Quantitative phosphotyrosine proteomics of EphB2 signaling by stable isotope labeling with amino acids in cell culture (SILAC)

Eph-related receptor tyrosine kinases (RTK) have been implicated in several biological functions including synaptic plasticity, axon guidance, and morphogenesis, yet the details of the signal transduction pathways that produce these specific biological functions after ligand-receptor interaction remain unclear. We used Stable Isotope Labeling by Amino Acids in Cell Culture (SILAC) in combination with LC-MS/MS to characterize cellular signaling following stimulation by ephrinB1-Fc of NG-108 cells that overexpress EphB2 receptors. Because tyrosine phosphorylation functions as a key regulatory event in RTK signaling, we used anti-phosphotyrosine immunoprecipitation (pY IP) of cell lysates to isolate potential participants in the EphB2 pathway. Our SILAC experiments identified 127 unique proteins, 40 of which demonstrated increased abundance in pY IPs from ephrinB1-Fc stimulated cells as compared with unstimulated cells. Six proteins demonstrated decreased abundance, and 81 did not change significantly in relative abundance. Western blotting analysis of five proteins after pY IP verified their SILAC results. On the basis of previously published work and use of PathwayAssist software, we proposed an interaction network downstream of EphB2 for the proteins with changed ratios.     

Using binding profiles to predict binding sites of target RNAs

Prediction of RNA-RNA interaction is a key to elucidating possible functions of small non-coding RNAs, and a number of computational methods have been proposed to analyze interacting RNA secondary structures. In this article, we focus on predicting binding sites of target RNAs that are expected to interact with regulatory antisense RNAs in a general form of interaction. For this purpose, we propose bistaRNA, a novel method for predicting multiple binding sites of target RNAs. bistaRNA employs binding profiles that represent scores for hybridized structures, leading to reducing the computational cost for interaction prediction. bistaRNA considers an ensemble of equilibrium interacting structures and seeks to maximize expected accuracy using dynamic programming. Experimental results on real interaction data validate good accuracy and fast computation time of bistaRNA as compared with several competitive methods. Moreover, we aim to find new targets given specific antisense RNAs, which provides interesting insights into antisense RNA regulation. bistaRNA is implemented in C++. The program and Supplementary Material are available at http://rna.naist.jp/program/bistarna/.     

Higher plant tetrapyrrols: Their biosynthesis and its regulation,tetrapyrrol role in transmission of retrograde signals

This review summarizes the current knowledge of the biosynthesis of tetrapyrroles in plants, key stages of this process regulation, as well as the available data on the participation of these compounds in the induction and transmission of retrograde signals. Possible sources of retrograde signals are considered. Particular attention is paid to the analysis of data on the signaling molecules that induce retrograde regulation. The interaction of retrograde signals with other cells signaling pathways is emphasized.     

Global Versus Local Centrality in Evolution of Yeast Protein Network

It is shown here that in the yeast protein interaction network the global centrality measure (betweenness) depends on the protein evolutionary age (i.e., on historic contingency) more weakly than the local centrality measure (degree). This phenomenon is not observed in mutational duplication-and-divergence models. The network domains responsible for this difference deal with DNA/RNA information processing, regulation, and cell cycle. A selection vector can operate in these domains, which integrates the network activity and thus compensates for the process of mutational divergence. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Intrinsically disordered proteins: regulation and disease

Intrinsically disordered proteins (IDPs) are enriched in signaling and regulatory functions because disordered segments permit interaction with several proteins and hence the re-use of the same protein in multiple pathways. Understanding IDP regulation is important because altered expression of IDPs is associated with many diseases. Recent studies show that IDPs are tightly regulated and that dosage-sensitive genes encode proteins with disordered segments. The tight regulation of IDPs may contribute to signaling fidelity by ensuring that IDPs are available in appropriate amounts and not present longer than needed. The altered availability of IDPs may result in sequestration of proteins through non-functional interactions involving disordered segments (i.e., molecular titration), thereby causing an imbalance in signaling pathways. We discuss the regulation of IDPs, address implications for signaling, disease and drug development, and outline directions for future research.     

cLoops2: a full-stack comprehensive analytical tool for chromatin interactions

Investigating chromatin interactions between regulatory regions such as enhancer and promoter elements is vital for understanding the regulation of gene expression. Compared to Hi-C and its variants, the emerging 3D mapping technologies focusing on enriched signals, such as TrAC-looping, reduce the sequencing cost and provide higher interaction resolution for cis-regulatory elements. A robust pipeline is needed for the comprehensive interpretation of these data, especially for loop-centric analysis. Therefore, we have developed a new versatile tool named cLoops2 for the full-stack analysis of these 3D chromatin interaction data. cLoops2 consists of core modules for peak-calling, loop-calling, differentially enriched loops calling and loops annotation. It also contains multiple modules for interaction resolution estimation, data similarity estimation, features quantification, feature aggregation analysis, and visualization. cLoops2 with documentation and example data are open source and freely available at GitHub: https://github.com/KejiZhaoLab/cLoops2.     

Current views on structure and function of alpha2-antiplasmin

Data available in literature on basic, primary, rapidly acting alpha 2-antiplasmin (a plasmin inhibitor) are generalized. Methods of its isolation from human blood plasma, properties, structure, kinetics of interaction with plasmin as well as the key role in the fibrinolysis regulation are considered.     

Mitochondrial content is central to nuclear gene expression: Profound implications for human health

We review a recent paper in Genome Research by Guantes et al. showing that nuclear gene expression is influenced by the bioenergetic status of the mitochondria. The amount of energy that mitochondria make available for gene expression varies considerably. It depends on: the energetic demands of the tissue; the mitochondrial DNA (mtDNA) mutant load; the number of mitochondria; stressors present in the cell. Hence, when failing mitochondria place the cell in energy crisis there are major effects on gene expression affecting the risk of degenerative diseases, cancer and ageing. In 2015 the UK parliament approved a change in the regulation of IVF techniques, allowing “Mitochondrial replacement therapy” to become a reproductive choice for women at risk of transmitting mitochondrial disease to their children. This is the first time that this technique will be available. Therefore understanding the interaction between mitochondria and the nucleus has never been more important.     

Modulation of the dopamine transporter by interaction with Secretory Carrier Membrane Protein 2

The monoamine transporters for dopamine (DAT), norepinephrine (NET) and serotonin (SERT) facilitate the homeostatic balance of neurotransmitters in the synaptic cleft and thus, play a fundamental role in regulating neuronal activity. Despite the importance of these monoamine transporters in controlling brain function, only relatively little information is available regarding the cellular and molecular regulation of these proteins. The monoamine transporters have been found to associate with a number of different proteins that regulate the function and subcellular localization of the transporters. We recently reported a functional interaction between SERT and the Secretory Carrier Membrane Protein 2 (SCAMP2). Here, we demonstrate that SCAMP2 also plays a role in the functional regulation of DAT. DAT and SCAMP2 interaction is here verified by co-immunoprecipitation and fluorescence resonance energy transfer (FRET) microscopy. Moreover, co-expression of DAT and SCAMP2 results in a decrease in DAT-mediated dopamine uptake caused by reduced levels of DAT molecules on the cell surface. Our finding that SCAMP2 interacts with and regulates the subcellular distribution of both DAT and SERT suggests that interaction with SCAMP2 may constitute an important mechanism for coordinating cell surface expression of monoamine transporters.     

A model of troponin-I in complex with troponin-C using hybrid experimental data: the inhibitory region is a beta-hairpin

We present a model for the skeletal muscle troponin-C (TnC)/troponin-I (TnI) interaction, a critical molecular switch that is responsible for calcium-dependent regulation of the contractile mechanism. Despite concerted efforts by multiple groups for more than a decade, attempts to crystallize troponin-C in complex with troponin-I, or in the ternary troponin-complex, have not yet delivered a high-resolution structure. Many groups have pursued different experimental strategies, such as X-ray crystallography, NMR, small-angle scattering, chemical cross-linking, and fluorescent resonance energy transfer (FRET) to gain insights into the nature of the TnC/TnI interaction. We have integrated the results of these experiments to develop a model of the TnC/TnI interaction, using an atomic model of TnC as a scaffold. The TnI sequence was fit to each of two alternate neutron scattering envelopes: one that winds about TnC in a left-handed sense (Model L), and another that winds about TnC in a right-handed sense (Model R). Information from crystallography and NMR experiments was used to define segments of the models. Tests show that both models are consistent with available cross-linking and FRET data. The inhibitory region TnI(95-114) is modeled as a flexible beta-hairpin, and in both models it is localized to the same region on the central helix of TnC. The sequence of the inhibitory region is similar to that of a beta-hairpin region of the actin-binding protein profilin. This similarity supports our model and suggests the possibility of using an available profilin/actin crystal structure to model the TnI/actin interaction. We propose that the beta-hairpin is an important structural motif that communicates the Ca2+-activated troponin regulatory signal to actin.     

Simulation of daytime vigilance by the additive interaction of a homeostatic and a circadian process

The two-process model of sleep regulation postulates that a homeostatic and a circadian process underlie sleep regulation. The timing of sleep and waking is accounted for by the interaction of these two processes. The assumptions of two separate processes or of a single process resulting from their additive interaction are mathematically equivalent but conceptually different. Based on an additive interaction, subjective alertness ratings in a forced desynchrony protocol and subjective sleepiness ratings in a photoperiod experiment were simulated. The correspondence between empirical and simulated data supports the basic assumption of the model. Received: 7 October 1993/Accepted in revised form: 22 January 1994     

Adenylyl cyclase-associated protein 1: Structure,regulation, and participation in cellular processes

This review summarizes information available to date about the structural organization, regulation of functional activity of adenylyl cyclase-associated protein 1 (CAP1), and its participation in cellular processes. Numerous data are generalized on the role of CAP1 in the regulation of actin cytoskeleton and its interactions with many actin-binding proteins. Attention is drawn to the similarity of the structure of CAP1 and its contribution to the remodeling of actin filaments in prokaryotes and eukaryotes, as well as to the difference in the interaction of CAP1 with adenylyl cyclase in these cells. In addition, we discuss the participation of CAP1 in various pathological processes.     

CGI: a new approach for prioritizing genes by combining gene expression and protein-protein interaction data

MOTIVATION: Identifying candidate genes associated with a given phenotype or trait is an important problem in biological and biomedical studies. Prioritizing genes based on the accumulated information from several data sources is of fundamental importance. Several integrative methods have been developed when a set of candidate genes for the phenotype is available. However, how to prioritize genes for phenotypes when no candidates are available is still a challenging problem. RESULTS: We develop a new method for prioritizing genes associated with a phenotype by Combining Gene expression and protein Interaction data (CGI). The method is applied to yeast gene expression data sets in combination with protein interaction data sets of varying reliability. We found that our method outperforms the intuitive prioritizing method of using either gene expression data or protein interaction data only and a recent gene ranking algorithm GeneRank. We then apply our method to prioritize genes for Alzheimer's disease. AVAILABILITY: The code in this paper is available upon request.