Protein Trafficking Pathways Research Articles

Structural Basis for the Interaction between the Ezrin FERM-Domain and Human Aquaporins.

The Ezrin/Radixin/Moesin (ERM) family of proteins act as cross-linkers between the plasma membrane and the actin cytoskeleton. This mechanism plays an essential role in processes related to membrane remodeling and organization, such as cell polarization, morphogenesis and adhesion, as well as in membrane protein trafficking and signaling pathways. For several human aquaporin (AQP) isoforms, an interaction between the ezrin band Four-point-one, Ezrin, Radixin, Moesin (FERM)-domain and the AQP C-terminus has been demonstrated, and this is believed to be important for AQP localization in the plasma membrane. Here, we investigate the structural basis for the interaction between ezrin and two human AQPs: AQP2 and AQP5. Using microscale thermophoresis, we show that full-length AQP2 and AQP5 as well as peptides corresponding to their C-termini interact with the ezrin FERM-domain with affinities in the low micromolar range. Modelling of the AQP2 and AQP5 FERM complexes using ColabFold reveals a common mode of binding in which the proximal and distal parts of the AQP C-termini bind simultaneously to distinct binding sites of FERM. While the interaction at each site closely resembles other FERM-complexes, the concurrent interaction with both sites has only been observed in the complex between moesin and its C-terminus which causes auto-inhibition. The proposed interaction between AQP2/AQP5 and FERM thus represents a novel binding mode for extrinsic ERM-interacting partners.

Sit4 Genetically Interacts with Vps27 to Regulate Mitochondrial Function and Lifespan in Saccharomyces cerevisiae.

The Sit4 protein phosphatase plays a key role in orchestrating various cellular processes essential for maintaining cell viability during aging. We have previously shown that SIT4 deletion promotes vacuolar acidification, mitochondrial derepression, and oxidative stress resistance, increasing yeast chronological lifespan. In this study, we performed a proteomic analysis of isolated vacuoles and yeast genetic interaction analysis to unravel how Sit4 influences vacuolar and mitochondrial function. By employing high-resolution mass spectrometry, we show that sit4Δ vacuolar membranes were enriched in Vps27 and Hse1, two proteins that are part of the endosomal sorting complex required for transport-0. In addition, SIT4 exhibited a negative genetic interaction with VPS27, as sit4∆vps27∆ double mutants had a shortened lifespan compared to sit4∆ and vps27∆ single mutants. Our results also show that Vps27 did not increase sit4∆ lifespan by improving protein trafficking or vacuolar sorting pathways. However, Vps27 was critical for iron homeostasis and mitochondrial function in sit4∆ cells, as sit4∆vps27∆ double mutants exhibited high iron levels and impaired mitochondrial respiration. These findings show, for the first time, cross-talk between Sit4 and Vps27, providing new insights into the mechanisms governing chronological lifespan.

The relationship between intraflagellar transport and upstream protein trafficking pathways and macrocyclic lactone resistance in Caenorhabditis elegans.

Parasitic nematodes are globally important and place a heavy disease burden on infected humans, crops, and livestock, while commonly administered anthelmintics used for treatment are being rendered ineffective by increasing levels of resistance. It has recently been shown in the model nematode Caenorhabditis elegans that the sensory cilia of the amphid neurons play an important role in resistance toward macrocyclic lactones such as ivermectin (an avermectin) and moxidectin (a milbemycin) either through reduced uptake or intertissue signaling pathways. This study interrogated the extent to which ciliary defects relate to macrocyclic lactone resistance and dye-filling defects using a combination of forward genetics and targeted resistance screening approaches and confirmed the importance of intraflagellar transport in this process. This approach also identified the protein trafficking pathways used by the downstream effectors and the components of the ciliary basal body that are required for effector entry into these nonmotile structures. In total, 24 novel C. elegans anthelmintic survival-associated genes were identified in this study. When combined with previously known resistance genes, there are now 46 resistance-associated genes that are directly involved in amphid, cilia, and intraflagellar transport function.

Colocalising proteins and polysaccharides in plants for cell wall and trafficking studies.

Plant cell walls (PCWs) are intricate structures with complex polysaccharides delivered by distinct trafficking routes. Unravelling the intricate trafficking pathways of polysaccharides and proteins involved in PCW biosynthesis is a crucial first step towards understanding the complexities of plant growth and development. This study investigated the feasibility of employing a multi-modal approach that combines transmission electron microscopy (TEM) with molecular-genetic tagging and antibody labelling techniques to differentiate these pathways at the nanoscale. The genetically encoded electron microscopy (EM) tag APEX2 was fused to Arabidopsis thaliana cellulose synthase 6 (AtCESA6) and Nicotiana alata ARABINAN DEFICIENT LIKE 1 (NaARADL1), and these were transiently expressed in Nicotiana benthamiana leaves. APEX2 localization was then combined with immunolabeling using pectin-specific antibodies (JIM5 and JIM7). Our results demonstrate distinct trafficking patterns for AtCESA6 and NaARADL, with AtCESA6 localized primarily to the plasma membrane and vesicles, while NaARADL1 was found in the trans-Golgi network and cytoplasmic vesicles. Pectin epitopes were observed near the plasma membrane, in Golgi-associated vesicles, and in secretory vesicle clusters (SVCs) with both APEX2 constructs. Notably, JIM7 labelling was found in vesicles adjacent to APEX2-AtCESA6 vesicles, suggesting potential co-trafficking. This integrative approach offers a powerful tool for elucidating the dynamic interactions between PCW components at the nanoscale level. The methodology presented here facilitates the precise mapping of protein and polysaccharide trafficking pathways, advancing our understanding of PCW biosynthesis and providing avenues for future research aimed at engineering plant cell walls for various applications.

Deep-Sea Sponges and Corals off the Western Coast of Florida-Intracellular Mechanisms of Action of Bioactive Compounds and Technological Advances Supporting the Drug Discovery Pipeline.

The majority of natural products utilized to treat a diverse array of human conditions and diseases are derived from terrestrial sources. In recent years, marine ecosystems have proven to be a valuable resource of diverse natural products that are generated to defend and support their growth. Such marine sources offer a large opportunity for the identification of novel compounds that may guide the future development of new drugs and therapies. Using the National Oceanic and Atmospheric Administration (NOAA) portal, we explore deep-sea coral and sponge species inhabiting a segment of the U.S. Exclusive Economic Zone, specifically off the western coast of Florida. This area spans ~100,000 km2, containing coral and sponge species at sea depths up to 3000 m. Utilizing PubMed, we uncovered current knowledge on and gaps across a subset of these sessile organisms with regards to their natural products and mechanisms of altering cytoskeleton, protein trafficking, and signaling pathways. Since the exploitation of such marine organisms could disrupt the marine ecosystem leading to supply issues that would limit the quantities of bioactive compounds, we surveyed methods and technological advances that are necessary for sustaining the drug discovery pipeline including in vitro aquaculture systems and preserving our natural ecological community in the future. Collectively, our efforts establish the foundation for supporting future research on the identification of marine-based natural products and their mechanism of action to develop novel drugs and therapies for improving treatment regimens of human conditions and diseases.

The seven rice vacuolar sorting receptors localize to prevacuolar compartments

Vacuolar sorting is critically important in plants as it regulates the mobilization of proteins and plays a major role in important agricultural traits like yield and seed protein content. Vacuolar sorting receptors (VSRs) are integral membrane proteins that mediate protein trafficking from the Golgi apparatus to the vacuole via the intermediate membrane-bound prevacuolar compartment (PVC)/multivesicular body (MVB). VSR proteins, such as an 80 kD (BP-80) from pea, also serve as markers for PVC/MVB. Dissecting VSR-mediated protein trafficking pathways may provide ways to enhance agronomic traits and crop yield. Green fluorescence protein (GFP) fusions with the seven Arabidopsis (Arabidopsis thaliana) VSRs were previously shown to localize to PVCs in transgenic tobacco BY-2 cells. The Rice (Oryza sativa) genome contains seven VSRs (OsVSR1-7), but little is known about their subcellular localizations. Here we studied the subcellular localization of OsVSR1-7 b y using a reporter approach, in which GFP-OsVSR1-7 fusions containing the transmembrane domain (TMD) and cytoplasmic tail (CT) of individual OsVSR were expressed in the protoplasts of rice, transgenic tobacco BY-2 cells and transgenic rice plants. Immunofluorescent labelling studies and confocal laser scanning microscope observation demonstrated that the seven OsVSRs are localized to PVCs and form ring-like structures upon wortmannin treatment. Therefore, we have verified the subcellular localization of OsVSR1-7 in this study. The OsVSRs tagged with GFP can serve as PVCs/MVBs markers in rice for the future studies.

The Osiris family genes function as novel regulators of the tube maturation process in the Drosophila trachea.

Drosophila trachea is a premier model to study tube morphogenesis. After the formation of continuous tubes, tube maturation follows. Tracheal tube maturation starts with an apical secretion pulse that deposits extracellular matrix components to form a chitin-based apical luminal matrix (aECM). This aECM is then cleared and followed by the maturation of taenidial folds. Finally, air fills the tubes. Meanwhile, the cellular junctions are maintained to ensure tube integrity. Previous research has identified several key components (ER, Golgi, several endosomes) of protein trafficking pathways that regulate the secretion and clearance of aECM, and the maintenance of cellular junctions. The Osiris (Osi) gene family is located at the Triplo-lethal (Tpl) locus on chromosome 3R 83D4-E3 and exhibits dosage sensitivity. Here, we show that three Osi genes (Osi9, Osi15, Osi19), function redundantly to regulate adherens junction (AJ) maintenance, luminal clearance, taenidial fold formation, tube morphology, and air filling during tube maturation. The localization of Osi proteins in endosomes (Rab7-containing late endosomes, Rab11-containing recycling endosomes, Lamp-containing lysosomes) and the reduction of these endosomes in Osi mutants suggest the possible role of Osi genes in tube maturation through endosome-mediated trafficking. We analyzed tube maturation in zygotic rab11 and rab7 mutants, respectively, to determine whether endosome-mediated trafficking is required. Interestingly, similar tube maturation defects were observed in rab11 but not in rab7 mutants, suggesting the involvement of Rab11-mediated trafficking, but not Rab7-mediated trafficking, in this process. To investigate whether Osi genes regulate tube maturation primarily through the maintenance of Rab11-containing endosomes, we overexpressed rab11 in Osi mutant trachea. Surprisingly, no obvious rescue was observed. Thus, increasing endosome numbers is not sufficient to rescue tube maturation defects in Osi mutants. These results suggest that Osi genes regulate other aspects of endosome-mediated trafficking, or regulate an unknown mechanism that converges or acts in parallel with Rab11-mediated trafficking during tube maturation.

TRAPPC1 is essential for the maintenance and differentiation of common myeloid progenitors inmice.

Myeloid cell development in bone marrow is essential for the maintenance of peripheral immune homeostasis. However, the role of intracellular protein trafficking pathways during myeloid cell differentiation is currently unknown. By mining bioinformatics data, we identify trafficking protein particle complex subunit 1 (TRAPPC1) as continuously upregulated during myeloid cell development. Using inducible ER-TRAPPC1 knockout mice and bone marrow chimeric mouse models, we demonstrate that TRAPPC1 deficiency causes severe monocyte and neutrophil defects, accompanied by a selective decrease in common myeloid progenitors (CMPs) and subsequent cell subsets in bone marrow. TRAPPC1-deleted CMPs differentiate poorly into monocytes and neutrophils invivo and invitro, in addition to exhibiting enhanced endoplasmic reticulum stress and apoptosis via a Ca2+ -mitochondria-dependent pathway. Cell cycle arrest and senescence of TRAPPC1-deleted CMPs are mediated by the activation of pancreatic endoplasmic reticulum kinase and the upregulation of cyclin-dependent kinase inhibitor p21. This study reveals the essential role of TRAPPC1 in the maintenance and differentiation of CMPs and highlights the significance of protein processing and trafficking processes in myeloid cell development.

Transcriptome Analysis Reveals Potential Mechanism in Storage Protein Trafficking within Developing Grains of Common Wheat

Gluten proteins are the major storage protein fraction in the mature wheat grain. They are restricted to the starchy endosperm, which defines the viscoelastic properties of wheat dough. The synthesis of these storage proteins is controlled by the endoplasmic reticulum (ER) and is directed into the vacuole via the Golgi apparatus. In the present study, transcriptome analysis was used to explore the potential mechanism within critical stages of grain development of wheat cultivar "Shaannong 33" and its sister line used as the control (CK). Samples were collected at 10 DPA (days after anthesis), 14 DPA, 20 DPA, and 30 DPA for transcriptomic analysis. The comparative transcriptome analysis identified that a total of 18,875 genes were differentially expressed genes (DEGs) between grains of four groups "T10 vs. CK10, T14 vs. CK14, T20 vs. CK20, and T30 vs. CK30", including 2824 up-regulated and 5423 down-regulated genes in T30 vs. CK30. Further, the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment highlighted the maximum number of genes regulating protein processing in the endoplasmic reticulum (ER) during grain enlargement stages (10-20 DPA). In addition, KEGG database analysis reported 1362 and 788 DEGs involved in translation, ribosomal structure, biogenesis, flavonoid biosynthesis pathway and intracellular trafficking, secretion, and vesicular transport through protein processing within ER pathway (ko04141). Notably, consistent with the higher expression of intercellular storage protein trafficking genes at the initial 10 DPA, there was relatively low expression at later stages. Expression levels of nine randomly selected genes were verified by qRT-PCR, which were consistent with the transcriptome data. These data suggested that the initial stages of "cell division" played a significant role in protein quality control within the ER, thus maintaining the protein quality characteristics at grain maturity. Furthermore, our data suggested that the protein synthesis, folding, and trafficking pathways directed by a different number of genes during the grain enlargement stage contributed to the observed high-quality characteristics of gluten protein in Shaannong 33 (Triticum aestivum L.).

Advances in vesicle trafficking of membrane proteins and their regulatory mechanisms

<p indent="0mm">Membrane proteins are essential components of cell membrane structures and can be transported through vesicles as signaling molecules. The vesicle trafficking of membrane proteins is crucial for maintaining biological processes such as plant growth and development, substance exchange, cell recognition, immune response, and signal transduction. Both vesicle exocytosis and endocytosis play integral roles in regulating the activities and turnover of plasma membrane proteins required for signal triggering or attenuation at the cell surface. Exocytosis involves the transport of newly synthesized or recycled material to the plasma membrane (PM) in vesicles where they are released into the extracellular space. Recent investigations have shown that changing the balance between conventional and unconventional protein secretion can be an efficient strategy to respond to stressful conditions in eukaryotes. With the aid of an N-terminal located signal peptide (SP), conventional protein secretion delivers newly synthesized proteins from the endoplasmic reticulum (ER) to the Golgi apparatus and the <italic>trans</italic>-Golgi network (TGN) and finally to the PM or extracellular space via secretory vesicles or secretory granules. Unconventional protein secretion is responsible for the secretion of such SP-lacking protein. Notably, several unconventional protein secretion pathways mediated by distinct mechanisms have been reported in plants, most of which are related to stress responses. Conversely to exocytosis, endocytosis is the process by which material associated with the cellular surface is uptaken by invagination of the plasma membrane and internalized as an endocytic vesicle. In recent years, substantial progress has been made in the characterization of the plant components involved in clathrin-mediated and membrane microdomain-associated endocytic pathways. Moreover, the coordination of exocytic and endocytic trafficking has been shown to be important for various plant cell functions. An increasing pool of evidence has indicated that multiple cellular processes require tightly regulated exocytosis-endocytosis coupling, including nutrient uptake, signaling transduction, and plant–microbe interactions. However, compared to animal systems in which multiple mechanisms have been described, the functions and regulation of coupled trafficking processes in plants remain poorly understood. All proteins in the cells of an organism and most extracellular proteins are continually being degraded and replaced. The autophagy pathway is a conserved protein auto degradation pathway that delivers dysfunctional organelles or other cytoplasmic components to vacuoles for degradation and recycling in plants. In addition, autophagy is reported to be involved in plant development as part of processes such as root tip cell growth and differentiation. Recent observations have revealed possible interplay between vesicle trafficking and autophagic pathways in regulating vacuolar degradation in plants. In recent years, with constant advances in super-resolution microscopy and labeling methods, much progress has been made in mechanistic studies of membrane protein transportation, and a diversity of exocytic and endocytic trafficking pathways have been uncovered in plants. Although a variety of methods and techniques have been used to study plant vesicle trafficking, a systematic summary of vesicle trafficking pathways of plant membrane proteins has not been published. This review first introduces the relevant organelles involved in membrane protein vesicle trafficking, and then summarizes the different plant vesicle trafficking pathways for membrane proteins. Based on the review, we systematically consolidated the chemical treatments and mutants used in the study of plant vesicle trafficking. Finally, future directions for research in vesicle trafficking pathways of plant membrane proteins are discussed. With recent advancements, we expect major breakthroughs in our understanding of the regulatory mechanisms that determine the perception and adaptability of plants to their environments in the coming years.

Modulation of Cytoskeleton, Protein Trafficking, and Signaling Pathways by Metabolites from Cucurbitaceae, Ericaceae, and Rosaceae Plant Families.

One promising frontier within the field of Medical Botany is the study of the bioactivity of plant metabolites on human health. Although plant metabolites are metabolic byproducts that commonly regulate ecological interactions and biochemical processes in plant species, such metabolites also elicit profound effects on the cellular processes of human and other mammalian cells. In this regard, due to their potential as therapeutic agents for a variety of human diseases and induction of toxic cellular responses, further research advances are direly needed to fully understand the molecular mechanisms induced by these agents. Herein, we focus our investigation on metabolites from the Cucurbitaceae, Ericaceae, and Rosaceae plant families, for which several plant species are found within the state of Florida in Hillsborough County. Specifically, we compare the molecular mechanisms by which metabolites and/or plant extracts from these plant families modulate the cytoskeleton, protein trafficking, and cell signaling to mediate functional outcomes, as well as a discussion of current gaps in knowledge. Our efforts to lay the molecular groundwork in this broad manner hold promise in supporting future research efforts in pharmacology and drug discovery.

Multifaceted Roles of Retromer in EGFR Trafficking and Signaling Activation.

Mammalian retromer complex contributes to multiple early endosome-associated trafficking pathways whose origins are dependent on which sorting nexin (SNX) they are complexed with. In an attempt to dissect out the contribution of individual retromer-SNX complexes, we examined the trafficking of EGFR in detail within a series of KO cell line models. We demonstrated that the depletion of retromer subunit Vps35 leads to decreased EGFR protein levels in resting cells with enhanced association of EGFR with lysosomal compartments. Compared to control cells, the addition of EGF to Vps35 KO cells resulted in a reduced rate of EGFR degradation; AKT activation and cell prolferation rates were elevated, while ERK activation remained relatively unchanged. These observations are consistent with a prolonged temporal association of EGFR within early endosomes due to the inefficiency of early endosome-associated protein trafficking pathways or organelle maturation due to retromer absence. We did not fully delineate the discrete contributions from retromer-associated SNXs to the phenotypes observed from retromer Vps35 depletion. While each of the knock-outs of SNX1/2, SNX3, or SNX27 promotes the enhanced association of EGFR with early endosomal compartments, only the decreased EGF-mediated EGFR degradation was observed in SNX1/2 dKO cells, while the enhanced AKT activation was only increased in SNX3 KO or SNX27 KO cells. Despite this, each of the knock-outs showed increased EGF-stimulated cell proliferation rates.

A Phytophthora capsici RXLR effector manipulates plant immunity by targeting RAB proteins and disturbing the protein trafficking pathway.

The oomycete pathogen Phytophthora capsici encodes hundreds of RXLR effectors that enter the plant cells and suppress host immunity. Only a few of these genes are conserved across different strains and species. Such core effectors might target hub genes and immune pathways in hosts. Here, we describe the functional characterization of the core P. capsici RXLR effector RXLR242. The expression of RXLR242 was up-regulated during infection, and its ectopic expression in Nicotiana benthamiana, an experimental plant host, further promoted Phytophthora infection. RXLR242 physically interacted with a group of RAB proteins that belong to the small GTPase family and play a role in regulating transport pathways in the intracellular membrane trafficking system. In addition, RXLR242 impeded the secretion of PATHOGENESIS-RELATED 1 (PR1) protein to the apoplast. This phenomenon resulted from the competitive binding of RXLR242 to RABE1-7. We also found that RXLR242 interfered with the association between RABA4-3 and its binding protein, thereby disrupting the trafficking of the membrane receptor FLAGELLIN-SENSING 2. Thus, RXLR242 manipulates plant immunity by targeting RAB proteins and disrupting protein trafficking in the host plants.

The Roles of Ubiquitination in Pathogenesis of Influenza Virus Infection.

The ubiquitin system denotes a potent post-translational modification machinery that is capable of activation or deactivation of target proteins through reversible linkage of a single ubiquitin or ubiquitin chains. Ubiquitination regulates major cellular functions such as protein degradation, trafficking and signaling pathways, innate immune response, antiviral defense, and virus replication. The RNA sensor RIG-I ubiquitination is specifically induced by influenza A virus (IAV) to activate type I IFN production. Influenza virus modulates the activity of major antiviral proteins in the host cell to complete its full life cycle. Its structural and non-structural proteins, matrix proteins and the polymerase complex can regulate host immunity and antiviral response. The polymerase PB1-F2 of mutated 1918 IAV, adapts a novel IFN antagonist function by sending the DDX3 into proteasomal degradation. Ultimately the fate of virus is determined by the outcome of interplay between viral components and host antiviral proteins and ubiquitination has a central role in the encounter of virus and its host cell.

α-synuclein and phosphoinositide-binding proteins: α-synuclein inhibits the association of PX- but not FYVE-containing proteins with vesicles in vivo

By an unknown mechanism, alpha-synuclein (α-syn) inhibits autophagy in yeast and human cells. Herein, using the yeast Saccharomyces cerevisiae, we tested the hypothesis that α-syn disrupts autophagy by inhibiting the required association of sorting nexin 4 (Snx4) with phagophores. Snx4 contains a phox (PX) homology domain that selectively binds membranes enriched in phosphatidylinositol 3-phosphate (PI3P). Using fluorescence microscopy, we show that upon nitrogen starvation, 70% of the cells exhibited green puncta (phagophores); whereas identically treated cells expressing α-syn exhibited a significantly lower percentage of cells (30%) with such puncta. Our interpretation is that α-syn outcompetes Snx4 for binding to membranes enriched in PI3P, resulting in fewer phagophores and consequently inefficient induction of autophagy. As a control, we tested whether α-syn disrupts the binding of Vps27-GFP to late endosomes/multivesicular bodies (MVBs). Vps27 contains a PI3P-binding domain called FYVE. α-Syn did not disrupt the binding of Vps27-GFP to late endosomes. α-Syn likely inhibits the binding of PX- but not FYVE-containing proteins to PI3P because FYVE domains bind more than two-orders of magnitude tighter than PX domains. We propose that in all cells, whether yeast or human, α-syn has the potential to inhibit protein trafficking pathways that are dependent on PX-domain proteins such as sorting nexins.

New Insights into the Chloroplast Outer Membrane Proteome and Associated Targeting Pathways.

Plastids are a dynamic class of organelle in plant cells that arose from an ancient cyanobacterial endosymbiont. Over the course of evolution, most genes encoding plastid proteins were transferred to the nuclear genome. In parallel, eukaryotic cells evolved a series of targeting pathways and complex proteinaceous machinery at the plastid surface to direct these proteins back to their target organelle. Chloroplasts are the most well-characterized plastids, responsible for photosynthesis and other important metabolic functions. The biogenesis and function of chloroplasts rely heavily on the fidelity of intracellular protein trafficking pathways. Therefore, understanding these pathways and their regulation is essential. Furthermore, the chloroplast outer membrane proteome remains relatively uncharted territory in our understanding of protein targeting. Many key players in the cytosol, receptors at the organelle surface, and insertases that facilitate insertion into the chloroplast outer membrane remain elusive for this group of proteins. In this review, we summarize recent advances in the understanding of well-characterized chloroplast outer membrane protein targeting pathways as well as provide new insights into novel targeting signals and pathways more recently identified using a bioinformatic approach. As a result of our analyses, we expand the known number of chloroplast outer membrane proteins from 117 to 138.

MetaNetwork Enhances Biological Insights from Quantitative Proteomics Differences by Combining Clustering and Enrichment Analyses.

Interpreting proteomics data remains challenging due to the large number of proteins that are quantified by modern mass spectrometry methods. Weighted gene correlation network analysis (WGCNA) can identify groups of biologically related proteins using only protein intensity values by constructing protein correlation networks. However, WGCNA is not widespread in proteomic analyses due to challenges in implementing workflows. To facilitate the adoption of WGCNA by the proteomics field, we created MetaNetwork, an open-source, R-based application to perform sophisticated WGCNA workflows with no coding skill requirements for the end user. We demonstrate MetaNetwork's utility by employing it to identify groups of proteins associated with prostate cancer from a proteomic analysis of tumor and adjacent normal tissue samples. We found a decrease in cytoskeleton-related protein expression, a known hallmark of prostate tumors. We further identified changes in module eigenproteins indicative of dysregulation in protein translation and trafficking pathways. These results demonstrate the value of using MetaNetwork to improve the biological interpretation of quantitative proteomics experiments with 15 or more samples.

Golgi Metal Ion Homeostasis in Human Health and Diseases.

The Golgi apparatus is a membrane organelle located in the center of the protein processing and trafficking pathway. It consists of sub-compartments with distinct biochemical compositions and functions. Main functions of the Golgi, including membrane trafficking, protein glycosylation, and sorting, require a well-maintained stable microenvironment in the sub-compartments of the Golgi, along with metal ion homeostasis. Metal ions, such as Ca2+, Mn2+, Zn2+, and Cu2+, are important cofactors of many Golgi resident glycosylation enzymes. The homeostasis of metal ions in the secretory pathway, which is required for proper function and stress response of the Golgi, is tightly regulated and maintained by transporters. Mutations in the transporters cause human diseases. Here we provide a review specifically focusing on the transporters that maintain Golgi metal ion homeostasis under physiological conditions and their alterations in diseases.

The disturbance of protein synthesis/degradation homeostasis is a common trait of age-related neurodegenerative disorders.

Protein homeostasis or "proteostasis" represent the process that regulates the balance of the intracellular functional and "healthy" proteins. Proteostasis is fundamental to preserve physiological metabolic processes in the cell and it allow to respond to any given stimulus as the expression of components of the proteostasis network is customized according to the proteomic demands of different cellular environments. In conditions that promote unfolding/misfolding of proteins chaperones act as signaling molecules inducing extreme measures to either fix the problem or destroy unfolded proteins. When the chaperone machinery fails under pathological insults unfolded proteins induce the endoplasmic reticulum (ER) stress activating the unfolded protein response (UPR) machinery. The activation of the UPR restores ER proteostasis primarily through the transcriptional remodeling of ER protein folding, trafficking, and degradation pathways, such as the ubiquitin proteasome system (UPS). If these mechanisms do not manage to clear the aberrant proteins, proteasome overload and become defective, and misfolded proteins may form aggregates thus extending the UPR mechanism. These aggregates are then attempted to be cleared by macroautophagy. Impaired proteostasis promote the accumulation of misfolded proteins that exacerbate the damage to chaperones, surveillance systems and/or degradative activities. Remarkably, the removal of toxic misfolded proteins is critical for all cells, but it is especially significant in neurons since these cannot be readily replaced. In neurons, the maintenance of efficient proteostasis is essential to healthy aging since the dysregulation of the proteostasis network can lead to neurodegenerative disease. Each of these brain pathologies is characterized by the repeated misfolding of one of more peculiar proteins, which evade both the protein folding machinery and cellular degradation mechanisms and begins to form aggregates that nucleate out into large fibrillar aggregates. In this chapter we describe the mechanisms, associated with faulty proteostasis, that promote the formation of protein aggregates, amyloid fibrils, intracellular, and extracellular inclusions in the most common nondegenerative disorders also referred to as protein misfolding disorders.

How Influenza A Virus NS1 Deals with the Ubiquitin System to Evade Innate Immunity.

Ubiquitination is a post-translational modification regulating critical cellular processes such as protein degradation, trafficking and signaling pathways, including activation of the innate immune response. Therefore, viruses, and particularly influenza A virus (IAV), have evolved different mechanisms to counteract this system to perform proper infection. Among IAV proteins, the non-structural protein NS1 is shown to be one of the main virulence factors involved in these viral hijackings. NS1 is notably able to inhibit the host’s antiviral response through the perturbation of ubiquitination in different ways, as discussed in this review.