Integrated expression profiling of multiple RNA species by real-time PCR.

MicroRNAs (miRNAs) are endogenous, non-coding RNAs comprising approximately 21-23 nucleotides that regulate gene expression by binding to and targeting messenger RNA (mRNA) for translational repression or degradation. miRNAs have been shown to regulate cellular processes including proliferation, differentiation, and development and to play an important role in immune system function. The expression of miRNAs is misregulated in numerous diseases, including cancers of immunological origin.

Cdc45 limits replicon usage from a low density of preRCs in mammalian cells.

Little is known about mammalian preRC stoichiometry, the number of preRCs on chromosomes, and how this relates to replicon size and usage. We show here that, on average, each 100-kb of the mammalian genome contains a preRC composed of approximately one ORC hexamer, 4-5 MCM hexamers, and 2 Cdc6. Relative to these subunits, ∼0.

Cell cycle arrest by transforming growth factor beta1 near G1/S is mediated by acute abrogation of prereplication complex activation involving an Rb-MCM interaction.

Understanding inhibitory mechanisms of transforming growth factor beta1 (TGF-beta1) has provided insight into cell cycle regulation and how TGF-beta1 sensitivity is lost during tumorigenesis. We show here that TGF-beta1 utilizes a previously unknown mechanism targeting the function of prereplication complexes (pre-RCs) to acutely block S-phase entry when added to cells in late G(1), after most G(1) events have occurred. TGF-beta1 treatment in early G(1) suppresses Myc and CycE-Cdk2 and blocks pre-RC assembly.

Mammalian MCM loading in late-G(1) coincides with Rb hyperphosphorylation and the transition to post-transcriptional control of progression into S-phase.

Background: Control of the onset of DNA synthesis in mammalian cells requires the coordinated assembly and activation of the pre-Replication Complex. In order to understand the regulatory events controlling preRC dynamics, we have investigated how the timing of preRC assembly relates temporally to other biochemical events governing progress into S-phase.

Methodology/principal Finding: In murine and Chinese hamster (CHO) cells released from quiescence, the loading of the replicative MCM helicase onto chromatin occurs in the final 3-4 hrs of G(1).

In vivo chromatin decondensation assays: molecular genetic analysis of chromatin unfolding characteristics of selected proteins.

Of critical importance to many of the events underlying transcriptional control of gene expression are modifications to core and linker histones that regulate the accessibility of trans-acting factors to the DNA substrate within the context of chromatin. Likewise, control over the initiation of DNA replication, as well as the ability of the replication machinery to proceed during elongation through the multiple levels of chromatin condensation that are likely to be encountered, is almost certain to involve the creation of chromatin accessibility. In the latter case in particular, chromatin access will likely need to be a transient event so as to prevent total genomic unraveling of the chromatin that would be deleterious to cells.

Expression of the circadian clock genes Per1 and Per2 in sporadic and familial breast tumors.

There is a growing body of evidence implicating aberrant circadian clock expression in the development of cancer. Based on our initial experiments identifying a putative interaction between BRCA1 and the clock proteins Per1 and Per2, as well as the reported involvement of the circadian clock in the development of cancer, we have performed an expression analysis of the circadian clock genes Per1 and Per2 in both sporadic and familial primary breast tumors and normal breast tissues using real-time polymerase chain reaction. Significantly decreased levels of Per1 were observed between sporadic tumors and normal samples (P < .

The interaction of PP1 with BRCA1 and analysis of their expression in breast tumors.

Background: The breast cancer susceptibility gene, BRCA1, is implicated in multiple cellular processes including DNA repair, the transactivation of genes, and the ubiquitination of proteins; however its precise functions remain to be fully understood. Identification and characterization of BRCA1 protein interactions may help to further elucidate the function and regulation of BRCA1. Additionally, detection of changes in the expression levels of BRCA1 and its interacting proteins in primary human breast tumors may further illuminate their role in the development of breast cancer.