Publications by authors named "Roseline Godbout"

66 Publications

DDX1 vesicles control calcium-dependent mitochondrial activity in mouse embryos.

Nat Commun 2022 07 1;13(1):3794. Epub 2022 Jul 1.

Department of Oncology, Cross Cancer Institute, University of Alberta, Edmonton, AB, T6G 1Z2, Canada.

The DEAD box protein DDX1, previously associated with 3'-end RNA processing and DNA repair, forms large aggregates in the cytoplasm of early mouse embryos. Ddx1 knockout causes stalling of embryos at the 2-4 cell stages. Here, we identify a DDX1-containing membrane-bound calcium-containing organelle with a nucleic acid core. We show that aggregates of these organelles form ring-like structures in early-stage embryos which we have named Membrane Associated RNA-containing Vesicles. We present evidence that DDX1 is required for the formation of Membrane Associated RNA-containing Vesicles which in turn regulate the spatial distribution of calcium in embryos. We find that Ddx1 knockout in early embryos disrupts calcium distribution, and increases mitochondria membrane potential, mitochondrial activity, and reactive oxygen species. Sequencing analysis of embryos from Ddx1 heterozygote crosses reveals downregulation of a subset of RNAs involved in developmental and mitochondrial processes in the embryos with low Ddx1 RNA. We propose a role for Membrane Associated RNA-containing Vesicles in calcium-controlled mitochondrial functions that are essential for embryonic development.
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http://dx.doi.org/10.1038/s41467-022-31497-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9249788PMC
July 2022

DEAD box 1 (DDX1) protein binds to and protects cytoplasmic stress response mRNAs in cells exposed to oxidative stress.

J Biol Chem 2022 Jun 23;298(8):102180. Epub 2022 Jun 23.

Department of Oncology, Cross Cancer Institute, University of Alberta, Edmonton, Alberta, Canada. Electronic address:

The integrated stress response is a network of highly orchestrated pathways activated when cells are exposed to environmental stressors. While global repression of translation is a well-recognized hallmark of the integrated stress response, less is known about the regulation of mRNA stability during stress. DEAD box proteins are a family of RNA unwinding/remodeling enzymes involved in every aspect of RNA metabolism. We previously showed that DEAD box 1 (DDX1) protein accumulates at DNA double-strand breaks during genotoxic stress and promotes DNA double-strand break repair via homologous recombination. Here, we examine the role of DDX1 in response to environmental stress. We show that DDX1 is recruited to stress granules (SGs) in cells exposed to a variety of environmental stressors, including arsenite, hydrogen peroxide, and thapsigargin. We also show that DDX1 depletion delays resolution of arsenite-induced SGs. Using RNA immunoprecipitation sequencing, we identify RNA targets bound to endogenous DDX1, including RNAs transcribed from genes previously implicated in stress responses. We show the amount of target RNAs bound to DDX1 increases when cells are exposed to stress, and the overall levels of these RNAs are increased during stress in a DDX1-dependent manner. Even though DDX1's RNA-binding property is critical for maintenance of its target mRNA levels, we found RNA binding is not required for localization of DDX1 to SGs. Furthermore, DDX1 knockdown does not appear to affect RNA localization to SGs. Taken together, our results reveal a novel role for DDX1 in maintaining cytoplasmic mRNA levels in cells exposed to oxidative stress.
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http://dx.doi.org/10.1016/j.jbc.2022.102180DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9293777PMC
June 2022

FABP7 Facilitates Uptake of Docosahexaenoic Acid in Glioblastoma Neural Stem-like Cells.

Nutrients 2021 Jul 30;13(8). Epub 2021 Jul 30.

Department of Oncology, Cross Cancer Institute, University of Alberta, Edmonton, AB T6G 1Z2, Canada.

Glioblastoma (GBM) is an aggressive tumor with a dismal prognosis. Neural stem-like cells contribute to GBM's poor prognosis by driving drug resistance and maintaining cellular heterogeneity. GBM neural stem-like cells express high levels of brain fatty acid-binding protein (FABP7), which binds to polyunsaturated fatty acids (PUFAs) ω-6 arachidonic acid (AA) and ω-3 docosahexaenoic acid (DHA). Similar to brain, GBM tissue is enriched in AA and DHA. However, DHA levels are considerably lower in GBM tissue compared to adult brain. Therefore, it is possible that increasing DHA content in GBM, particularly in neural stem-like cells, might have therapeutic value. Here, we examine the fatty acid composition of patient-derived GBM neural stem-like cells grown as neurosphere cultures. We also investigate the effect of AA and DHA treatment on the fatty acid profiles of GBM neural stem-like cells with or without FABP7 knockdown. We show that DHA treatment increases DHA levels and the DHA:AA ratio in GBM neural stem-like cells, with FABP7 facilitating the DHA uptake. We also found that an increased uptake of DHA inhibits the migration of GBM neural stem-like cells. Our results suggest that increasing DHA content in the GBM microenvironment may reduce the migration/infiltration of FABP7-expressing neural stem-like cancer cells.
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http://dx.doi.org/10.3390/nu13082664DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8402214PMC
July 2021

Super resolution microscopy reveals DHA-dependent alterations in glioblastoma membrane remodelling and cell migration.

Nanoscale 2021 Jun;13(21):9706-9722

Cross Cancer Institute, University of Alberta, Department of Oncology, 11560 University Avenue, Edmonton, Alberta, Canada.

Brain fatty acid binding protein (FABP7; B-FABP) promotes glioblastoma (GBM) cell migration and is associated with tumor infiltration, properties associated with a poor prognosis in GBM patients. FABP7-expressing neural stem-like cells are known to drive tumor migration/infiltration and resistance to treatment. We have previously shown that FABP7's effects on cell migration can be reversed when GBM cells are cultured in medium supplemented with the omega-3 fatty acid, docosahexaenoic acid (DHA). Here, we use super-resolution imaging on patient-derived GBM stem-like cells to examine the importance of FABP7 and its fatty acid ligands in mitigating GBM cell migration. As FABPs are involved in fatty acid transport from membrane to cytosol, we focus on the effect of FABP7 and its ligand DHA on GBM membrane remodeling, as well as FABP7 nanoscale domain formation on GBM membrane. Using quantitative plasma membrane lipid order imaging, we show that FABP7 expression in GBM cells correlates with increased membrane lipid order, with DHA dramatically decreasing lipid order. Using super-resolution stimulated emission depletion (STED) microscopy, we observe non-uniform distribution of FABP7 on the surface of GBM cells, with FABP7 forming punctate nanoscale domains of ∼100 nm in diameter. These nanodomains are particularly enriched at the migrating front of GBM cells. Interestingly, FABP7 nanodomains are disrupted when GBM cells are cultured in DHA-supplemented medium. We demonstrate a tight link between cell migration, a higher membrane lipid order and increased FABP7 nanoscale domains. We propose that DHA-mediated disruption of membrane lipid order and FABP7 nanodomains forms the basis of FABP7/DHA-mediated inhibition of cell migration in GBM.
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http://dx.doi.org/10.1039/d1nr02128aDOI Listing
June 2021

An Amplified Fatty Acid-Binding Protein Gene Cluster in Prostate Cancer: Emerging Roles in Lipid Metabolism and Metastasis.

Cancers (Basel) 2020 Dec 18;12(12). Epub 2020 Dec 18.

Department of Oncology, Cross Cancer Institute, University of Alberta, Edmonton, AB T6G 1Z2, Canada.

Treatment for early stage and localized prostate cancer (PCa) is highly effective. Patient survival, however, drops dramatically upon metastasis due to drug resistance and cancer recurrence. The molecular mechanisms underlying PCa metastasis are complex and remain unclear. It is therefore crucial to decipher the key genetic alterations and relevant molecular pathways driving PCa metastatic progression so that predictive biomarkers and precise therapeutic targets can be developed. Through PCa cohort analysis, we found that a fatty acid-binding protein (FABP) gene cluster (containing five FABP family members) is preferentially amplified and overexpressed in metastatic PCa. All five FABP genes reside on chromosome 8 at 8q21.13, a chromosomal region frequently amplified in PCa. There is emerging evidence that these FABPs promote metastasis through distinct biological actions and molecular pathways. In this review, we discuss how these FABPs may serve as drivers/promoters for PCa metastatic transformation using patient cohort analysis combined with a review of the literature.
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http://dx.doi.org/10.3390/cancers12123823DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7766576PMC
December 2020

The FABP12/PPARγ pathway promotes metastatic transformation by inducing epithelial-to-mesenchymal transition and lipid-derived energy production in prostate cancer cells.

Mol Oncol 2020 12 23;14(12):3100-3120. Epub 2020 Oct 23.

Department of Oncology, Cross Cancer Institute, University of Alberta, Edmonton, AB, Canada.

Early stage localized prostate cancer (PCa) has an excellent prognosis; however, patient survival drops dramatically when PCa metastasizes. The molecular mechanisms underlying PCa metastasis are complex and remain unclear. Here, we examine the role of a new member of the fatty acid-binding protein (FABP) family, FABP12, in PCa progression. FABP12 is preferentially amplified and/or overexpressed in metastatic compared to primary tumors from both PCa patients and xenograft animal models. We show that FABP12 concurrently triggers metastatic phenotypes (induced epithelial-to-mesenchymal transition (EMT) leading to increased cell motility and invasion) and lipid bioenergetics (increased fatty acid uptake and accumulation, increased ATP production from fatty acid β-oxidation) in PCa cells, supporting increased reliance on fatty acids for energy production. Mechanistically, we show that FABP12 is a driver of PPARγ activation which, in turn, regulates FABP12's role in lipid metabolism and PCa progression. Our results point to a novel role for a FABP-PPAR pathway in promoting PCa metastasis through induction of EMT and lipid bioenergetics.
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http://dx.doi.org/10.1002/1878-0261.12818DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7718947PMC
December 2020

Cytoplasmic aggregation of DDX1 in developing embryos: Early embryonic lethality associated with Ddx1 knockout.

Dev Biol 2019 11 19;455(2):420-433. Epub 2019 Jul 19.

Department of Oncology, Cross Cancer Institute, University of Alberta, Edmonton, Alberta, T6G 1Z2, Canada. Electronic address:

Temporally-regulated maternal RNA translation is essential for embryonic development, with defective degradation resulting in stalled 2-cell embryos. We show that DDX1, a DEAD box protein implicated in RNA transport, may be a key regulator of maternal RNA utilization. DDX1 protein localizes exclusively to cytoplasmic granules in both oocytes and early stage mouse embryos, with DDX1 requiring RNA for retention at these sites. Homozygous knockout of Ddx1 causes stalling of mouse embryos at the 2-4 cell stages. These results suggest a maternal RNA-dependent role for DDX1 in the progression of embryos past the 2-4 cell stage. The change in appearance of DDX1-containing granules in developing embryos further supports a role in temporally-regulated degradation of RNAs. We carried out RNA-immunoprecipitations (RNA-IPs) to identify mRNAs bound to DDX1 in 2-cell embryos, focusing on 16 maternal genes previously shown to be essential for embryonic development past the 1- to 2-cell stages. Five of these RNAs were preferentially bound by DDX1: Ago2, Zar1, Tle6, Floped and Tif1α. We propose that DDX1 controls access to subsets of key maternal RNAs required for early embryonic development.
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http://dx.doi.org/10.1016/j.ydbio.2019.07.014DOI Listing
November 2019

A positive feedback loop involving nuclear factor IB and calpain 1 suppresses glioblastoma cell migration.

J Biol Chem 2019 08 1;294(34):12638-12654. Epub 2019 Jul 1.

Cross Cancer Institute, Department of Oncology, University of Alberta, Edmonton, Alberta T6G 1Z2, Canada

Glioblastoma (GBM) is a brain tumor that remains largely incurable because of its highly-infiltrative properties. Nuclear factor I (NFI)-type transcription factors regulate genes associated with GBM cell migration and infiltration. We have previously shown that NFI activity depends on the NFI phosphorylation state and that calcineurin phosphatase dephosphorylates and activates NFI. Calcineurin is cleaved and activated by calpain proteases whose activity is, in turn, regulated by an endogenous inhibitor, calpastatin (CAST). The gene is a target of NFI in GBM cells, with differentially phosphorylated NFIs regulating the levels of transcript variants. Here, we uncovered an NFIB-calpain 1-positive feedback loop mediated through CAST and calcineurin. In NFI-hyperphosphorylated GBM cells, NFIB expression decreased the CAST-to-calpain 1 ratio in the cytoplasm. This reduced ratio increased autolysis and activity of cytoplasmic calpain 1. Conversely, in NFI-hypophosphorylated cells, NFIB expression induced differential subcellular compartmentalization of CAST and calpain 1, with CAST localizing primarily to the cytoplasm and calpain 1 to the nucleus. Overall, this altered compartmentalization increased nuclear calpain 1 activity. We also show that nuclear calpain 1, by cleaving and activating calcineurin, induces NFIB dephosphorylation. Of note, knockdown of calpain 1, NFIB, or both increased GBM cell migration and up-regulated the pro-migratory factors fatty acid-binding protein 7 (FABP7) and Ras homolog family member A (RHOA). In summary, our findings reveal bidirectional cross-talk between NFIB and calpain 1 in GBM cells. A physiological consequence of this positive feedback loop appears to be decreased GBM cell migration.
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http://dx.doi.org/10.1074/jbc.RA119.008291DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6709629PMC
August 2019

Non-canonical BAD activity regulates breast cancer cell and tumor growth via 14-3-3 binding and mitochondrial metabolism.

Oncogene 2019 05 11;38(18):3325-3339. Epub 2019 Jan 11.

Department of Biochemistry, University of Alberta, Edmonton, AB, T6G 2H7, Canada.

The Bcl-2-associated death promoter BAD is a prognostic indicator for good clinical outcome of breast cancer patients; however, whether BAD affects breast cancer biology is unknown. Here we showed that BAD increased cell growth in breast cancer cells through two distinct mechanisms. Phosphorylation of BAD at S118 increased S99 phosphorylation, 14-3-3 binding and AKT activation to promote growth and survival. Through a second, more prominent pathway, BAD stimulated mitochondrial oxygen consumption in a novel manner that was downstream of substrate entry into the mitochondria. BAD stimulated complex I activity that facilitated enhanced cell growth and sensitized cells to apoptosis in response to complex I blockade. We propose that this dependence on oxidative metabolism generated large but nonaggressive cancers. This model identifies a non-canonical role for BAD and reconciles BAD-mediated tumor growth with favorable outcomes in BAD-high breast cancer patients.
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http://dx.doi.org/10.1038/s41388-018-0673-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6756016PMC
May 2019

Effects of nuclear factor I phosphorylation on calpastatin () gene variant expression and subcellular distribution in malignant glioma cells.

J Biol Chem 2019 01 30;294(4):1173-1188. Epub 2018 Nov 30.

Department of Oncology, University of Alberta, Cross Cancer Institute, Edmonton, Alberta T6G 1Z2, Canada. Electronic address:

Malignant glioma (MG) is the most lethal primary brain tumor. In addition to having inherent resistance to radiation treatment and chemotherapy, MG cells are highly infiltrative, rendering focal therapies ineffective. Genes involved in MG cell migration and glial cell differentiation are up-regulated by hypophosphorylated nuclear factor I (NFI), which is dephosphorylated by the phosphatase calcineurin in MG cells. Calcineurin is cleaved and thereby activated by calpain proteases, which are, in turn, inhibited by calpastatin (CAST). Here, we show that the gene is a target of NFI and has NFI-binding sites in its intron 3 region. We also found that NFI-mediated regulation of depends on NFI's phosphorylation state. We noted that occupation of intron 3 by hypophosphorylated NFI results in increased activation of an alternative promoter. This activation resulted in higher levels of transcript variants, leading to increased levels of CAST protein that lacks the N-terminal XL domain. CAST was primarily present in the cytoplasm of NFI-hypophosphorylated MG cells, with a predominantly perinuclear immunostaining pattern. NFI knockdown in NFI-hypophosphorylated MG cells increased CAST levels at the plasma membrane. These results suggest that NFI plays an integral role in the regulation of variants and CAST subcellular distribution. Along with the previous findings indicating that NFI activity is regulated by calcineurin, these results provide a foundation for further investigations into the possibility of regulatory cross-talk between NFI and the CAST/calpain/calcineurin signaling pathway in MG cells.
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http://dx.doi.org/10.1074/jbc.RA118.004787DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6349116PMC
January 2019

NFIB promotes cell survival by directly suppressing p21 transcription in TP53-mutated triple-negative breast cancer.

J Pathol 2019 02 18;247(2):186-198. Epub 2018 Dec 18.

Department of Oncology, University of Alberta, Cross Cancer Institute, Edmonton, Canada.

Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype with limited treatment options and poor prognosis. There is an urgent need to identify and understand the key factors and signalling pathways driving TNBC tumour progression, relapse, and treatment resistance. In this study, we report that gene copy numbers and expression levels of nuclear factor IB (NFIB), a recently identified oncogene in small cell lung cancer, are preferentially increased in TNBC compared to other breast cancer subtypes. Furthermore, increased levels of NFIB are significantly associated with high tumour grade, poor prognosis, and reduced chemotherapy response. Concurrent TP53 mutations and NFIB overexpression (z-scores > 0) were observed in 77.9% of TNBCs, in contrast to 28.5% in non-TNBCs. Depletion of NFIB in TP53-mutated TNBC cell lines promotes cell death, cell cycle arrest, and enhances sensitivity to docetaxel, a first-line chemotherapeutic drug in breast cancer treatment. Importantly, these alterations in growth properties were accompanied by induction of CDKN1A, the gene encoding p21, a downstream effector of p53. We show that NFIB directly interacts with the CDKN1A promoter in TNBC cells. Furthermore, knockdown of combined p21 and NFIB reverses the docetaxel-induced cell growth inhibition observed upon NFIB knockdown, indicating that NFIB's effect on chemotherapeutic drug response is mediated through p21. Our results indicate that NFIB is an important TNBC factor that drives tumour cell growth and drug resistance, leading to poor clinical outcomes. Thus, targeting NFIB in TP53-mutated TNBC may reverse oncogenic properties associated with mutant p53 by restoring p21 activity. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
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http://dx.doi.org/10.1002/path.5182DOI Listing
February 2019

Nuclear Factor I Represses the Notch Effector HEY1 in Glioblastoma.

Neoplasia 2018 10 6;20(10):1023-1037. Epub 2018 Sep 6.

Department of Oncology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, Alberta, Canada, T6G 1Z2. Electronic address:

Glioblastomas (GBMs) are highly aggressive brain tumors with a dismal prognosis. Nuclear factor I (NFI) is a family of transcription factors that controls glial cell differentiation in the developing central nervous system. NFIs have previously been shown to regulate the expression of astrocyte markers such as glial fibrillary acidic protein (GFAP) in both normal brain and GBM cells. We used chromatin immunoprecipitation (ChIP)-on-chip to identify additional NFI targets in GBM cells. Analysis of our ChIP data revealed ~400 putative NFI target genes including an effector of the Notch signaling pathway, HEY1, implicated in the maintenance of neural stem cells. All four NFIs (NFIA, NFIB, NFIC, and NFIX) bind to NFI recognition sites located within 1 kb upstream of the HEY1 transcription site. We further showed that NFI negatively regulates HEY1 expression, with knockdown of all four NFIs in GBM cells resulting in increased HEY1 RNA levels. HEY1 knockdown in GBM cells decreased cell proliferation, increased cell migration, and decreased neurosphere formation. Finally, we found a general correlation between elevated levels of HEY1 and expression of the brain neural stem/progenitor cell marker B-FABP in GBM cell lines. Knockdown of HEY1 resulted in an increase in the RNA levels of the GFAP astrocyte differentiation marker. Overall, our data indicate that HEY1 is negatively regulated by NFI family members and is associated with increased proliferation, decreased migration, and increased stem cell properties in GBM cells.
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http://dx.doi.org/10.1016/j.neo.2018.08.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6138789PMC
October 2018

ω-3 and ω-6 Fatty Acids Modulate Conventional and Atypical Protein Kinase C Activities in a Brain Fatty Acid Binding Protein Dependent Manner in Glioblastoma Multiforme.

Nutrients 2018 Apr 6;10(4). Epub 2018 Apr 6.

Department of Oncology, Cross Cancer Institute, University of Alberta, Edmonton, AB T6G 1Z2, Canada.

Glioblastoma multiforme (GBM) is a highly infiltrative brain cancer with a dismal prognosis. High levels of brain fatty acid binding protein (B-FABP) are associated with increased migration/infiltration in GBM cells, with a high ratio of arachidonic acid (AA) to docosahexaenoic acid (DHA) driving B-FABP-mediated migration. Since several protein kinase Cs (PKCs) are overexpressed in GBM and linked to migration, we explored a possible relationship between B-FABP and levels/activity of different PKCs, as a function of AA and DHA supplementation. We report that ectopic expression of B-FABP in U87 cells alters the levels of several PKCs, particularly PKC. Upon analysis of RNA levels in a panel of GBM cell lines and patient-derived GBM neurospheres, we observed a trend towards moderate positive correlation ( = 0.624, = 0.054) between B-FABP and RNA levels. Analysis of PKC activity in U87 GBM cells revealed decreased typical PKC activity (23.4%) in B-FABP-expressing cells compared with nonexpressing cells, with no difference in novel and atypical PKC activities. AA and DHA modulated both conventional and atypical PKC activities in a B-FABP-dependent manner, but had no effect on novel PKC activity. These results suggest that conventional and atypical PKCs are potential downstream effectors of B-FABP/fatty acid-mediated alterations in GBM growth properties.
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http://dx.doi.org/10.3390/nu10040454DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5946239PMC
April 2018

AP-2ε Expression in Developing Retina: Contributing to the Molecular Diversity of Amacrine Cells.

Sci Rep 2018 02 21;8(1):3386. Epub 2018 Feb 21.

Department of Oncology, University of Alberta, Cross Cancer Institute, Edmonton, Alberta, Canada.

AP-2 transcription factors play important roles in the regulation of gene expression during development. Four of the five members of the AP-2 family (AP-2α, AP-2β, AP-2γ and AP-2δ) have previously been shown to be expressed in developing retina. Mouse knockouts have revealed roles for AP-2α, AP-2β and AP-2δ in retinal cell specification and function. Here, we show that the fifth member of the AP-2 family, AP-2ε, is also expressed in amacrine cells in developing mammalian and chicken retina. Our data indicate that there are considerably fewer AP-2ε-positive cells in the developing mouse retina compared to AP-2α, AP-2β and AP-2γ-positive cells, suggesting a specialized role for AP-2ε in a subset of amacrine cells. AP-2ε, which is restricted to the GABAergic amacrine lineage, is most commonly co-expressed with AP-2α and AP-2β, especially at early stages of retinal development. Co-expression of AP-2ε and AP-2γ increases with differentiation. Analysis of previously published Drop-seq data from single retinal cells supports co-expression of multiple AP-2s in the same cell. Since AP-2s bind to their target sequences as either homodimers or heterodimers, our work suggests spatially- and temporally-coordinated roles for combinations of AP-2 transcription factors in amacrine cells during retinal development.
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http://dx.doi.org/10.1038/s41598-018-21822-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5821864PMC
February 2018

Alternative Splicing of Disabled-1 Controls Multipolar-to-Bipolar Transition of Migrating Neurons in the Neocortex.

Cereb Cortex 2018 10;28(10):3457-3467

Department of Neurobiology, Key Laboratory of Medical Neurobiology of Ministry of Health of China, Key Laboratory of Neurobiology of Zhejiang Province, Zhejiang University School of Medicine, Hangzhou, China.

Multipolar-to-bipolar transition (MBT) is crucial for the neuronal migration and positioning in the neocortex. Reelin-Disabled-1 (Dab1) signaling plays a pivotal role in neuronal migration, yet how Dab1 coordinatively regulates downstream molecules to affect MBT remains unclear. We have previously found that alternative splicing produces multiple Dab1 isoforms with different tyrosine motifs and differential ability to recruit downstream effectors. Here, we report that splicing of Dab1 exons 7 and 8 and 9bc dynamically regulates the inclusion and activities of Dab1 tyrosine motifs in the neocortex. By in utero electroporation, we show that expression of Dab1 isoforms missing exons 7 and 8 or retaining exons 9bc in WT neurons resulted in neuronal migration defects with attenuated Dab1 tyrosine phosphorylation, disrupted leading process extension, and disorientated multipolar neurons in the multipolar accumulation zone. Introducing the canonical Dab1 form, but not those missing exons 7 and 8 or retaining exons 9bc, into Dab1-deficient neurons promoted MBT and rescued neuronal migration defects, suggesting that alternative splicing of Dab1 modulates the tyrosine motif switch and mediates MBT of cortical neurons. Our study reveals a critical mechanism by which Dab1 alternative splicing coordinately controls MBT and neuronal migration in a spatiotemporal manner.
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http://dx.doi.org/10.1093/cercor/bhx212DOI Listing
October 2018

Role for RIF1-interacting partner DDX1 in BLM recruitment to DNA double-strand breaks.

DNA Repair (Amst) 2017 07 13;55:47-63. Epub 2017 May 13.

Department of Oncology, Cross Cancer Institute, University of Alberta, 11560 University Avenue, Edmonton Alberta T6G 1Z2, Canada. Electronic address:

Human Rap1-interacting factor 1 (RIF1) is an important player in the repair of DNA double strand breaks (DSBs). RIF1 acts downstream of 53BP1, with well-documented roles in class switch recombination in B-cells and inhibition of end resection initiation in BRCA1-defective cells. Here, we report that DEAD Box 1 (DDX1), a RNA helicase also implicated in DSB repair, interacts with RIF1, with co-localization of DDX1 and RIF1 observed throughout interphase. Recruitment of DDX1 to DSBs is dependent on RIF1, with RIF1 depletion abolishing DDX1-mediated facilitation of homologous recombination at DSBs. As previously demonstrated for RIF1, DDX1 is also required for chromatin loading of Bloom syndrome helicase (BLM) to ionizing radiation-induced DSBs, a RIF1-related activity that is independent of 53BP1. We show that DDX1 and RIF1 have different nucleic acid requirements for accumulation at DSBs, with RNA-DNA hybrids required for DDX1 accrual at DSBs, and single-strand RNA required for accumulation of RIF1 at these sites. Our data suggest both convergent and divergent roles for DDX1 and RIF1 in DSB repair, and may help explain why RIF1 depletion does not fully mimic 53BP1 ablation in the restoration of homologous recombination defects in BRCA1-deficient cells.
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http://dx.doi.org/10.1016/j.dnarep.2017.05.001DOI Listing
July 2017

Functional assessment of von Willebrand factor expression by cancer cells of non-endothelial origin.

Oncotarget 2017 Feb;8(8):13015-13029

Department of Medicine, University of Alberta, Edmonton, Alberta, Canada.

Von Willebrand factor (VWF) is a highly adhesive procoagulant molecule that mediates platelet adhesion to endothelial and subendothelial surfaces. Normally it is expressed exclusively in endothelial cells (ECs) and megakaryocytes. However, a few studies have reported VWF detection in cancer cells of non-endothelial origin, including osteosarcoma. A role for VWF in cancer metastasis has long been postulated but evidence supporting both pro- and anti-metastatic roles for VWF has been presented. We hypothesized that the role of VWF in cancer metastasis is influenced by its cellular origin and that cancer cell acquisition of VWF expression may contribute to enhanced metastatic potential. We demonstrated de novo expression of VWF in glioma as well as osteosarcoma cells. Endothelial monolayer adhesion, transmigration and extravasation capacities of VWF expressing cancer cells were shown to be enhanced compared to non-VWF expressing cells, and were significantly reduced as a result of VWF knock down. VWF expressing cancer cells were also detected in patient tumor samples of varying histologies. Analyses of the mechanism of transcriptional activation of the VWF in cancer cells demonstrated a pattern of trans-activating factor binding and epigenetic modifications consistent overall with that observed in ECs. These results demonstrate that cancer cells of non-endothelial origin can acquire de novo expression of VWF, which can enhance processes, including endothelial and platelet adhesion and extravasation, that contribute to cancer metastasis.
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http://dx.doi.org/10.18632/oncotarget.14273DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5355073PMC
February 2017

DEAD Box 1 Facilitates Removal of RNA and Homologous Recombination at DNA Double-Strand Breaks.

Mol Cell Biol 2016 11 28;36(22):2794-2810. Epub 2016 Oct 28.

Department of Oncology, University of Alberta, Cross Cancer Institute, Edmonton, Alberta, Canada

Although RNA and RNA-binding proteins have been linked to double-strand breaks (DSBs), little is known regarding their roles in the cellular response to DSBs and, if any, in the repair process. Here, we provide direct evidence for the presence of RNA-DNA hybrids at DSBs and suggest that binding of RNA to DNA at DSBs may impact repair efficiency. Our data indicate that the RNA-unwinding protein DEAD box 1 (DDX1) is required for efficient DSB repair and cell survival after ionizing radiation (IR), with depletion of DDX1 resulting in reduced DSB repair by homologous recombination (HR). While DDX1 is not essential for end resection, a key step in homology-directed DSB repair, DDX1 is required for maintenance of the single-stranded DNA once generated by end resection. We show that transcription deregulation has a significant effect on DSB repair by HR in DDX1-depleted cells and that RNA-DNA duplexes are elevated at DSBs in DDX1-depleted cells. Based on our combined data, we propose a role for DDX1 in resolving RNA-DNA structures that accumulate at DSBs located at sites of active transcription. Our findings point to a previously uncharacterized requirement for clearing RNA at DSBs for efficient repair by HR.
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http://dx.doi.org/10.1128/MCB.00415-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5086521PMC
November 2016

Characterization of femtosecond-laser pulse induced cell membrane nanosurgical attachment.

Biomed Opt Express 2016 Jul 22;7(7):2749-58. Epub 2016 Jun 22.

Department of Electrical and Computer Engineering, Ultrafast Photonics and Nano-Optics Laboratory, University of Alberta, Edmonton, AB T6G 2V4, Canada.

This article provides insight into the mechanism of femtosecond laser nanosurgical attachment of cells. We have demonstrated that during the attachment of two retinoblastoma cells using sub-10 femtosecond laser pulses, with 800 nm central wavelength, the phospholipid molecules of both cells hemifuse and form one shared phospholipid bilayer, at the attachment location. In order to verify the hypothesis that hemifusion takes place, transmission electron microscope images of the cell membranes of retinoblastoma cells were taken. It is shown that at the attachment interface, the two cell membranes coalesce and form one single membrane shared by both cells. Thus, further evidence is provided to support the hypothesis that laser-induced ionization process led to an ultrafast reversible destabilization of the phospholipid layer of the cellular membrane, which resulted in cross-linking of the phospholipid molecules in each membrane. This process of hemifusion occurs throughout the entire penetration depth of the femtosecond laser pulse train. Thus, the attachment between the cells takes place across a large surface area, which affirms our findings of strong physical attachment between the cells. The femtosecond laser pulse hemifusion technique can potentially provide a platform for precise molecular manipulation of cellular membranes. Manipulation of the cellular membrane is an important procedure that could aid in studying diseases such as cancer; where the expression level of plasma proteins on the cell membrane is altered.
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http://dx.doi.org/10.1364/BOE.7.002749DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4948627PMC
July 2016

Loss of AP-2delta reduces retinal ganglion cell numbers and axonal projections to the superior colliculus.

Mol Brain 2016 06 4;9(1):62. Epub 2016 Jun 4.

Department of Oncology, Cross Cancer Institute, University of Alberta, 11560 University Avenue, Edmonton, AB, T6G 1Z2, Canada.

Background: AP-2δ is the most divergent member of the Activating Protein-2 (TFAP2) family of transcription factors. AP-2δ is restricted to specific regions of the CNS, including a subset of ganglion cells in the retina. Retinal ganglion cells (RGCs), the only output neurons of the retina, are responsible for transmitting the visual signal to the brain.

Results: AP-2δ knockout results in loss of Brn3c (Pou4f3) expression in AP-2δ -positive RGCs. While AP-2δ-/- mice have morphologically normal retinas at birth, there is a significant reduction in retinal ganglion cell numbers by P21, after eye opening. Chromatin immunoprecipitation indicates that Brn3c is a target of AP-2δ in the retina. Using fluorochrome-conjugated cholera toxin subunit B to trace ganglion cell axons from the eye to the major visual pathways in the brain, we found 87 % and 32 % decreases in ipsilateral and contralateral projections, respectively, to the superior colliculus in AP-2δ-/- mice. In agreement with anatomical data, visually evoked responses recorded from the brain confirmed that retinal outputs to the brain are compromised.

Conclusions: AP-2δ is important for the maintenance of ganglion cell numbers in the retina. Loss of AP-2δ alters retinal axonal projections to visual centers of the brain, with ipsilaterial projections to the superior colliculus being the most dramatically affected. Our results have important implications for integration of the visual signal at the superior colliculus.
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http://dx.doi.org/10.1186/s13041-016-0244-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4893287PMC
June 2016

Notch and TGFβ form a positive regulatory loop and regulate EMT in epithelial ovarian cancer cells.

Cell Signal 2016 08 10;28(8):838-49. Epub 2016 Apr 10.

Department of Oncology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada; Department of Obstetrics and Gynecology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada. Electronic address:

Epithelial-mesenchymal transition (EMT) plays a critical role in the progression of epithelial ovarian cancer (EOC). However, the mechanisms that regulate EMT in EOC are not fully understood. Here, we report that activation of Notch1 induces EMT in EOC cells as evidenced by downregulation of E-cadherin and cytokeratins, upregulation of Slug and Snail, as well as morphological changes. Interestingly, activation of Notch1 increases TGFβ/Smad signaling by upregulating the expression of TGFβ and TGFβ type 1 receptor. Time course experiments demonstrate that inhibition of Notch by DAPT (a γ-secretase inhibitor) decreases TGFβ-induced phosphorylation of receptor Smads at late, but not at early, timepoints. These results suggest that Notch activation plays a role in sustaining TGFβ/Smad signaling in EOC cells. Furthermore, inhibition of Notch by DAPT decreases TGFβ induction of Slug and repression of E-cadherin and knockdown of Notch1 decreases TGFβ-induced repression of E-cadherin, indicating that Notch is required, at least in part, for TGFβ-induced EMT in EOC cells. On the other hand, TGFβ treatment increases the expression of Notch ligand Jagged1 and Notch target gene HES1 in EOC cells. Functionally, the combination of Notch1 activation and TGFβ treatment is more potent in promoting motility and migration of EOC cells than either stimulation alone. Taken together, our results indicate that Notch and TGFβ form a reciprocal positive regulatory loop and cooperatively regulate EMT and promote EOC cell motility and migration.
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http://dx.doi.org/10.1016/j.cellsig.2016.03.016DOI Listing
August 2016

Association between cytoplasmic CRABP2, altered retinoic acid signaling, and poor prognosis in glioblastoma.

Glia 2016 Jun 19;64(6):963-76. Epub 2016 Feb 19.

Department of Oncology, University of Alberta, Cross Cancer Institute, 11560 University Avenue, Edmonton, Alberta, T6G 1Z2, Canada.

Retinoic acid (RA), a metabolite of vitamin A, is required for the regulation of growth and development. Aberrant expression of molecules involved in RA signaling has been reported in various cancer types including glioblastoma multiforme (GBM). Cellular retinoic acid-binding protein 2 (CRABP2) has previously been shown to play a key role in the transport of RA to retinoic acid receptors (RARs) to activate their transcription regulatory activity. Here, we demonstrate that CRABP2 is predominantly located in the cytoplasm of GBM tumors. Cytoplasmic, but not nuclear, CRABP2 levels in GBM tumors are associated with poor patient survival. Treatment of malignant glioma cell lines with RA results in a dose-dependent increase in accumulation of CRABP2 in the cytoplasm. CRABP2 knockdown reduces proliferation rates of malignant glioma cells, and enhances RA-induced RAR activation. Levels of CRYAB, a small heat shock protein with anti-apoptotic activity, and GFAP, an astrocyte-specific intermediate filament protein, are greatly reduced in CRABP2-depleted cells. Restoration of CRYAB expression partially but significantly reversed the effect of CRABP2 depletion on RAR activation. Our combined in vivo and in vitro data indicate that: (i) CRABP2 is an important determinant of clinical outcome in GBM patients, and (ii) the mechanism of action of CRABP2 in GBM involves sequestration of RA in the cytoplasm and activation of an anti-apoptotic pathway, thereby enhancing proliferation and preventing RA-mediated cell death and differentiation. We propose that reducing CRABP2 levels may enhance the therapeutic index of RA in GBM patients.
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http://dx.doi.org/10.1002/glia.22976DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5595534PMC
June 2016

Oxidative stress contributes to the tamoxifen-induced killing of breast cancer cells: implications for tamoxifen therapy and resistance.

Sci Rep 2016 Feb 17;6:21164. Epub 2016 Feb 17.

Signal Transduction Research Group, Department of Biochemistry, University of Alberta, Edmonton, Alberta, T6G 2S2, Canada.

Tamoxifen is the accepted therapy for patients with estrogen receptor-α (ERα)-positive breast cancer. However, clinical resistance to tamoxifen, as demonstrated by recurrence or progression on therapy, is frequent and precedes death from metastases. To improve breast cancer treatment it is vital to understand the mechanisms that result in tamoxifen resistance. This study shows that concentrations of tamoxifen and its metabolites, which accumulate in tumors of patients, killed both ERα-positive and ERα-negative breast cancer cells. This depended on oxidative damage and anti-oxidants rescued the cancer cells from tamoxifen-induced apoptosis. Breast cancer cells responded to tamoxifen-induced oxidation by increasing Nrf2 expression and subsequent activation of the anti-oxidant response element (ARE). This increased the transcription of anti-oxidant genes and multidrug resistance transporters. As a result, breast cancer cells are able to destroy or export toxic oxidation products leading to increased survival from tamoxifen-induced oxidative damage. These responses in cancer cells also occur in breast tumors of tamoxifen-treated mice. Additionally, high levels of expression of Nrf2, ABCC1, ABCC3 plus NAD(P)H dehydrogenase quinone-1 in breast tumors of patients at the time of diagnosis were prognostic of poor survival after tamoxifen therapy. Therefore, overcoming tamoxifen-induced activation of the ARE could increase the efficacy of tamoxifen in treating breast cancer.
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http://dx.doi.org/10.1038/srep21164DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4756695PMC
February 2016

Novel Method for Neuronal Nanosurgical Connection.

Sci Rep 2016 Feb 5;6:20529. Epub 2016 Feb 5.

Department of Electrical and Computer Engineering, University of Alberta, Edmonton, AB, Canada.

Neuronal injury may cause an irreversible damage to cellular, organ and organism function. While preventing neural injury is ideal, it is not always possible. There are multiple etiologies for neuronal injury including trauma, infection, inflammation, immune mediated disorders, toxins and hereditary conditions. We describe a novel laser application, utilizing femtosecond laser pulses, in order to connect neuronal axon to neuronal soma. We were able to maintain cellular viability, and demonstrate that this technique is universal as it is applicable to multiple cell types and media.
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http://dx.doi.org/10.1038/srep20529DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4742801PMC
February 2016

Long-Term Effect of Docosahexaenoic Acid Feeding on Lipid Composition and Brain Fatty Acid-Binding Protein Expression in Rats.

Nutrients 2015 Oct 22;7(10):8802-17. Epub 2015 Oct 22.

Department of Oncology, Cross Cancer Institute, University of Alberta, 11560 University Avenue, Edmonton, AB T6G 1Z2, Canada.

Arachidonic (AA) and docosahexaenoic acid (DHA) brain accretion is essential for brain development. The impact of DHA-rich maternal diets on offspring brain fatty acid composition has previously been studied up to the weanling stage; however, there has been no follow-up at later stages. Here, we examine the impact of DHA-rich maternal and weaning diets on brain fatty acid composition at weaning and three weeks post-weaning. We report that DHA supplementation during lactation maintains high DHA levels in the brains of pups even when they are fed a DHA-deficient diet for three weeks after weaning. We show that boosting dietary DHA levels for three weeks after weaning compensates for a maternal DHA-deficient diet during lactation. Finally, our data indicate that brain fatty acid binding protein (FABP7), a marker of neural stem cells, is down-regulated in the brains of six-week pups with a high DHA:AA ratio. We propose that elevated levels of DHA in developing brain accelerate brain maturation relative to DHA-deficient brains.
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http://dx.doi.org/10.3390/nu7105433DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4632453PMC
October 2015

Loss of the Drosophila melanogaster DEAD box protein Ddx1 leads to reduced size and aberrant gametogenesis.

Dev Biol 2015 Nov 1;407(2):232-45. Epub 2015 Oct 1.

Departments of Oncology University of Alberta, Cross Cancer Institute, 11560 University Avenue, Edmonton, Alberta, T6G 1Z2, Canada. Electronic address:

Mammalian DDX1 has been implicated in RNA trafficking, DNA double-strand break repair and RNA processing; however, little is known about its role during animal development. Here, we report phenotypes associated with a null Ddx1 (Ddx1(AX)) mutation generated in Drosophila melanogaster. Ddx1 null flies are viable but significantly smaller than control and Ddx1 heterozygous flies. Female Ddx1 null flies have reduced fertility with egg chambers undergoing autophagy, whereas males are sterile due to disrupted spermatogenesis. Comparative RNA sequencing of control and Ddx1 null third instars identified several transcripts affected by Ddx1 inactivation. One of these, Sirup mRNA, was previously shown to be overexpressed under starvation conditions and implicated in mitochondrial function. We demonstrate that Sirup is a direct binding target of Ddx1 and that Sirup mRNA is differentially spliced in the presence or absence of Ddx1. Combining Ddx1 null mutation with Sirup dsRNA-mediated knock-down causes epistatic lethality not observed in either single mutant. Our data suggest a role for Drosophila Ddx1 in stress-induced regulation of splicing.
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http://dx.doi.org/10.1016/j.ydbio.2015.09.012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7094483PMC
November 2015

CRABP1 is associated with a poor prognosis in breast cancer: adding to the complexity of breast cancer cell response to retinoic acid.

Mol Cancer 2015 Jul 5;14:129. Epub 2015 Jul 5.

Department of Oncology, University of Alberta, Cross Cancer Institute, 11560 University Avenue, Edmonton, T6G 1Z2, AB, Canada.

Background: Clinical trials designed to test the efficacy of retinoic acid (RA) as an adjuvant for the treatment of solid cancers have been disappointing, primarily due to RA resistance. Estrogen receptor (ER)-negative breast cancer cells are more resistant to RA than ER-positive cells. The expression and subcellular distribution of two RA-binding proteins, FABP5 and CRABP2, has already been shown to play critical roles in breast cancer cell response to RA. CRABP1, a third member of the RA-binding protein family, has not previously been investigated as a possible mediator of RA action in breast cancer.

Methods: CRABP1 and CRABP2 expression in primary breast tumor tissues was analyzed using gene expression and tissue microarrays. CRABP1 levels were manipulated using siRNAs and by transient overexpression. RA-induced subcellular translocation of CRABPs was examined by immunofluorescence microscopy and immunoblotting. RA-induced transactivation of RAR was analyzed using a RA response element (RARE)-driven luciferase reporter system. Effects of CRABP1 expression and RA treatment on downstream gene expression were investigated by semi-quantitative RT-PCR analysis.

Results: Compared to normal mammary tissues, CRABP1 expression is significantly down-regulated in ER+ breast tumors, but maintained in triple-negative breast cancers. Elevated CRABP1 levels are associated with poor patient prognosis, high Ki67 immunoreactivity and high tumor grade in breast cancer. The prognostic significance of CRABP1 is attributed to its cytoplasmic localization. We demonstrate that CRABP1 expression attenuates RA-induced cell growth arrest and inhibits RA signalling in breast cancer cells by sequestering RA in the cytoplasm. We also show that CRABP1 affects the expression of genes involved in RA biosynthesis, trafficking and metabolism.

Conclusions: CRABP1 is an adverse factor for clinical outcome in triple-negative breast cancer and a potent inhibitor of RA signalling in breast cancer cells. Our data indicate that CRABP1, in conjunction with previously identified CRABP2 and FABP5, plays a key role in breast cancer cell response to RA. We propose that these three RA-binding proteins can serve as biomarkers for predicting triple-negative breast cancer response to RA, with elevated levels of either cytoplasmic CRABP1 or FABP5 associated with RA resistance, and elevated levels of nuclear CRABP2 associated with sensitivity to RA.
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http://dx.doi.org/10.1186/s12943-015-0380-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4491424PMC
July 2015

Ddx1 knockout results in transgenerational wild-type lethality in mice.

Sci Rep 2015 Apr 24;5:9829. Epub 2015 Apr 24.

Department of Oncology, University of Alberta, Cross Cancer Institute, 11560 University Avenue, Edmonton, Alberta T6G 1Z2, Canada.

DEAD box 1 (DDX1) is a member of the DEAD box family of RNA helicases which are involved in all aspects of RNA metabolism. DDX1 has been implicated in a variety of biological processes, including 3'-end processing of mRNA, DNA repair, microRNA processing, tRNA maturation and mRNA transport. To study the role of DDX1 during development, we have generated mice carrying a constitutive Ddx1 knock-out allele. Ddx1(+/-) mice have no obvious phenotype and express similar levels of DDX1 as wild-type mice indicating compensation from the intact Ddx1 allele. Heterozygote matings produce no viable Ddx1(-/-) progeny, with Ddx1(-/-) embryos dying prior to embryonic day (E) 3.5. Intriguingly, the number of wild-type progeny is significantly decreased in heterozygote crosses, with two different heterozygote populations identified based on parental genotype: (i) normal Ddx1(+/-) mice which generate the expected number of wild-type progeny and (ii) Ddx1*(/-) mice (with * signifying a non-genetically altered allele) which generate a significantly reduced number of wild-type mice. The transgenerational inheritance of wild-type lethality observed upon crossing Ddx1*(/-) mice is independent of parental sex and occurs in cis through a mechanism that is different from other types of previously reported transgenerational epigenetic inheritance.
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http://dx.doi.org/10.1038/srep09829DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4408975PMC
April 2015

Aldehyde dehydrogenase 1A3 influences breast cancer progression via differential retinoic acid signaling.

Mol Oncol 2015 Jan 24;9(1):17-31. Epub 2014 Jul 24.

Department of Pathology, Dalhousie University, Halifax, NS, Canada; Department of Microbiology & Immunology, Dalhousie University, Halifax, NS, Canada. Electronic address:

Aldehyde dehydrogenase (ALDH) 1A enzymes produce retinoic acid (RA), a transcription induction molecule. To investigate if ALDH1A1 or ALDH1A3-mediated RA signaling has an active role in breast cancer tumorigenesis, we performed gene expression and tumor xenograft studies. Analysis of breast patient tumors revealed that high levels of ALDH1A3 correlated with expression of RA-inducible genes with retinoic acid response elements (RAREs), poorer patient survival and triple-negative breast cancers. This suggests a potential link between ALDH1A3 expression and RA signaling especially in aggressive and/or triple-negative breast cancers. In MDA-MB-231, MDA-MB-468 and MDA-MB-435 cells, ALDH1A3 and RA increased expression of RA-inducible genes. Interestingly, ALDH1A3 had opposing effects in tumor xenografts, increasing tumor growth and metastasis of MDA-MB-231 and MDA-MB-435 cells, but decreasing tumor growth of MDA-MB-468 cells. Exogenous RA replaced ALDH1A3 in inducing the same opposing tumor growth and metastasis effects, suggesting that ALDH1A3 mediates these effects by promoting RA signaling. Genome expression analysis revealed that ALDH1A3 induced largely divergent gene expression in MDA-MB-231 and MDA-MB-468 cells which likely resulted in the opposing tumor growth effects. Treatment with DNA methylation inhibitor 5-aza-2'deoxycytidine restored uniform RA-inducibility of RARE-containing HOXA1 and MUC4 in MDA-MB-231 and MDA-MB-468 cells, suggesting that differences in epigenetic modifications contribute to differential ALDH1A3/RA-induced gene expression in breast cancer. In summary, ALDH1A3 induces differential RA signaling in breast cancer cells which affects the rate of breast cancer progression.
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http://dx.doi.org/10.1016/j.molonc.2014.07.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5528683PMC
January 2015

Femtosecond laser-induced cell-cell surgical attachment.

Lasers Surg Med 2014 Apr 18;46(4):335-41. Epub 2014 Feb 18.

Department of Electrical and Computer Engineering, Ultrafast Photonics and Nano-Optics Laboratory, University of Alberta, Edmonton, AB, T6G 2V4, Canada.

Background And Objective: Laser-induced cell-cell surgical attachment using femtosecond laser pulses is reported.

Study Design/materials And Methods: We have demonstrated the ability to attach single cells using sub-10 femtosecond laser pulses, with 800 nm central wavelength delivered from a Ti:Sapphire laser. To check that the cells did not go through a cell-fusion process, a fluorescent dye Calcein AM was used to verify that the fluorescent dye did not migrate from a dyed cell to a non-dyed cell. The mechanical integrity of the attached joint was assessed using an optical tweezer.

Results: Attachment of cells was performed without the induction of cell-cell fusion, with attachment efficiency of 95%, and while preserving the cells' viability. Cell-cell attachment was achieved by delivery of one to two trains of femtosecond laser pulses lasting 15 ms each.

Conclusions: Laser-induced ionization process led to an ultrafast reversible destabilization of the phospholipid layer of the cellular membrane. The inner cell membrane remained intact during the attachment procedure, and isolation of the cells' cytoplasm from the surrounding medium was obtained. A strong physical attachment between the cells was obtained due to the bonding of the membranes' ionized phospholipid molecules and the formation of a joint cellular membrane at the connection point. The cellular attachment technique, femtosecond laser-induced cell-cell surgical attachment, can potentially provide a platform for the creation of engineered tissue and cell cultures.
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http://dx.doi.org/10.1002/lsm.22230DOI Listing
April 2014
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