recombination mediator protein, uvsY, is necessary for the proper assembly of the T4 presynaptic filament (uvsX recombinase cooperatively bound to ssDNA), leading to the recombination-primed initiation of leading strand DNA synthesis. In the lagging strand synthesis component of RDR, replication mediator protein gp59 is required for the assembly of gp41, the DNA helicase component of the T4 primosome, onto lagging strand ssDNA. Together, uvsY and gp59 mediate the productive coupling of homologous recombination events to the initiation of T4 RDR. UvsY promotes presynaptic filament formation on 3 ssDNA-tailed chromosomes, the physiological primers for T4 RDR, and recent results suggest that uvsY also may serve as a coupling factor between presynapsis and the nucleolytic resection of double-stranded DNA ends. Other results indicate that uvsY stabilizes uvsX bound to the invading strand, effectively preventing primosome assembly there. Instead, gp59 directs primosome assembly to the displaced strand of the D loop͞replication fork. This partitioning mechanism enforced by the T4 recombination͞replication mediator proteins guards against antirecombination activity of the helicase component and ensures that recombination intermediates formed by uvsX͞uvsY will efficiently be converted into semiconservative DNA replication forks. Although the major mode of T4 RDR is semiconservative, we present biochemical evidence that a conservative ''bubble migration'' mode of RDR could play a role in lesion bypass by the T4 replication machinery. Bacteriophage T4 provides an excellent model system for biochemical and genetic studies of recombinationdependent replication (RDR), because DNA replication and recombination are closely coupled throughout much of the phage life cycle. After infecting a host Escherichia coli cell, T4 first replicates its genome via an origin-dependent replication initiation pathway. This pathway is shut off after a few rounds of replication, after expression of the T4 uvsW RNA͞DNA helicase, which resolves R loops required for origin function (1). T4 then relies on a recombination-dependent mechanism to initiate DNA synthesis, and this pathway accounts for a large fraction of the total DNA synthesis observed during T4 infection. In the T4 RDR pathway (reviewed in refs. 2 and 3), branched recombination intermediates generated by the phage homologous recombination machinery are captured and converted into semiconservative DNA replication forks. T4 RDR requires all of the major phage-encoded DNA replication and recombination enzymes including: gp43 (DNA polymerase), gp45 (sliding clamp), gp44͞62 (clamp loader), gp32 [single-stranded DNA (ssDNA) binding protein or ssb], gp61 (primase), gp41 (DNA helicase), gp59 (helicase loader; replication mediator protein or RMP...

Semi-conservative DNA synthesis reactions catalyzed by the bacteriophage T4 DNA polymerase holoenzyme are initiated by a strand displacement mechanism requiring gp32, the T4 single-stranded DNA (ssDNA)-binding protein, to sequester the displaced strand. After initiation, DNA helicase acquisition by the nascent replication fork leads to a dramatic increase in the rate and processivity of leading strand DNA synthesis. In vitro studies have established that either of two T4-encoded DNA helicases, gp41 or dda, is capable of stimulating strand displacement synthesis. The acquisition of either helicase by the nascent replication fork is modulated by other protein components of the fork including gp32 and, in the case of the gp41 helicase, its mediator/ loading protein gp59. Here, we examine the relationships between gp32 and the gp41/gp59 and dda helicase systems, respectively, during T4 replication using altered forms of gp32 defective in either protein-protein or protein-ssDNA interactions. We show that optimal stimulation of DNA synthesis by gp41/gp59 helicase requires gp32-gp59 interactions and is strongly dependent on the stability of ssDNA binding by gp32. Fluorescence assays demonstrate that gp59 binds stoichiometrically to forked DNA molecules; however, gp59-forked DNA complexes are destabilized via protein-protein interactions with the C-terminal "A-domain" fragment of gp32. These and previously published results suggest a model in which a mobile gp59-gp32 cluster bound to lagging strand ssDNA is the target for gp41 helicase assembly. In contrast, stimulation of DNA synthesis by dda helicase requires direct gp32-dda protein-protein interactions and is relatively unaffected by mutations in gp32 that destabilize its ssDNA binding activity. The latter data support a model in which protein-protein interactions with gp32 maintain dda in a proper active state for translocation at the replication fork. The relationship between dda and gp32 proteins in T4 replication appears similar to the relationship observed between the UL9 helicase and ICP8 ssDNA-binding protein in herpesvirus replication.DNA helicase acquisition by a nascent DNA replication fork is essential for reconstituting rapid, processive DNA synthesis. This principle is illustrated dramatically by the bacteriophage T4 DNA replication system (1-2). T4 encodes two DNA helicases known to affect the movement and properties of DNA replication forks; the gene 41 protein (gp41) and the dda protein, respectively. gp41 is the essential replicative helicase of the T4 phage. This hexameric enzyme translocates processively in a 5Ј 3 3Ј direction on the displaced lagging strand of the dsDNA 1 template, thus enhancing the rate and processivity of leading strand DNA synthesis catalyzed by the T4 DNA polymerase holoenzyme (gp43, gp44/62, and gp45 proteins) (1-3). In addition, gp41 is an obligatory component of the T4 primosome (helicase/primase complex) and is, thus, essential for lagging strand DNA synthesis (4 -5). A second T4-encoded DNA helicase, dda protein, stimula...

Cilia are microscopic hair-like external cell organelles that are ubiquitously present in nature, also within the human body. They fulfill crucial biological functions: motile cilia provide transportation of fluids and...

Colorectal cancer (CRC) remains a major cause of carcinoma‐related deaths worldwide. MicroRNA‐498 (miR‐498) modulates the development of a variety of biological events, including tumorigenesis. Nevertheless, it is unclear whether miR‐498 plays a role in CRC. This study was designed to elucidate the underlying mechanism and role of miR‐498 in modulation of the viability and invasiveness of CRC cells. We report that CRC tissues and cells exhibited decreased expression of miR‐498, and that overexpression of miR‐498 resulted in reduced proliferation of CRC cells, concomitant with increased apoptosis. Furthermore, bioinformatic prediction and dual‐luciferase reporter assay revealed that miR‐498 targeted the 3′‐UTR of Bcl‐2 for silencing. However, Bcl‐2 overexpression suppressed the proapoptosis of miR‐498 on CRC cells. In summary, we describe a possible role of miR‐498 in CRC, which may lead to the identification of new targets for treatment of this malignancy.

To investigate the clinical characteristics, survival, prognostic factors, and treatment of brain metastasis (BM) from colorectal cancer (CRC). Twenty-one patients with BM from CRC were retrospectively reviewed. Predictive factors for BM and prognostic factors after the diagnosis of BM were examined by univariate and multivariate COX analysis. The time from the development of extracranial metastases, including lung, bone, and liver, to the occurrence of BM was recorded separately. The median overall survival time was 7 months. In univariate prognostic analysis, median survival with multimodal therapy was better than that with unimodal therapy (10 months vs 3 months, P = .000). In addition, median survival with Karnofsky performance status (KPS) < 70, 1 BM lesion, primary tumor stage of II-III, extracranial lesions < 2, and no extracranial metastasis were much better than the other groups (P < .05 of all). Although there was not a significant difference in median survival between patients receiving combination treatment with bevacizumab and those who did not, treatment with bevacizumab was associated with better survival (10 months vs 5 months, P = .436). The time intervals from bone, liver, and lung metastases to BM were 3, 6.5, and 11 months, respectively. Based on multivariate Cox analysis, KPS and treatment modalities were independent prognosis factors (P = .039 and P = .000, respectively). CRC patients with a high KPS and multimodal treatment have improved survival.Abbreviations: BM = brain metastasis, CI = confidence interval, CRC = colorectal cancer, KPS = Karnofskyperformance status, KRAS = Kirsten rat sarcoma viral oncogene homolog, OS = overall survival, RAS = rat sarcoma viral oncogene homolog, WBRT = whole-brain radiotherapy.

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