The BRCA1 tumor suppressor binds to inositol 1,4,5-trisphosphate receptors to stimulate apoptotic calcium release.

The inositol 1,4,5-trisphosphate receptor (IP3R) is a ubiquitously expressed endoplasmic reticulum (ER)-resident calcium channel. Calcium release mediated by IP3Rs influences many signaling pathways, including those regulating apoptosis. IP3R activity is regulated by protein-protein interactions, including binding to proto-oncogenes and tumor suppressors to regulate cell death.

The ubiquitin ligases RNF8 and RNF168 display rapid but distinct dynamics at DNA repair foci in living cells.

Rapid assembly of DNA damage response (DDR) proteins at nuclear "repair" foci is a hallmark response of ionizing radiation (IR)-treated cells. The ubiquitin E3 ligases RNF8 and RNF168 are critical for foci formation, and here we aim to determine their dynamic mobility and abundance at individual foci in living cells. To this end, YFP-tagged RNF8 and RNF168 were expressed at physiological levels in MCF-7 cells, then analyzed by fluorescence recovery after photobleaching (FRAP) assays, nuclear retention measurement, and virus-like particles (VLPs)-based quantification.

Targeting the DNA replication checkpoint by pharmacologic inhibition of Chk1 kinase: a strategy to sensitize APC mutant colon cancer cells to 5-fluorouracil chemotherapy.

5-fluorouracil (5-FU) is the first line component used in colorectal cancer (CRC) therapy however even in combination with other chemotherapeutic drugs recurrence is common. Mutations of the adenomatous polyposis coli (APC) gene are considered as the initiating step of transformation in familial and sporadic CRCs. We have previously shown that APC regulates the cellular response to DNA replication stress and recently hypothesized that APC mutations might therefore influence 5-FU resistance.

Neural processing of facial expressions of emotion in first onset psychosis.

Schizophrenia is characterized by deficits in face and facial emotion processing. This is the first study using event-related potentials (ERPs) to investigate the corresponding neural activation in first onset psychosis. ERPs for 108 first onset psychosis participants and 108 matched healthy controls were recorded while they viewed facial expressions.

Targeting the β-catenin nuclear transport pathway in cancer.

The nuclear localization of specific proteins is critical for cellular processes such as cell division, and in recent years perturbation of the nuclear transport cycle of key proteins has been linked to cancer. In particular, specific gene mutations can alter nuclear transport of tumor suppressing and oncogenic proteins, leading to cell transformation or cancer progression. This review will focus on one such factor, β-catenin, a key mediator of the canonical wnt signaling pathway.

Wnt signaling proteins associate with the nuclear pore complex: implications for cancer.

Several components of the Wnt signaling pathway have in recent years been linked to the nuclear pore complex. β-catenin, the primary transducer of Wnt signals from the plasma membrane to the nucleus, has been shown to transiently associate with different FG-repeat containing nucleoporins (Nups) and to translocate bidirectionally through pores of the nuclear envelope in a manner independent of classical transport receptors and the Ran GTPase. Two key regulators of β-catenin, IQGAP1 and APC, have also been reported to bind specific Nups or to locate at the nuclear pore complex.

The scaffolding protein IQGAP1 co-localizes with actin at the cytoplasmic face of the nuclear envelope: implications for cytoskeletal regulation.

IQGAP1 is an important cytoskeletal regulator, known to act at the plasma membrane to bundle and cap actin filaments, and to tether the cortical actin meshwork to microtubules via plus-end binding proteins. Here we describe the novel subcellular localization of IQGAP1 at the cytoplasmic face of the nuclear envelope, where it co-located with F-actin. The IQGAP1 and F-actin staining overlapped that of microtubules at the nuclear envelope, revealing a pattern strikingly similar to that observed at the plasma membrane.

The in vivo dynamic interplay of MDC1 and 53BP1 at DNA damage-induced nuclear foci.

MDC1 (NFBD1) and 53BP1 are critical mediators of the mammalian DNA damage response (DDR) at nuclear foci. Here we show by quantitative imaging assays that MDC1 and 53BP1 are similar in total copy number (~1200 copies per focus), but differ substantially in dynamics at both replication-associated nuclear bodies in normal cells and DNA repair foci in ionizing radiation (IR)-damaged cells. The majority of MDC1 (~80%) is extremely mobile and under continuous exchange, with only a small fraction (~20%) remaining immobile at foci irrespective of IR treatment.

Three-dimensional imaging reveals the spatial separation of γH2AX-MDC1-53BP1 and RNF8-RNF168-BRCA1-A complexes at ionizing radiation-induced foci.

Background And Purpose: Ionizing radiation (IR)-induced DNA damage causes the accumulation of DNA damage response (DDR) proteins as visible foci in cell nuclei. Despite the identified functional roles in DNA repair, the spatial relationships of different DDR proteins at foci have not been explicitly examined. This study aims to systematically compare the distribution of DDR proteins at IR-induced foci.

Wnt signaling from membrane to nucleus: β-catenin caught in a loop.

β-catenin is the central nuclear effector of the Wnt signaling pathway, and regulates other cellular processes including cell adhesion. Wnt stimulation of cells culminates in the nuclear translocation of β-catenin and transcriptional activation of target genes that function during both normal and malignant development. Constitutive activation of the Wnt pathway leads to inappropriate nuclear accumulation of β-catenin and gene transactivation, an important step in cancer progression.

The in vivo dynamic organization of BRCA1-A complex proteins at DNA damage-induced nuclear foci.

The breast cancer associated gene 1 (BRCA1)-A protein complex assembles at DNA damage-induced nuclear foci to facilitate repair of double-stranded breaks. Here, we describe the first systematic comparison of the dynamics, copy number and organization of its core components at foci. We show that the protein pools at individual foci generally comprise a small immobile fraction (∼20%) and larger mobile fraction (∼80%), which together occupy the same focal space but exist at different densities.

The BRCA1 Breast Cancer Suppressor: Regulation of Transport, Dynamics, and Function at Multiple Subcellular Locations.

Inherited mutations in the BRCA1 gene predispose to a higher risk of breast/ovarian cancer. The BRCA1 tumor suppressor is a 1863 amino acid protein with multiple protein interaction domains that facilitate its roles in regulating DNA repair and maintenance, cell cycle progression, transcription, and cell survival/apoptosis. BRCA1 was first identified as a nuclear phosphoprotein, but has since been shown to contain different transport sequences including nuclear export and nuclear localization signals that enable it to shuttle between specific sites within the nucleus and cytoplasm, including DNA repair foci, centrosomes, and mitochondria.

APC as a mobile scaffold: regulation and function at the nucleus, centrosomes, and mitochondria.

Genetic mutations of adenomatous polyposis coli (APC) predispose to high risk of human colon cancer. APC is a large tumor suppressor protein and truncating mutations disrupt its normal roles in regulating cell migration, DNA replication/repair, mitosis, apoptosis, and turnover of oncogenic β-catenin. APC is targeted to multiple subcellular sites, and here we discuss recent evidence implicating novel protein interactions and functions of APC in the nucleus and at centrosomes and mitochondria.

Characterization of BARD1 targeting and dynamics at the centrosome: the role of CRM1, BRCA1 and the Q564H mutation.

BARD1 heterodimerizes with BRCA1, forming an E3 ubiquitin ligase that functions at nuclear foci to repair DNA damage and the centrosome to regulate mitosis. We compared BARD1 recruitment at these structures using fluorescence recovery after photobleaching assays to measure YFP-BARD1 dynamics in live cells. In nuclei at ionizing radiation-induced foci, 20% of the BARD1 pool was immobile and 80% of slow mobility exhibiting a recovery time >500 s.

Adenomatous polyposis coli protein regulates the cellular response to DNA replication stress.

The adenomatous polyposis coli (APC) tumor suppressor traffics between nucleus and cytoplasm to perform distinct functions. Here we identify a specific role for APC in the DNA replication stress response. The silencing of APC caused an accumulation of asynchronous cells in early S phase and delayed S phase progression in cells released from hydroxyurea-mediated replication arrest.

Regulation of β-catenin nuclear dynamics by GSK-3β involves a LEF-1 positive feedback loop.

Nuclear localization of β-catenin is integral to its role in Wnt signaling and cancer. Cellular stimulation by Wnt or lithium chloride (LiCl) inactivates glycogen synthase kinase-3β (GSK-3β), causing nuclear accumulation of β-catenin and transactivation of genes that transform cells. β-catenin is a shuttling protein; however, the mechanism by which GSK-3β regulates β-catenin nuclear dynamics is poorly understood.

Methods to measure nuclear export of beta-catenin using fixed and live cell assays.

beta-Catenin is a multifunctional protein which is overexpressed in several types of cancers. The subcellular location of beta-catenin at the membrane junctions or in the nucleus determines its function in cell adhesion and transcription activation, respectively. The aberrant localization of beta-catenin in the nucleus activates transcription of a number of genes involved in cancer progression.

Nuclear export and centrosome targeting of the protein phosphatase 2A subunit B56alpha: role of B56alpha in nuclear export of the catalytic subunit.

Protein phosphatase (PP) 2A is a heterotrimeric enzyme regulated by specific subunits. The B56 (or B'/PR61/PPP2R5) class of B-subunits direct PP2A or its substrates to different cellular locations, and the B56alpha, -beta, and -epsilon isoforms are known to localize primarily in the cytoplasm. Here we studied the pathways that regulate B56alpha subcellular localization.

Differential modulation of BRCA1 and BARD1 nuclear localisation and foci assembly by DNA damage.

The BRCA1/BARD1 heterodimer regulates genomic maintenance and contributes to the DNA damage checkpoint response. We previously reported that BRCA1 and BARD1 can shuttle between nucleus and cytoplasm. In this study, we evaluated the localisation patterns of BRCA1 and BARD1 in response to different types of DNA damaging agents and chemotherapeutic drugs.

A comparison of BRCA1 nuclear localization with 14 DNA damage response proteins and domains: identification of specific differences between BRCA1 and 53BP1 at DNA damage-induced foci.

BRCA1 is an important mediator of the DNA damage response pathway. Previous studies have identified a number of proteins that associate with BRCA1 at nuclear foci after ionizing radiation (IR)-induced DNA damage. However, the co-localization patterns of BRCA1 and various DNA damage response proteins have not yet been systematically quantified and compared within the same experimental system.

IQGAP1 regulation and roles in cancer.

IQGAP1 is a key mediator of several distinct cellular processes, in particular cytoskeletal rearrangements. Recent studies have implicated a potential role for IQGAP1 in cancer, supported by the over-expression and distinct membrane localisation of IQGAP1 observed in a range of tumours. IQGAP1 is thought to contribute to the transformed cancer cell phenotype by regulating signalling pathways involved in cell proliferation and transformation, weakening of cell:cell adhesion contacts and stimulation of cell motility and invasion.

Regulation of beta-catenin trafficking to the membrane in living cells.

Beta-catenin is a key mediator of the Wnt signaling process and accumulates in the nucleus and at the membrane in response to Wnt-mediated inhibition of GSK-3beta. In this study we used live cell photobleaching experiments to determine the dynamics and rate of recruitment of beta-catenin at membrane adherens junctions (cell adhesion) and membrane ruffles (cell migration). First, we confirmed the nuclear-cytoplasmic shuttling of GFP-tagged beta-catenin, and found that a small mobile pool of beta-catenin can move from the nucleus to membrane ruffles in NIH 3T3 fibroblasts with a t(0.

APC shuttling to the membrane, nucleus and beyond.

The adenomatous polyposis coli (APC) tumor suppressor is a multi-functional protein, the mutation of which triggers colon cancer progression through de-regulation of the canonical Wnt signaling pathway and disruption of the mitotic spindle checkpoint. In recent years, APC has been detected at several unexpected intracellular locations, implicating APC in multiple roles that now include the regulation of directed cell migration, apoptosis and DNA repair. In this review, we discuss the intracellular trafficking pathway of APC and describe how truncated cancer-mutant forms of APC display frequent changes in sub-cellular localization and function.

BARD1 regulates BRCA1-mediated transactivation of the p21WAF1/CIP1 and Gadd45 promoters.

BRCA1 regulates gene transcription as part of its tumor suppressor function. Prior studies on BRCA1 transactivation did not account for the impact of its binding partner, BARD1. Here we tested the effect of BARD1 on BRCA1 transactivation of the p21 and Gadd45 promoters.