Macrophage migration inhibitory factor family proteins are multitasking cytokines in tissue injury.

The family of macrophage migration inhibitory factor (MIF) proteins in humans consist of MIF, its functional homolog D-dopachrome tautomerase (D-DT, also known as MIF-2) and the relatively unknown protein named DDT-like (DDTL). MIF is a pleiotropic cytokine with multiple properties in tissue homeostasis and pathology. MIF was initially found to associate with inflammatory responses and therefore established a reputation as a pro-inflammatory cytokine. However, increasing evidence demonstrates that MIF influences many different intra- and extracellular molecular processes important for the maintenance of cellular homeostasis, such as promotion of cellular survival, antioxidant signaling, and wound repair. In contrast, studies on D-DT are scarce and on DDTL almost nonexistent and their functions remain to be further investigated as it is yet unclear how similar they are compared to MIF. Importantly, the many and sometimes opposing functions of MIF suggest that targeting MIF therapeutically should be considered carefully, taking into account timing and severity of tissue injury. In this review, we focus on the latest discoveries regarding the role of MIF family members in tissue injury, inflammation and repair, and highlight the possibilities of interventions with therapeutics targeting or mimicking MIF family proteins.

The planar cell polarity Vangl2 protein: From genetics to cellular and molecular functions.

Planar cell polarity (PCP) refers to the capacity of a tissue, typically, but not exclusively, an epithelium, to transmit directional information across the tissue plane such that its cellular constituents can differentiate, divide or move in a coordinated manner and along a common axis, generally orthogonal to the apical-basal axis. PCP relies on a core module of highly conserved proteins originally identified in Drosophila which can act intra- and extracellularly. In this review, we focus on the vertebrate ortholog of one of these core PCP components, namely the Vangl2 protein. After a brief historical perspective, we discuss novel cellular settings for which a cellular Vangl2 requirement has been recently documented, with a particular emphasis on adult tissues that rely on Vangl2 for the maintenance of their regenerative capacity or their physiological functions. Finally we compile the most recent data about Vangl2 interacting proteins.

The anterior gradient-2 interactome.

The anterior gradient-2 (AGR2) is an endoplasmic reticulum (ER)-resident protein belonging to the protein disulfide isomerase family that mediates the formation of disulfide bonds and assists the protein quality control in the ER. In addition to its role in proteostasis, extracellular AGR2 is responsible for various cellular effects in many types of cancer, including cell proliferation, survival, and metastasis. Various OMICs approaches have been used to identify AGR2 binding partners and to investigate the functions of AGR2 in the ER and outside the cell. Emerging data showed that AGR2 exists not only as monomer, but it can also form homodimeric structure and thus interact with different partners, yielding different biological outcomes. In this review, we summarize the AGR2 "interactome" and discuss the pathological and physiological role of such AGR2 interactions.

Binding partners of NRF2: Functions and regulatory mechanisms.

NRF2 is a redox-sensitive transcription factor that plays an important role in protecting organisms against diverse types of electrophiles or oxidants. The level of NRF2 is maintained low in normal cells, but highly elevated in cancer provoking chemoresistance or radioresistance. It is now recognized that NRF2 does not merely maintain the redox balance, but also plays significant roles in autophagy, apoptosis, cell cycle progression, and stem cell differentiation, all of which could be possibly attributable to the existence of multiple binding proteins. In the present manuscript, we summarize direct binding partners of NRF2 and illustrate how they bind to NRF2 and regulate its stability or activity.

Unconventional Myosins: How Regulation Meets Function.

Unconventional myosins are multi-potent molecular motors that are assigned important roles in fundamental cellular processes. Depending on their mechano-enzymatic properties and structural features, myosins fulfil their roles by acting as cargo transporters along the actin cytoskeleton, molecular anchors or tension sensors. In order to perform such a wide range of roles and modes of action, myosins need to be under tight regulation in time and space. This is achieved at multiple levels through diverse regulatory mechanisms: the alternative splicing of various isoforms, the interaction with their binding partners, their phosphorylation, their applied load and the composition of their local environment, such as ions and lipids. This review summarizes our current knowledge of how unconventional myosins are regulated, how these regulatory mechanisms can adapt to the specific features of a myosin and how they can converge with each other in order to ensure the required tight control of their function.

Identification of new FGF1 binding partners-Implications for its intracellular function.

Besides its classical mode of action through activation of specific receptors at the cell surface, fibroblast growth factor 1 (FGF1) can also cross the cellular membrane and translocate into the cytosol and further to the nucleus. The mechanism of this translocation is described partially, but the role of FGF1 inside the cell remains unknown. The aim of our work was to identify novel binding partners of FGF1 to predict its intracellular functions. We combined three methods of identification of such partners based on different principles: yeast two-hybrid screen and mass spectrometry (MS) analysis of complexes obtained by Tandem Affinity Purification (TAP) or by co-precipitation from cell lysate using recombinant FGF1. Altogether, we identified twenty novel intracellular proteins interacting with FGF1. For selected proteins, their direct interaction with FGF1 was confirmed by pull-down assays and SPR measurements. Interestingly, half of the proteins found are involved in processes related to cell viability, such as apoptosis, cell proliferation, and cell cycle regulation. Thus, our study indicates that the role of intracellular FGF1 is to protect the cell against stress conditions by providing an additional signal for cell survival, independently of receptor-activated signaling cascades.

Cellular localization and characterization of cytosolic binding partners for Gla domain-containing proteins PRRG4 and PRRG2.

The genes encoding a family of proteins termed proline-rich γ-carboxyglutamic acid (PRRG) proteins were identified and characterized more than a decade ago, but their functions remain unknown. These novel membrane proteins have an extracellular γ-carboxyglutamic acid (Gla) protein domain and cytosolic WW binding motifs. We screened WW domain arrays for cytosolic binding partners for PRRG4 and identified novel protein-protein interactions for the protein. We also uncovered a new WW binding motif in PRRG4 that is essential for these newly found protein-protein interactions. Several of the PRRG-interacting proteins we identified are essential for a variety of physiologic processes. Our findings indicate possible novel and previously unidentified functions for PRRG proteins.

MAP3K4 kinase action and dual role in cancer.

It is commonly known that the MAPK pathway is involved in translating environmental inputs, regulating downstream reactions, and maintaining the intrinsic dynamic balance. Numerous essential elements and regulatory processes are included in this pathway, which are essential to its functionality. Among these, MAP3K4, a member of the serine/threonine kinases family, plays vital roles throughout the organism's life cycle, including the regulation of apoptosis and autophagy. Moreover, MAP3K4 can interact with key partners like GADD45, which affects organism's growth and development. Notably, MAP3K4 functions as both a tumor promotor and suppressor, being activated by a variety of factors and triggering diverse downstream pathways that differently influence cancer progression. The aim of this study is to provide a brief overview of physiological functions of MAP3K4 and shed light on its contradictory roles in tumorigenesis.

Progesterone Receptor Membrane Component 1 and its Accomplice: Emerging Therapeutic Targets in Lung Cancer.

Progesterone receptor membrane component 1 (PGRMC1) is a trans-membrane evolutionarily conserved protein with a cytochrome b5 like heme/steroid binding domain. PGRMC1 clinical levels are strongly suggested to correlate with poor patient survival and lung cancer prognosis. PGRMC1 has been reported to possess pleiotropic functions, such as participating in cellular and membrane trafficking, steroid hormone signaling, cholesterol metabolism and steroidogenesis, glycolysis and mitochondrial energy metabolism, heme transport and homeostasis, neuronal movement and synaptic function, autophagy, anti-apoptosis, stem cell survival and the list is still expanding. PGRMC1 mediates its pleiotropic functions through its ability to interact with multiple binding partners, such as epidermal growth factor receptor (EGFR), sterol regulatory element binding protein cleavage activating protein (SCAP), insulin induced gene-1 protein (Insig-1), heme binding proteins (hepcidin, ferrochelatase and cyp450 members), plasminogen activator inhibitor 1 RNA binding protein (PAIR-BP1). In this review, we provide a comprehensive overview of PGRMC1 and its associated pleiotropic functions that are indispensable for lung cancer promotion and progression, suggesting it as a prospective therapeutic target for intervention. Notably, we have compiled and reported various preclinical studies wherein prospective agonists and antagonists had been tested against PGRMC1 expressing cancer cell lines, suggesting it as a prospective therapeutic target for cancer intervention.

Vascular Endothelial Growth Factor Receptor, fms-Like Tyrosine Kinase-1 (Flt-1), as a Novel Binding Partner for SARS-CoV-2 Spike Receptor-Binding Domain.

Angiotensin-converting enzyme 2 (ACE2) and neuropilin 1, a vascular endothelial growth factor (VEGF) receptor, were identified to bind to the SARS-CoV-2 spike receptor-binding domain (spike RBD). In silico analysis based on 3D structure, multiple sequence alignment, and molecular docking of second domain of soluble Flt-1 (sFlt-1) and spike RBD revealed structural similarities, sequence homology, and protein-protein interaction. Interaction and binding of recombinant spike RBD (rspike RBD) and recombinant sFlt-1 (rsFlt-1) in vitro induced a conformational change, as revealed by spectrofluorimetric data, with increased fluorescence intensity in emission spectra as compared to either of the proteins alone. Results on ELISA confirmed the binding and cross-reactivity of rspike-RBD and rsFlt-1 as determined by using either specific antibodies towards each protein or immunized human serum. We found that polyclonal or monoclonal anti-spike RBD antibodies can recognize either rsFlt-1 or rspike RBD, showing cross-reactivity for the two proteins in a dose-dependent binding response. Recognition of bound rspike RBD or rsFlt-1 by anti-Flt-1 or anti-spike RBD antibodies, respectively, as observed by immunoblotting, further confirmed interaction between the two proteins. Immunoprecipitation and immunoblot analysis demonstrated the identification of rspike RBD binding to the Flt-1 receptor on A549 cells. Further, the binding of rspike RBD to Flt-1 receptor was shown using immunofluorescence on 2D-culture or 3D-spheroid of MDA-MB-231 cells, which over-express Flt-1 receptor. Together, our study concludes that the Flt-1 receptor is a novel binding partner for SARS-CoV-2 spike RBD.

Tumor suppressor gene DLC1: Its modifications, interactive molecules, and potential prospects for clinical cancer application.

Deleted in liver cancer 1 (DLC1) is a recognized tumor suppressor gene that negatively regulates Rho family proteins by hydrolyzing the active GTP-bound state to its inactive GDP-bound state. Active Rho proteins play a positive role in tumorigenesis. Numerous in vitro and in vivo experiments have shown that DLC1 is downregulated or inactivated in various solid tumors, which may be due to the following five reasons: genomic deletion, epigenetic modification and ubiquitin-dependent proteasomal degradation may cause DLC1 underexpression; phosphorylation at the post-translation level may cause DLC1 inactivation; and failure to localize at focal adhesions (FAs) may prevent DLC1 from exerting full activity. All of the causes could be attributed to molecular binding. Experimental evidence suggests that direct or indirect targeting of DLC1 is feasible for cancer treatment. Therefore, elucidating the interaction of DLC1 with its binding partners might provide novel targeted therapies for cancer. In this review, we summarized the binding partners of DLC1 at both the gene and protein levels and expounded a variety of anticancer drugs targeting DLC1 to provide information about DLC1 as a cancer diagnostic indicator or therapeutic target.

Regulation of acid-sensing ion channels by protein binding partners.

Acid-sensing ion channels (ASICs) are a family of proton-gated cation channels that contribute to a diverse array of functions including pain sensation, cell death during ischemia, and more broadly to neurotransmission in the central nervous system. There is an increasing interest in understanding the physiological regulatory mechanisms of this family of channels. ASICs have relatively short N- and C-termini, yet a number of proteins have been shown to interact with these domains both in vitro and in vivo. These proteins can impact ASIC gating, localization, cell-surface expression, and regulation. Like all ion channels, it is important to understand the cellular context under which ASICs function in neurons and other cells. Here we will review what is known about a number of these potentially important regulatory molecules.

Human Galectin-1 in Multiple Cancers: A Privileged Molecular Target in Oncology.

Galectin-1 (Gal-1), a 14kDa carbohydrate-binding protein of the galectin family found in humans, affects intracellular signaling pathways upon interaction with ß-galactosides on cell-surface, cytosol, and nucleus. The structural information reveals that it consists of a monovalent dimer composed of subunits with one Carbohydrate Recognition Domain (CRD), which is the main active site to interact with various glycoproteins, and carbohydrates in the body to regulate cellular functions. Gal-1 contributes towards different events associated with cancer biology, including tumor transformation, cell cycle regulation, apoptosis, cell adhesion, migration, and inflammation. The extracellular existence and function of Gal-1 have been well-established, and it is known to express in many tumor types, including astrocytoma, melanoma, prostate, colon, bladder, and ovarian carcinomas, etc. Several studies suggested the upregulation of Gal-1 levels in multiple cancer cells. Thus, Gal-1 is a promising molecular target for the development of new therapeutic tools. The present review focuses on the functions of Gal-1 in tumor progression in multiple cancers and its structural insights.

Intracellular partners of fibroblast growth factors 1 and 2 - implications for functions.

Fibroblast growth factors 1 and 2 (FGF1 and FGF2) are mainly considered as ligands of surface receptors through which they regulate a broad spectrum of biological processes. They are secreted in non-canonical way and, unlike other growth factors, they are able to translocate from the endosome to the cell interior. These unique features, as well as the role of the intracellular pool of FGF1 and FGF2, are far from being fully understood. An increasing number of reports address this problem, focusing on the intracellular interactions of FGF1 and 2. Here, we summarize the current state of knowledge of the FGF1 and FGF2 binding partners inside the cell and the possible role of these interactions. The partner proteins are grouped according to their function, including proteins involved in secretion, cell signaling, nucleocytoplasmic transport, binding and processing of nucleic acids, ATP binding, and cytoskeleton assembly. An in-depth analysis of the network of these binding partners could indicate novel, non-classical functions of FGF1 and FGF2 and uncover an additional level of a fine control of the well-known FGF-regulated cellular processes.

Identification of Binding Partners of CsaA - An Archaeal Chaperonic Protein of Picrophilus torridus.

BACKGROUND: CsaA is among the few chaperones which are present in both bacteria and archaea, but absent in eukaryotes. There are no reports on interactome analysis of CsaA from archaea, till date. Identification of binding partners of CsaA might be helpful in understanding CsaA-associated processes in Picrophilus torridus an extreme thermoacidophilic euryarchaeon. OBJECTIVES: The present study was conducted to identify the binding partners of CsaA of P. torridus (PtCsaA). METHODS: The binding partners of PtCsaA were isolated and identified using a pull down assay and liquid chromatography-mass spectrometry (LC-MS). RESULTS: The results revealed twelve potential binding partners of CsaA. These were thermosome subunits (Q6KZS2 and Q6L132), nascent polypeptide-associated complex protein (Q6L1N3), elongation factor 1-alpha (Q6L202), uncharacterized protein (Q6L0Y6), citrate synthase (Q6L0M8), asparaginyl- tRNA synthetase (Q6L0M5), succinyl-CoA synthetase beta chain (Q6L0B4), pyruvate ferredoxin oxidoreductase alpha and beta chain proteins (Q6KZA7 and Q6KZA6, respectively), malate dehydrogenase (Q6L0C3) and reversed fumarylacetoacetase (Q6KZ97). Functional categorization revealed that of these, six proteins were involved in energy metabolic pathways, three were archaeal chaperones, two were involved in translation and one might be a transcription regulator. STRING-based analysis of the protein-protein interactions of the experimental interactome revealed strong interactions among them. CONCLUSION: PtCsaA might be a multifaceted protein which besides translation might also play important role in metabolic processes of P. torridus. However, further experiments investigating the binding partners of CsaA in other archaea are required for a better understanding of CsaA-associated processes in archaea.

Gene expression regulations by long noncoding RNAs and their roles in cancer.

Cancers are deadly diseases. The general mystery of carcinogenesis, primary and acquired drug resistance and distant organ metastasis are still puzzles to be solved. Long noncoding RNAs (lncRNAs) are receiving more and more attention in recent years since their roles in transcriptional regulation have been unveiled. Detailed functional annotations of lncRNAs have showed that their regulatory roles are largely determined by their binding partners. LncRNAs directly bind to DNA, RNA and proteins and regulate gene expression at transcriptional, post-transcriptional and post-translational levels. Here we review the current understanding of molecular functions of lncRNAs, will emphasize on their binding partners and summarize their biological roles in cancer.

Insulin-Like Growth Factor Binding Protein-3 (IGFBP-3): Unraveling the Role in Mediating IGF-Independent Effects Within the Cell.

Insulin-like growth factor (IGF) binding protein-3 (IGFBP-3), one of the six members of the IGFBP family, is a key protein in the IGF pathway. IGFBP-3 can function in an IGF-dependent as well as in an IGF-independent manner. The IGF-dependent roles of IGFBP-3 include its endocrine role in the delivery of IGFs from the site of synthesis to the target cells that possess IGF receptors and the activation of associated downstream signaling. IGF-independent role of IGFBP-3 include its interactions with the proteins of the extracellular matrix and the proteins of the plasma membrane, its translocation through the plasma membrane into the cytoplasm and into the nucleus. The C-terminal domain of IGFBP-3 has the ability to undergo cell penetration therefore, generating a short 8-22-mer C-terminal domain peptides that can be conjugated to drugs or genes for effective intracellular delivery. This has opened doors for biotechnological applications of the molecule in molecular medicine. The aim of this this review is to summarize the complex roles of IGFBP-3 within the cell, including its mechanisms of cellular uptake and its translocation into the nucleus, various molecules with which it is capable of interacting, and its ability to regulate IGF-independent cell growth, survival and apoptosis. This would pave way into understanding the modus operandi of IGFBP-3 in regulating IGF-independent processes and its pleiotropic ability to bind with potential partners thus regulating several cellular functions implicated in metabolic diseases, including cancer.

Affinity Purification of NF1 Protein-Protein Interactors Identifies Keratins and Neurofibromin Itself as Binding Partners.

Neurofibromatosis Type 1 (NF1) is caused by pathogenic variants in the NF1 gene encoding neurofibromin. Definition of NF1 protein-protein interactions (PPIs) has been difficult and lacks replication, making it challenging to define binding partners that modulate its function. We created a novel tandem affinity purification (TAP) tag cloned in frame to the 3' end of the full-length murine Nf1 cDNA (mNf1). We show that this cDNA is functional and expresses neurofibromin, His-Tag, and can correct p-ERK/ERK ratios in NF1 null HEK293 cells. We used this affinity tag to purify binding partners with Strep-Tactin®XT beads and subsequently, identified them via mass spectrometry (MS). We found the tagged mNf1 can affinity purify human neurofibromin and vice versa, indicating that neurofibromin oligomerizes. We identify 21 additional proteins with high confidence of interaction with neurofibromin. After Metacore network analysis of these 21 proteins, eight appear within the same network, primarily keratins regulated by estrogen receptors. Previously, we have shown that neurofibromin levels negatively regulate keratin expression. Here, we show through pharmacological inhibition that this is independent of Ras signaling, as the inhibitors, selumetinib and rapamycin, do not alter keratin expression. Further characterization of neurofibromin oligomerization and binding partners could aid in discovering new neurofibromin functions outside of Ras regulation, leading to novel drug targets.

GST Pull-Down Assay to Measure Complex Formations.

The GST pull-down assay is an intuitive and fast in vitro method for analyzing protein-protein or protein-ligand interactions and is comprised of a "bait" which is a GST-fused protein expressed in E. coli host or a baculovirus expression system and a "prey" which comprises putative binding partner protein(s) or other ligand molecule(s). This method is suitable for examining the direct interaction between two purified proteins and estimating the extent of the affinity.

Novel insight into the function of tankyrase.

Tankyrases are multifunctional poly(ADP-ribose) polymerases that regulate a variety of cellular processes, including Wnt signaling, telomere maintenance and mitosis regulation. Tankyrases interact with target proteins and regulate their interactions and stability through poly(ADP-ribosyl) ation. In addition to their roles in telomere maintenance and regulation of mitosis, tankyrase proteins regulate tumor suppressors, including AXIN, phosphatase and tensin homolog and angiomotin. Therefore, tankyrases may be effective targets for cancer treatment. Tankyrase inhibitors could affect a variety of carcinogenic pathways that promote uncontrolled proliferation, including Wnt, AKT, yes-associated protein, telomere maintenance and mitosis regulation. Recently, novel aspects of the function and mechanism of tankyrases have been reported, and a number of tankyrase inhibitors have been identified. A combination of conventional chemotherapy agents with tankyrase inhibitors may have synergistic anticancer effects. Therefore, it is expected that more advanced and improved tankyrase inhibitors will be developed, enabling novel therapeutic strategies against cancer and other tankyrase-associated diseases. The present review discusses tankyrase function and the role of tankyrase inhibitors in the treatment of cancer.