Potential candidate biomarkers associated with osteoarthritis: Evidence from a comprehensive network and pathway analysis

Osteoarthritis (OA) is one of the most common forms of arthritis world widely. Some key genes and diagnostic markers have been reported due to the development of modern molecular biology technologies. However, the etiology and pathogenesis of OA remains unknown. In this study, an integrated network and pathway analysis towards the biological function of OA-associated genes was conducted to provide valuable information to further explore the etiology and pathogenesis of OA. A total of 2,548 genes which reported a statistically significant association with OA were screened. An integrated network and pathway analysis was performed to identify the pathways and genes most associated to OA. Moreover, OA-specific protein–protein interaction (PPI) network was constructed by cytocluster based on the Molecular Complex Detection Algorithm (MCODE) to screen its candidate biomarkers. Quantitative real-time polymerase chain reaction was used to confirm the expression levels and to validate the results of MCODE cluster analysis by six genes. The pathway networks suggested that extracellular matrix (ECM) organization, collagen degradation and collagen formation showed important associations with OA. In top two PPI clusters, 61 of the OA-associated genes were included in the OA-specific PPI network, which also included 23 candidate genes that are likely to be highly associated with OA based on MCODE clusters. Analysis of mRNA showed that the expression levels of COL9A1, COL9A2, ITGA3, COL9A3, ITGA2, and LAMA1 in the peripheral blood mononuclear cells of OA patients were significantly lower than those of the normal controls (p<0.005). To our knowledge, this is the first comprehensive and systematic report based on OA-related genes demonstrating that the functional destruction of collagen in cartilage may be a very important contributing factor to OA. Quantitative detection of collagen synthesis may be of great help in early identification and prediction of OA. Maintaining the quality and quantity of collagen can be a potential target for clinical treatment of OA in the future practice.     

自适应蚁群算法在DNA中的应用

主要讨论了自适应蚁群算法在DNA序列比对中的应用，主要的过程是：首先，我们设一个计分函数和一个得分策略，在任意给出一对DNA序列，建立一个序列比对矩阵．现由4只蚂蚁从左上角向右下角移动，并且最终到达右下角，那么这4只蚂蚁随意走出4条路径，根据4条路径得出4对等长的比对，再依照计分函数分别计算出4条路径的比对得分，再由1．3式进一步验证4条路径的平均得分值，取其中得分最高（即最优路径）路径；进行第二次信息素增量的调整，方法是根据蚂蚁所走过的方向和该方向上得分比例计算出来的，信息素的变化量利用矩阵来存储，那么下一次蚂蚁所选的路径就要根据以前在各条路径上的信息素浓度总和的大小选择移动方向，最终经过有限次迭代，蚂蚁就会找到一条最优路径，也就是一条与原来DNA最相似的DNA链．     

A Cargo-centered Perspective on the PEX5 Receptor-mediated Peroxisomal Protein Import Pathway

Peroxisomal matrix proteins are synthesized on cytosolic ribosomes and post-translationally targeted to the organelle by PEX5, the peroxisomal shuttling receptor. The pathway followed by PEX5 during this process is known with reasonable detail. After recognizing cargo proteins in the cytosol, the receptor interacts with the peroxisomal docking/translocation machinery, where it gets inserted; PEX5 is then monoubiquitinated, extracted back to the cytosol and, finally, deubiquitinated. However, despite this information, the exact step of this pathway where cargo proteins are translocated across the organelle membrane is still ill-defined. In this work, we used an in vitro import system to characterize the translocation mechanism of a matrix protein possessing a type 1 targeting signal. Our results suggest that translocation of proteins across the organelle membrane occurs downstream of a reversible docking step and upstream of the first cytosolic ATP-dependent step (i.e. before ubiquitination of PEX5), concomitantly with the insertion of the receptor into the docking/translocation machinery.     

HIV-1 Nef Binds a Subpopulation of MHC-I throughout Its Trafficking Itinerary and Down-regulates MHC-I by Perturbing Both Anterograde and Retrograde Trafficking

The HIV protein Nef is thought to mediate immune evasion and promote viral persistence in part by down-regulating major histocompatibility complex class I protein (MHC-I or HLA-I) from the cell surface. Two different models have been proposed to explain this phenomenon as follows: 1) stimulation of MHC-I retrograde trafficking from and aberrant recycling to the plasma membrane, and 2) inhibition of anterograde trafficking of newly synthesized HLA-I from the endoplasmic reticulum to the plasma membrane. We show here that Nef simultaneously uses both mechanisms to down-regulate HLA-I in peripheral blood mononuclear cells or HeLa cells. Consistent with this, we found by using fluorescence correlation spectroscopy that a third of diffusing HLA-I at the endoplasmic reticulum, Golgi/trans-Golgi network, and the plasma membrane (PM) was associated with Nef. The binding of Nef was similarly avid for native HLA-I and recombinant HLA-I A2 at the PM. Nef binding to HLA-I at the PM was sensitive to specific inhibition of endocytosis. It was also attenuated by cyclodextrin disruption of PM lipid micro-domain architecture, a change that also retarded lateral diffusion and induced large clusters of HLA-I. In all, our data support a model for Nef down-regulation of HLA-I that involves both major trafficking itineraries and persistent protein-protein interactions throughout the cell.     

An unexpected role for PI4,5P2 in EGF receptor endosomal trafficking

In mammalian cells, three Cdc25 phosphatases A, B, C coordinate cell cycle progression through activating dephosphorylation of Cyclin-dependent kinases. Whereas Cdc25B is believed to trigger entry into mitosis, Cdc25C is thought to act at a later stage of mitosis and in the nucleus. We report that a fraction of Cdc25C localises to centrosomes in a cell cycle-dependent fashion, as of late S phase and throughout G2 and mitosis. Moreover, Cdc25C colocalises with Cyclin B1 at centrosomes in G2 and in prophase and Fluorescence Recovery after Photobleaching experiments reveal that they are both in dynamic exchange between the centrosome and the cytoplasm. The centrosomal localisation of Cdc25C is essentially mediated by its catalytic C-terminal domain, but does not require catalytic activity. In fact phosphatase-dead and substrate-binding hotspot mutants of Cdc25C accumulate at centrosomes together with phosphoTyr15-Cdk1 and behave as dominant negative forms that impair entry into mitosis. Taken together, our data suggest an unexpected function for Cdc25C at the G2/M transition, in dephosphorylation of Cdk1. We propose that Cdc25C may participate in amplification of Cdk1-Cyclin B1 activity following initial activation by Cdc25B, and that this process is initiated at the centrosome, then further propagated throughout the cytoplasm thanks to the dynamic behavior of both Cdc25C and Cyclin B1.     

Defining a mutational signature for endometrial cancer screening and early detection

IntroductionThe current availability of genomic information represents an opportunity to develop new strategies for early detection of cancer. New molecular tests for endometrial cancer may improve performance and failure rates of histological aspirate-based diagnosis, and provide promising perspectives for a potential screening scenario. However, the selection of relevant biomarkers to develop efficient strategies can be a challenge.Materials and methodsWe developed an algorithm to identify the largest number of patients with endometrial cancer using the minimum number of somatic mutations based on The Cancer Genome Atlas (TCGA) dataset.ResultsThe algorithm provided the number of subjects with mutations (sensitivity) for a given number of biomarkers included in the signature. For instance, by evaluating the 50 most representative point mutations, up to 81.9% of endometrial cancers can be identified in the TCGA dataset. At gene level, a 92.9% sensitivity can be obtained by interrogating five genes.DiscussionWe developed a computational method to aid in the selection of relevant genomic biomarkers in endometrial cancer that can be adapted to other cancer types or diseases.     

Estimation of stage-specific developmental times and survivorship from stage frequency data

An algorithm for estimating mean stage durations, the standard deviations associated with the means, and stage-specific survivorships from stage frequency data is presented. The algorithm is based on an age-structured, distributed delay simulation model which usesErlang distributions to determine the probability of maturity for individuals in each stage. If the data set extends beyond the first generation, the algorithm requires a fecundity rate, as well as stage frequencies, as input. Goodness-of-fit was measured using a weighted least squares calculation summed over all observed stages and all sampling dates.     

Carbamoyl phosphate synthetase 1 deficiency in Italy: clinical and genetic findings in a heterogeneous cohort

Carbamoyl Phosphate Synthetase 1 deficiency (CPS1D) is a rare autosomal recessive urea cycle disorder, potentially leading to lethal hyperammonemia. Based on the age of onset, there are two distinct phenotypes: neonatal and late form. The CPS1 enzyme, located in the mitochondrial matrix of hepatocytes and epithelial cells of intestinal mucosa, is encoded by the CPS1 gene. At present more than 220 clear-cut genetic lesions leading to CPS1D have been reported. As most of them are private mutations diagnosis is complicated.Here we report an overview of the main clinical findings and biochemical and molecular data of 13 CPS1D Italian patients. In two of them, one with the neonatal form and one with the late form, cadaveric auxiliary liver transplant was performed. Mutation analysis in these patients identified 17 genetic lesions, 9 of which were new confirming their “private” nature. Seven of the newly identified mutations were missense/nonsense changes. In order to study their protein level effects, we performed an in silico analysis whose results indicate that the amino acid substitutions occur at evolutionary conserved positions and affect residues necessary for enzyme stability or function.     

Generation and nuclear translocation of sumoylated transmembrane fragment of cell adhesion molecule L1

The functions of the cell adhesion molecule L1 in the developing and adult nervous system are triggered by homophilic and heterophilic interactions that stimulate signal transductions that activate cellular responses. Here, we show that stimulation of signaling by function-triggering L1 antibodies or L1-Fc leads to serine protease-dependent cleavage of full-length L1 at the plasma membrane and generation of a sumoylated transmembrane 70-kDa fragment comprising the intracellular and transmembrane domains and part of the extracellular domain. The 70-kDa transmembrane fragment is transported from the plasma membrane to a late endosomal compartment, released from endosomal membranes into the cytoplasm, and transferred from there into the nucleus by a pathway that depends on importin and chromatin-modifying protein 1. Mutation of the sumoylation site at Lys(1172) or of the nuclear localization signal at Lys(1147) abolished L1-stimulated generation or nuclear import of the 70-kDa fragment, respectively. Nuclear import of the 70-kDa fragment may activate cellular responses in parallel or in association with phosphorylation-dependent signaling pathways. Alterations in the levels of the 70-kDa fragment during development and in the adult after spinal cord injury or in a mouse model of Alzheimer disease suggest that this fragment is functionally implicated in development, regeneration, neurodegeneration, tumorigenesis, and possibly synaptic plasticity in the mature nervous system.     

ERdj4 protein is a soluble endoplasmic reticulum (ER) DnaJ family protein that interacts with ER-associated degradation machinery

Protein localization within cells regulates accessibility for interactions with co-factors and substrates. The endoplasmic reticulum (ER) BiP co-factor ERdj4 is up-regulated by ER stress and has been implicated in ER-associated degradation (ERAD) of multiple unfolded secretory proteins. Several other ERdj family members tend to interact selectively with nascent proteins, presumably because those ERdj proteins associate with the Sec61 translocon that facilitates entry of nascent proteins into the ER. How ERdj4 selects and targets terminally misfolded proteins for destruction remains poorly understood. In this study, we determined properties of ERdj4 that might aid in this function. ERdj4 was reported to retain its signal sequence and to be resistant to mild detergent extraction, suggesting that it was an integral membrane protein. However, live cell photobleaching analyses of GFP-tagged ERdj4 revealed that the protein exhibits diffusion coefficients uncommonly high for an ER integral membrane protein and more similar to the mobility of a soluble luminal protein. Biochemical characterization established that the ERdj4 signal sequence is cleaved to yield a soluble protein. Importantly, we found that both endogenous and overexpressed ERdj4 associate with the integral membrane protein, Derlin-1. Our findings now directly link ERdj4 to the ERAD machinery and suggest a model in which ERjd4 could help recruit clients from throughout the ER to ERAD sites.