Protein trafficking through the endosomal system prepares intracellular parasites for a home invasion

Toxoplasma (toxoplasmosis) and Plasmodium (malaria) use unique secretory organelles for migration, cell invasion, manipulation of host cell functions, and cell egress. In particular, the apical secretory micronemes and rhoptries of apicomplexan parasites are essential for successful host infection. New findings reveal that the contents of these organelles, which are transported through the endoplasmic reticulum (ER) and Golgi, also require the parasite endosome-like system to access their respective organelles. In this review, we discuss recent findings that demonstrate that these parasites reduced their endosomal system and modified classical regulators of this pathway for the biogenesis of apical organelles

The role of clathrin in post-golgi trafficking in toxoplasma gondii

Apicomplexan parasites are single eukaryotic cells with a highly polarised secretory system that contains unique secretory organelles (micronemes and rhoptries) that are required for host cell invasion. In contrast, the role of the endosomal system is poorly understood in these parasites. With many typical endocytic factors missing, we speculated that endocytosis depends exclusively on a clathrin-mediated mechanism. Intriguingly, in Toxoplasma gondii we were only able to observe the endogenous clathrin heavy chain 1 (CHC1) at the Golgi, but not at the parasite surface. For the functional characterisation of Toxoplasma gondii CHC1 we generated parasite mutants conditionally expressing the dominant negative clathrin Hub fragment and demonstrate that CHC1 is essential for vesicle formation at the trans-Golgi network. Consequently, the functional ablation of CHC1 results in Golgi aberrations, a block in the biogenesis of the unique secretory microneme and rhoptry organelles, and of the pellicle. However, we found no morphological evidence for clathrin mediating endocytosis in these parasites and speculate that they remodelled their vesicular trafficking system to adapt to an intracellular lifestyle

Loss of heterozygosity of TRIM3 in malignant gliomas

<p>Abstract</p> <p>Background</p> <p>Malignant gliomas are frequent primary brain tumors associated with poor prognosis and very limited response to conventional chemo- and radio-therapies. Besides sharing common growth features with other types of solid tumors, gliomas are highly invasive into adjacent brain tissue, which renders them particularly aggressive and their surgical resection inefficient. Therefore, insights into glioma formation are of fundamental interest in order to provide novel molecular targets for diagnostic purposes and potential anti-cancer drugs. Human <it>Tripartite motif protein 3 </it>(<it>TRIM3</it>) encodes a structural homolog of <it>Drosophila brain tumor </it>(<it>brat</it>) implicated in progenitor cell proliferation control and cancer stem cell suppression. <it>TRIM3 </it>is located within the loss of allelic heterozygosity (LOH) hotspot of chromosome segment 11p15.5, indicating a potential role in tumor suppression. ...</p> <p>Methods</p> <p>Here we analyze 70 primary human gliomas of all types and grades and report somatic deletion mapping as well as single nucleotide polymorphism analysis together with quantitative real-time PCR of chromosome segment 11p15.5.</p> <p>Results</p> <p>Our analysis identifies LOH in 17 cases (24%) of primary human glioma which defines a common 130 kb-wide interval within the <it>TRIM3 </it>locus as a minimal area of loss. We further detect altered genomic dosage of <it>TRIM3 </it>in two glioma cases with LOH at 11p15.5, indicating homozygous deletions of <it>TRIM3</it>.</p> <p>Conclusion</p> <p>Loss of heterozygosity of chromosome segment 11p15.5 in malignant gliomas suggests <it>TRIM3 </it>as a candidate brain tumor suppressor gene.</p

Age determines the prognostic role of the cancer stem cell marker aldehyde dehydrogenase-1 in breast cancer

<p>Abstract</p> <p>Background</p> <p>The purpose of this study was to compare the expression and the prognostic effect of the breast cancer stem cell marker aldehyde dehydrogenase-1 (ALDH1) in young and elderly breast cancer patients.</p> <p>Methods</p> <p>The study population (N = 574) consisted of all early breast cancer patients primarily treated with surgery in our center between 1985 and 1994. Median follow-up was 17.9 years (range: 0.1 to 23.5). Tissue microarray slides were immunohistochemically stained for ALDH1 expression and quantified by two independent observers who were blinded to clinical outcome. Assessment of the prognostic effect of ALDH1 expression was stratified according to age and systemic treatment.</p> <p>Results</p> <p>Complete lack of expression of ALDH1 was found in 40% of tumors. With increasing age more tumors showed complete absence of ALDH1 expression (<it>P </it>< .001). In patients aged > 65 years, ALDH1 status was not associated with any clinical outcome. Conversely, in patients aged < 65 years, ALDH1 positivity was an independent risk factor of worse outcome for relapse free period (hazard ratio = 1.71 (95% CI, 1.09 to 2.68); <it>P </it>= .021) and relative survival (relative excess risks of death = 2.36 (95% CI, 1.22 to 3.68); <it>P </it>= .016). Ten-year relative survival risk was 57% in ALDH1-positive patients compared to 83% in ALDH1-negative patients.</p> <p>Conclusion</p> <p>ALDH1 expression and its prognostic effect are age-dependent. Our results support the hypothesis that breast cancer biology is different in elderly patients compared to their younger counterparts and emphasizes the importance of taking into consideration age-specific interactions in breast cancer research.</p

Gene duplications and evolution of vertebrate voltage-gated sodium channels

Author Posting. © The Author(s), 2006. This is the author's version of the work. It is posted here by permission of Springer for personal use, not for redistribution. The definitive version was published in Journal of Molecular Evolution 63 (2006): 208-221, doi:10.1007/s00239-005-0287-9.Voltage-gated sodium channels underlie action potential generation in excitable tissue. To establish the evolutionary mechanisms that shaped the vertebrate sodium channel a-subunit (SCNA) gene family and their encoded Nav1 proteins, we identified all SCNA genes in several teleost species. Molecular cloning revealed that teleosts have eight SCNA genes, comparable to the number in another vertebrate lineage, mammals. Prior phylogenetic analyses had indicated that teleosts and tetrapods share four monophyletic groups of SCNA genes and that tandem duplications selectively expanded the number of genes in two of the four mammalian groups. However, the number of genes in each group varies between teleosts and tetrapods suggesting different evolutionary histories in the two vertebrate lineages. Our findings from phylogenetic analysis and chromosomal mapping of Danio rerio genes indicate that tandem duplications are an unlikely mechanism for generation of the extant teleost SCNA genes. Instead, analysis of other closely mapped genes in D. rerio supports the hypothesis that a whole genome duplication was involved in expansion of the SCNA gene family in teleosts. Interestingly, despite their different evolutionary histories, mRNA analyses demonstrated a conservation of expression patterns for SCNA orthologues in teleosts and tetrapods, suggesting functional conservation.The authors’ work was supported by NIH grants (NS 38937; AEN, ADT and ABR, NS 25513; HHZ and YL and NSF IBN 0236147; MCJ)

Identification of Novel Proteins in Neospora caninum Using an Organelle Purification and Monoclonal Antibody Approach

Neospora caninum is an important veterinary pathogen that causes abortion in cattle and neuromuscular disease in dogs. Neospora has also generated substantial interest because it is an extremely close relative of the human pathogen Toxoplasma gondii, yet does not appear to infect humans. While for Toxoplasma there are a wide array of molecular tools and reagents available for experimental investigation, relatively few reagents exist for Neospora. To investigate the unique biological features of this parasite and exploit the recent sequencing of its genome, we have used an organelle isolation and monoclonal antibody approach to identify novel organellar proteins and develop a wide array of probes for subcellular localization. We raised a panel of forty-six monoclonal antibodies that detect proteins from the rhoptries, micronemes, dense granules, inner membrane complex, apicoplast, mitochondrion and parasite surface. A subset of the proteins was identified by immunoprecipitation and mass spectrometry and reveal that we have identified and localized many of the key proteins involved in invasion and host interaction in Neospora. In addition, we identified novel secretory proteins not previously studied in any apicomplexan parasite. Thus, this organellar monoclonal antibody approach not only greatly enhances the tools available for Neospora cell biology, but also identifies novel components of the unique biological characteristics of this important veterinary pathogen

CD133, CD15/SSEA-1, CD34 or side populations do not resume tumor-initiating properties of long-term cultured cancer stem cells from human malignant glio-neuronal tumors

<p>Abstract</p> <p>Background</p> <p>Tumor initiating cells (TICs) provide a new paradigm for developing original therapeutic strategies.</p> <p>Methods</p> <p>We screened for TICs in 47 human adult brain malignant tumors. Cells forming floating spheres in culture, and endowed with all of the features expected from tumor cells with stem-like properties were obtained from glioblastomas, medulloblastoma but not oligodendrogliomas.</p> <p>Results</p> <p>A long-term self-renewal capacity was particularly observed for cells of malignant glio-neuronal tumors (MGNTs). Cell sorting, karyotyping and proteomic analysis demonstrated cell stability throughout prolonged passages. Xenografts of fewer than 500 cells in Nude mouse brains induced a progressively growing tumor. CD133, CD15/LeX/Ssea-1, CD34 expressions, or exclusion of Hoechst dye occurred in subsets of cells forming spheres, but was not predictive of their capacity to form secondary spheres or tumors, or to resist high doses of temozolomide.</p> <p>Conclusions</p> <p>Our results further highlight the specificity of a subset of high-grade gliomas, MGNT. TICs derived from these tumors represent a new tool to screen for innovative therapies.</p

The measurement and therapeutic implications of circulating tumour cells in breast cancer

Circulating tumours cells (CTCs) represent an important biologic link in the spread of breast cancer from primary to metastatic disease. CTCs are strong predictors of prognosis in patients with metastatic breast cancer. Research to date has focused on development of methods with adequate sensitivity and specificity to reproducibly identify these rare events. Future research will focus on the biologic phenotypes of these cells with goals to understand mechanisms of metastasis, to identify novel therapeutic targets, and to monitor response to therapy

CD146 expression is associated with a poor prognosis in human breast tumors and with enhanced motility in breast cancer cell lines.

INTRODUCTION: Metastasis is a complex process involving loss of adhesion, migration, invasion and proliferation of cancer cells. Cell adhesion molecules play a pivotal role in this phenomenon by regulating cell-cell and cell-matrix interactions. CD146 (MCAM) is associated with an advanced tumor stage in melanoma, prostate cancer and ovarian cancer. Studies of CD146 expression and function in breast cancer remain scarce except for a report concluding that CD146 could act as a tumor suppressor in breast carcinogenesis. METHODS: To resolve these apparent discrepancies in the role of CD146 in tumor cells, we looked at the association of CD146 expression with histoclinical features in human primary breast cancers using DNA and tissue microarrays. By flow cytometry, we characterized CD146 expression on different breast cancer cell lines. Using siRNA or shRNA technology, we studied functional consequences of CD146 downmodulation of MDA-MB-231 cells in migration assays. Wild-type, mock-transfected and downmodulated transfected cells were profiled using whole-genome DNA microarrays to identify genes whose expression was modified by CD146 downregulation. RESULTS: Microarray studies revealed the association of higher levels of CD146 with histoclinical features that belong to the basal cluster of human tumors. Expression of CD146 protein on epithelial cells was detected in a small subset of cancers with histoclinical features of basal tumors. CD146+ cell lines displayed a mesenchymal phenotype. Downmodulation of CD146 expression in the MDA-MB-231 cell line resulted in downmodulation of vimentin, as well as of a set of genes that include both genes associated with a poor prognosis in a variety of cancers and genes known to promote cell motility. In vitro functional assays revealed decreased migration abilities associated with decreased CD146 expression. CONCLUSIONS: In addition to its expression in the vascular compartment, CD146 is expressed on a subset of epithelial cells in malignant breast. CD146 may directly or indirectly contribute to tumor aggressiveness by promoting malignant cell motility. Changes in molecular signatures following downmodulation of CD146 expression suggest that CD146 downmodulation is associated with the reversal of several biological characteristics associated with epithelial to mesenchymal transition, and the phenomenon associated with the metastatic process.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are

Mesenchymal Stem Cells Promote Mammosphere Formation and Decrease E-Cadherin in Normal and Malignant Breast Cells

Normal and malignant breast tissue contains a rare population of multi-potent cells with the capacity to self-renew, referred to as stem cells, or tumor initiating cells (TIC). These cells can be enriched by growth as "mammospheres" in three-dimensional cultures.We tested the hypothesis that human bone-marrow derived mesenchymal stem cells (MSC), which are known to support tumor growth and metastasis, increase mammosphere formation.We found that MSC increased human mammary epithelial cell (HMEC) mammosphere formation in a dose-dependent manner. A similar increase in sphere formation was seen in human inflammatory (SUM149) and non-inflammatory breast cancer cell lines (MCF-7) but not in primary inflammatory breast cancer cells (MDA-IBC-3). We determined that increased mammosphere formation can be mediated by secreted factors as MSC conditioned media from MSC spheroids significantly increased HMEC, MCF-7 and SUM149 mammosphere formation by 6.4 to 21-fold. Mammospheres grown in MSC conditioned media had lower levels of the cell adhesion protein, E-cadherin, and increased expression of N-cadherin in SUM149 and HMEC cells, characteristic of a pro-invasive mesenchymal phenotype. Co-injection with MSC in vivo resulted in a reduced latency time to develop detectable MCF-7 and MDA-IBC-3 tumors and increased the growth of MDA-IBC-3 tumors. Furthermore, E-cadherin expression was decreased in MDA-IBC-3 xenografts with co-injection of MSC.MSC increase the efficiency of primary mammosphere formation in normal and malignant breast cells and decrease E-cadherin expression, a biologic event associated with breast cancer progression and resistance to therapy