JoVE Visualize What is visualize?
Stop Reading. Start Watching.
Advanced Search
Stop Reading. Start Watching.
Regular Search
Find video protocols related to scientific articles indexed in Pubmed.
A network-based method to assess the statistical significance of mild co-regulation effects.
PLoS ONE
PUBLISHED: 01-01-2013
Show Abstract
Hide Abstract
Recent development of high-throughput, multiplexing technology has initiated projects that systematically investigate interactions between two types of components in biological networks, for instance transcription factors and promoter sequences, or microRNAs (miRNAs) and mRNAs. In terms of network biology, such screening approaches primarily attempt to elucidate relations between biological components of two distinct types, which can be represented as edges between nodes in a bipartite graph. However, it is often desirable not only to determine regulatory relationships between nodes of different types, but also to understand the connection patterns of nodes of the same type. Especially interesting is the co-occurrence of two nodes of the same type, i.e., the number of their common neighbours, which current high-throughput screening analysis fails to address. The co-occurrence gives the number of circumstances under which both of the biological components are influenced in the same way. Here we present SICORE, a novel network-based method to detect pairs of nodes with a statistically significant co-occurrence. We first show the stability of the proposed method on artificial data sets: when randomly adding and deleting observations we obtain reliable results even with noise exceeding the expected level in large-scale experiments. Subsequently, we illustrate the viability of the method based on the analysis of a proteomic screening data set to reveal regulatory patterns of human microRNAs targeting proteins in the EGFR-driven cell cycle signalling system. Since statistically significant co-occurrence may indicate functional synergy and the mechanisms underlying canalization, and thus hold promise in drug target identification and therapeutic development, we provide a platform-independent implementation of SICORE with a graphical user interface as a novel tool in the arsenal of high-throughput screening analysis.
Related JoVE Video
MicroRNA-200c represses migration and invasion of breast cancer cells by targeting actin-regulatory proteins FHOD1 and PPM1F.
Mol. Cell. Biol.
PUBLISHED: 12-05-2011
Show Abstract
Hide Abstract
MicroRNA-200c (miR-200c) has been shown to suppress epithelial-mesenchymal transition (EMT), which is attributed mainly to targeting of ZEB1/ZEB2, repressors of the cell-cell contact protein E-cadherin. Here we demonstrated that modulation of miR-200c in breast cancer cells regulates cell migration, cell elongation, and transforming growth factor ? (TGF-?)-induced stress fiber formation by impacting the reorganization of cytoskeleton that is independent of the ZEB/E-cadherin axis. We identified FHOD1 and PPM1F, direct regulators of the actin cytoskeleton, as novel targets of miR-200c. Remarkably, expression levels of FHOD1 and PPM1F were inversely correlated with the level of miR-200c in breast cancer cell lines, breast cancer patient samples, and 58 cancer cell lines of various origins. Furthermore, individual knockdown/overexpression of these target genes phenocopied the effects of miR-200c overexpression/inhibition on cell elongation, stress fiber formation, migration, and invasion. Mechanistically, targeting of FHOD1 by miR-200c resulted in decreased expression and transcriptional activity of serum response factor (SRF), mediated by interference with the translocation of the SRF coactivator mycocardin-related transcription factor A (MRTF-A). This finally led to downregulation of the expression and phosphorylation of the SRF target myosin light chain 2 (MLC2) gene, required for stress fiber formation and contractility. Thus, miR-200c impacts on metastasis by regulating several EMT-related processes, including a novel mechanism involving the direct targeting of actin-regulatory proteins.
Related JoVE Video
Utilization of RNAi to validate antibodies for reverse phase protein arrays.
Methods Mol. Biol.
PUBLISHED: 09-09-2011
Show Abstract
Hide Abstract
Reverse phase protein arrays (RPPAs) emerged as a very useful tool for high-throughput screening of protein expression in large numbers of small specimen. Similar to other protein chemistry methods, antibody specificity is also a major concern for RPPA. Currently, testing antibodies on Western blot for specificity and applying serial dilution curves to determine signal/concentration linearity of RPPA signals are most commonly employed to validate antibodies for RPPA applications. However, even the detection antibodies fulfilling both requirements do not always give the expected result. Chemically synthesized small interfering RNAs (siRNAs) are one of the most promising and time-efficient tools for loss-of-function studies by specifically targeting the gene of interest resulting in a reduction at the protein expression level, and are therefore used to dissect biological processes. Here, we report the utilization of siRNA-treated sample lysates for the quantification of a protein of interest as a useful and reliable tool to validate antibody specificity for RPPAs. As our results indicate, we recommend the use of antibodies which give the highest dynamic range between the control siRNA-treated samples and the target protein (here: EGFR) siRNA-treated ones on RPPAs, to be able to quantify even small differences of protein abundance with high confidence.
Related JoVE Video
RNAi-based validation of antibodies for reverse phase protein arrays.
Proteome Sci
PUBLISHED: 10-10-2010
Show Abstract
Hide Abstract
Reverse phase protein arrays (RPPA) have been demonstrated to be a useful experimental platform for quantitative protein profiling in a high-throughput format. Target protein detection relies on the readout obtained from a single detection antibody. For this reason, antibody specificity is a key factor for RPPA. RNAi allows the specific knockdown of a target protein in complex samples and was therefore examined for its utility to assess antibody performance for RPPA applications.
Related JoVE Video
Arctic-alpine distributions--metapopulations on a continental scale?
Am. Nat.
PUBLISHED: 02-10-2009
Show Abstract
Hide Abstract
Cold-adapted species in the Northern Hemisphere frequently show arctic-alpine discontinuous ranges at high latitudes and on mountains farther south, but area connectivity through current and historical gene flow remains unclear. We used the coalescent-based program IMa (Isolation with Migration-analytic) to test for migration among disjunct European areas of arctic-alpine wolf spiders of the Pardosa saltuaria group. Mitochondrial (ND1) and nuclear (ITS1, 5.8S rDNA, ITS2) markers were analyzed simultaneously. Gene flow was unidirectional from Scandinavia to the Alps and the Carpathians, complex with respect to intermediate relict areas in central Europe, and very limited in outlying areas in the Balkans and Pyrenees. Population connectivity may have been greater during glacial events that might alternatively account for the inferred arctic-alpine links. A simulation study under various demographic histories (using a new module in the Mesquite package, which models episodic migration) showed that the empirical results are equally consistent with moderate levels of ongoing (continuous) migration or, alternatively, with strong migration bursts at the last glacial maximum but not at earlier times. Habitat connectivity was probably maximal during glacial events, illustrating the potential influence of ecology and life history on organismal responses to past climatic change.
Related JoVE Video
Global microRNA level regulation of EGFR-driven cell-cycle protein network in breast cancer.
Mol. Syst. Biol.
Show Abstract
Hide Abstract
The EGFR-driven cell-cycle pathway has been extensively studied due to its pivotal role in breast cancer proliferation and pathogenesis. Although several studies reported regulation of individual pathway components by microRNAs (miRNAs), little is known about how miRNAs coordinate the EGFR protein network on a global miRNA (miRNome) level. Here, we combined a large-scale miRNA screening approach with a high-throughput proteomic readout and network-based data analysis to identify which miRNAs are involved, and to uncover potential regulatory patterns. Our results indicated that the regulation of proteins by miRNAs is dominated by the nucleotide matching mechanism between seed sequences of the miRNAs and 3-UTR of target genes. Furthermore, the novel network-analysis methodology we developed implied the existence of consistent intrinsic regulatory patterns where miRNAs simultaneously co-regulate several proteins acting in the same functional module. Finally, our approach led us to identify and validate three miRNAs (miR-124, miR-147 and miR-193a-3p) as novel tumor suppressors that co-target EGFR-driven cell-cycle network proteins and inhibit cell-cycle progression and proliferation in breast cancer.
Related JoVE Video
Collective Network of Binary Classifier Framework for Polarimetric SAR Image Classification: An Evolutionary Approach.
IEEE Trans Syst Man Cybern B Cybern
Show Abstract
Hide Abstract
Terrain classification over polarimetric synthetic aperture radar (SAR) images has been an active research field where several features and classifiers have been proposed up to date. However, some key questions, e.g., 1) how to select certain features so as to achieve highest discrimination over certain classes?, 2) how to combine them in the most effective way?, 3) which distance metric to apply?, 4) how to find the optimal classifier configuration for the classification problem in hand?, 5) how to scale/adapt the classifier if large number of classes/features are present?, and finally, 6) how to train the classifier efficiently to maximize the classification accuracy?, still remain unanswered. In this paper, we propose a collective network of (evolutionary) binary classifier (CNBC) framework to address all these problems and to achieve high classification performance. The CNBC framework adapts a "Divide and Conquer" type approach by allocating several NBCs to discriminate each class and performs evolutionary search to find the optimal BC in each NBC. In such an (incremental) evolution session, the CNBC body can further dynamically adapt itself with each new incoming class/feature set without a full-scale retraining or reconfiguration. Both visual and numerical performance evaluations of the proposed framework over two benchmark SAR images demonstrate its superiority and a significant performance gap against several major classifiers in this field.
Related JoVE Video

What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

How does it work?

We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

Video X seems to be unrelated to Abstract Y...

In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.