In Saccharomyces cerevisiae chromosomal DNA replication initiates at intervals of ϳ40 kb and depends upon the activity of autonomously replicating sequence (ARS) elements. The identification of ARS elements and analysis of their function as chromosomal replication origins requires the use of functional assays because they are not sufficiently similar to identify by DNA sequence analysis. To complete the systematic identification of ARS elements on S. cerevisiae chromosome III, overlapping clones covering 140 kb of the right arm were tested for their ability to promote extrachromosomal maintenance of plasmids. Examination of chromosomal replication intermediates of each of the seven ARS elements identified revealed that their efficiencies of use as chromosomal replication origins varied widely, with four ARS elements active in Յ10% of cells in the population and two ARS elements active in Ն90% of the population. Together with our previous analysis of a 200-kb region of chromosome III, these data provide the first complete analysis of ARS elements and DNA replication origins on an entire eukaryotic chromosome.
INTRODUCTIONThe replication of eukaryotic chromosomes initiates at multiple replication origins spaced at intervals of 40 -100 kb. In the budding yeast, Saccharomyces cerevisiae, replication origins depend upon cis-acting replicators called autonomously replicating sequence (ARS) elements, which are recognized by their ability to maintain extrachromosomal plasmids. The initiation of replication at individual replicators is a tightly regulated process. Replication initiations are confined to S phase, and individual replicators initiate at reproducible times during S phase, some early and some late (Reynolds et al., 1989, and references therein;Friedman et al., 1997;Donaldson et al., 1998b). Moreover, the efficiencies of initiation vary from one replicator to another, the extreme case being ARS elements that are not active as replication origins in their normal chromosomal positions (Dubey et al., 1991;Friedman et al., 1997;Yamashita et al., 1997).Replicators become competent to initiate replication during the G1 phase of the cell cycle through the stepwise assembly of prereplicative complexes on replicators that are bound by origin recognition complex (ORC) (reviewed by Dutta and Bell, 1997). The actual initiation events require the activities of at least two protein kinases, the cyclin-dependent kinase (CDK) Cdc28p associated with cyclin B (Clb5p or Clb6p) and the Cdc7p kinase associated with its regulatory subunit Dbf4p. The assembly of prereplicative complexes is prevented by the activity of cyclin B-associated CDK, effectively preventing reinitiation at origins during a single S phase (reviewed by Diffley, 1996). Timing determinants also appear to be specified during G1 of the cell cycle (Raghuraman et al., 1997), although the relationship between establishment of the prereplication complex and the specification of initiation timing is unclear. In the case of a latereplicating region of chromosome XIV, DN...
Cell-based fluorescence assays in leukemia/lymphoma evaluations typically rely on qualitative approaches for the identification and enumeration of the target cell population(s), but other equally important diagnostic assays are quantitative or semiquantitative. Qualitative assays usually give an overall picture of the composite phenotype based on the expression level of a set of antigens on particular cell lineages that render diagnostic patterns. Conversely, quantitative flow cytometry precisely measures the antigen density or absolute target cell count, and semi-quantitative assays quantify the abnormal target cell population above a certain threshold relative to its normal counterpart or total cells. The following sections attempt to provide an overview of the different types of flow cytometric evaluation of normal and pathological specimens, providing details of
Anaplastic large cell lymphoma (ALCL) is a distinct type of T/null-cell non-Hodgkin lymphoma that commonly involves nodal and extranodal sites. The World Health Organization of lymphoid neoplasms recognizes two types: anaplastic lymphoma kinase (ALK) positive or ALK negative, the former as a result of abnormalities involving the ALK gene at chromosome 2p23. Patients with ALCL rarely develop a leukemic phase of disease, either at the time of initial presentation or during the clinical course. Described herein is a patient with ALK+ ALCL, small cell variant, associated with the t(2;5)(p23;q35), who initially presented with leukemic involvement and an extraordinarily high leukocyte count of 529 x 10(9)/L, which subsequently peaked at 587 x 10(9)/L. Despite chemotherapy the patient died 2(1/2) months after diagnosis. In the literature review 20 well-documented cases are identified of ALCL in leukemic phase reported previously, with a WBC ranging from 15 to 151 x 10(9)/L. Leukemic phase of ALCL occurs almost exclusively in patients with ALK+ ALCL, most often associated with the small cell variant and the t(2;5)(p23;q35), similar to the present case. Patients with leukemic phase ALK+ ALCL appear to have a poorer prognosis than most patients with ALK+ ALCL.
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