Mining TCGA to Reveal Immunotherapy-related Genes for Soft Tissue Sarcoma

Objective: By mining the TCGA database to look for immunotherapy targets of soft tissue sarcoma, and analyzed their biological behavior. Methods: The data of 265 samples were downloaded from the TCGA database to analyze the expression profile of soft tissue sarcomas. Research methods include immune and stromal scores, calculating DEGs, volcano maps and differential gene survival curves, gene enrichment analysis.Results: Kaplan-Meier survival curves showed that in the high immune score group, the total survival time was generally higher than that in the low immune score group. Analysis of the top ten terms resulted in the minimum P values for immune and inflammatory responses, plasma membrane, receptor activity, and chemokine activity. By plotting the K-M curves, we obtained 86 survivals related DEGs. Finally, the genes that can be used as independent risk factors for prognosis of soft tissue sarcoma were obtained by multivariate analysis of the DEGs. Conclusion: We believe that these genes are expected to be new targets for sarcoma immunotherapy and key genes for the analysis of prognosis of sarcoma.

In Vitro Characterization of Guanylyl Cyclase BdPepR2 from Brachypodium distachyon Identified through a Motif-Based Approach

In recent years, cyclic guanosine 3′,5′-cyclic monophosphate (cGMP) and guanylyl cyclases (GCs), which catalyze the formation of cGMP, were implicated in a growing number of plant processes, including plant growth and development and the responses to various stresses. To identify novel GCs in plants, an amino acid sequence of a catalytic motif with a conserved core was designed through bioinformatic analysis. In this report, we describe the performed analyses and consider the changes caused by the introduced modification within the GC catalytic motif, which eventually led to the description of a plasma membrane receptor of peptide signaling molecules—BdPepR2 in Brachypodium distachyon. Both in vitro GC activity studies and structural and docking analyses demonstrated that the protein could act as a GC and contains a highly conserved 14-aa GC catalytic center. However, we observed that in the case of BdPepR2, this catalytic center is altered where a methionine instead of the conserved lysine or arginine residues at position 14 of the motif, conferring higher catalytic activity than arginine and alanine, as confirmed through mutagenesis studies. This leads us to propose the expansion of the GC motif to cater for the identification of GCs in monocots. Additionally, we show that BdPepR2 also has in vitro kinase activity, which is modulated by cGMP.

Inhibition by Thyroid Hormones of Cell Migration Activated by IGF-1 and MCP-1 in THP-1 Monocytes: Focus on Signal Transduction Events Proximal to Integrin αvβ3

Interaction between thyroid hormones and the immune system is reported in the literature. Thyroid hormones, thyroxine, T4, but also T3, act non-genomically through mechanisms that involve a plasma membrane receptor αvβ3 integrin, a co-receptor for insulin-like growth factor-1 (IGF-1). Previous data from our laboratory show a crosstalk between thyroid hormones and IGF-1 because thyroid hormones inhibit the IGF-1-stimulated glucose uptake and cell proliferation in L-6 myoblasts, and the effects are mediated by integrin αvβ3. IGF-1 also behaves as a chemokine, being an important factor for tissue regeneration after damage. In the present study, using THP-1 human leukemic monocytes, expressing αvβ3 integrin in their cell membrane, we focused on the crosstalk between thyroid hormones and either IGF-1 or monocyte chemoattractant protein-1 (MCP-1), studying cell migration and proliferation stimulated by the two chemokines, and the role of αvβ3 integrin, using inhibitors of αvβ3 integrin and downstream pathways. Our results show that IGF-1 is a potent chemoattractant in THP-1 monocytes, stimulating cell migration, and thyroid hormone inhibits the effect through αvβ3 integrin. Thyroid hormone also inhibits IGF-1-stimulated cell proliferation through αvβ3 integrin, an example of a crosstalk between genomic and non-genomic effects. We also studied the effects of thyroid hormone on cell migration and proliferation induced by MCP-1, together with the pathways involved, by a pharmacological approach and docking simulation. Our findings show a different downstream signaling for IGF-1 and MCP-1 in THP-1 monocytes mediated by the plasma membrane receptor of thyroid hormones, integrin αvβ3.

Orthology and synteny analysis of receptor-like kinases “RLK” and receptor-like proteins “RLP” in legumes

Background Legume species are an important plant model because of their protein-rich physiology. The adaptability and productivity of legumes are limited by major biotic and abiotic stresses. Responses to these stresses directly involve plasma membrane receptor proteins known as receptor-like kinases and receptor-like proteins. Evaluating the homology relations among RLK and RLP for seven legume species, and exploring their presence among synteny blocks allow an increased understanding of evolutionary relations, physical position, and chromosomal distribution in related species and their shared roles in stress responses. Results Typically, a high proportion of RLK and RLP legume proteins belong to orthologous clusters, which is confirmed in this study, where between 66 to 90% of the RLKs and RLPs per legume species were classified in orthologous clusters. One-third of the evaluated syntenic blocks had shared RLK/RLP genes among both legumes and non-legumes. Among the legumes, between 75 and 98% of the RLK/RLP were present in syntenic blocks. The distribution of chromosomal segments between Phaseolus vulgaris and Vigna unguiculata, two species that diverged ~ 8 mya, were highly similar. Among the RLK/RLP synteny clusters, seven experimentally validated resistance RLK/RLP genes were identified in syntenic blocks. The RLK resistant genes FLS2, BIR2, ERECTA, IOS1, and AtSERK1 from Arabidopsis and SLSERK1 from Solanum lycopersicum were present in different pairwise syntenic blocks among the legume species. Meanwhile, only the LYM1- RLP resistant gene from Arabidopsis shared a syntenic blocks with Glycine max. Conclusions The orthology analysis of the RLK and RLP suggests a dynamic evolution in the legume family, with between 66 to 85% of RLK and 83 to 88% of RLP belonging to orthologous clusters among the species evaluated. In fact, for the 10-species comparison, a lower number of singleton proteins were reported among RLP compared to RLK, suggesting that RLP positions are more physically conserved compared to RLK. The identification of RLK and RLP genes among the synteny blocks in legumes revealed multiple highly conserved syntenic blocks on multiple chromosomes. Additionally, the analysis suggests that P. vulgaris is an appropriate anchor species for comparative genomics among legumes.

Ipomoeassin-F inhibits the in vitro biogenesis of the SARS-CoV-2 spike protein and its host cell membrane receptor

ABSTRACTIn order to produce proteins essential for their propagation, many pathogenic human viruses, including SARS-CoV-2, the causative agent of COVID-19 respiratory disease, commandeer host biosynthetic machineries and mechanisms. Three major structural proteins, the spike, envelope and membrane proteins, are amongst several SARS-CoV-2 components synthesised at the endoplasmic reticulum (ER) of infected human cells prior to the assembly of new viral particles. Hence, the inhibition of membrane protein synthesis at the ER is an attractive strategy for reducing the pathogenicity of SARS-CoV-2 and other obligate viral pathogens. Using an in vitro system, we demonstrate that the small molecule inhibitor ipomoeassin F (Ipom-F) potently blocks the Sec61-mediated ER membrane translocation and/or insertion of three therapeutic protein targets for SARS-CoV-2 infection; the viral spike and ORF8 proteins together with angiotensin-converting enzyme 2, the host cell plasma membrane receptor. Our findings highlight the potential for using ER protein translocation inhibitors such as Ipom-F as host-targeting, broad-spectrum antiviral agents.This article has an associated First Person interview with the first author of the paper.

Ipomoeassin-F inhibits the in vitro biogenesis of the SARS-CoV-2 spike protein and its host cell membrane receptor

In order to produce proteins essential for their propagation, many pathogenic human viruses, including SARS-CoV-2 the causative agent of COVID-19 respiratory disease, commandeer host biosynthetic machineries and mechanisms. Three major structural proteins, the spike, envelope and membrane proteins, are amongst several SARS-CoV-2 components synthesised at the endoplasmic reticulum (ER) of infected human cells prior to the assembly of new viral particles. Hence, the inhibition of membrane protein synthesis at the ER is an attractive strategy for reducing the pathogenicity of SARS-CoV-2 and other obligate viral pathogens. Using an in vitro system, we demonstrate that the small molecule inhibitor ipomoeassin F (Ipom-F) potently blocks the Sec61-mediated ER membrane translocation/insertion of three therapeutic protein targets for SARS-CoV-2 infection; the viral spike and ORF8 proteins together with angiotensin-converting enzyme 2, the host cell plasma membrane receptor. Our findings highlight the potential for using ER protein translocation inhibitors such as Ipom-F as host-targeting, broad-spectrum, antiviral agents.

The IL1-IL1RAP axis plays an important role in the inflammatory leukemic niche that favors acute myeloid leukemia proliferation over normal hematopoiesis.

Upregulation of the plasma membrane receptor IL1RAP in Acute Myeloid Leukemia (AML) has been reported but its role in the context of the leukemic bone marrow niche is unclear. Here, we studied the signaling events downstream of IL1RAP in relation to leukemogenesis and normal hematopoiesis. High IL1RAP expression was associated with a leukemic GMP-like state, and knockdown of IL1RAP in AML reduced colony-forming capacity. Stimulation with IL1β resulted in the induction of multiple chemokines and an inflammatory secretome via the p38 MAPK and NFκB signaling pathways in IL1RAP-expressing AML cells, but IL1β-induced signaling was dispensable for AML cell proliferation and NFκB-driven survival. IL1RAP was also expressed in stromal cells where IL1β induced expression of inflammatory chemokines and cytokines as well. Intriguingly, the IL1β-induced inflammatory secretome of IL1RAPexpressing AML cells grown on a stromal layer of mesenchymal stem cells affected normal hematopoiesis including hematopoietic stem/progenitor cells while AML cell proliferation was not affected. The addition of Anakinra, an FDA-approved IL1 receptor antagonist, could reverse this effect. Therefore, blocking the IL1-IL1RAP signaling axis might be a good therapeutic approach to reduce inflammation in the bone marrow niche and thereby promote normal hematopoietic recovery over AML proliferation after chemotherapy.

Activation of Rab11a and endocytosis by Phosphatidylinositol 4-kinase III beta promotes oncogenic signaling in breast cancer

ABSTRACTEndosomes are now recognized as important sites for regulating signal transduction. Here we show that the lipid kinase phosphatidylinositol 4-kinase III beta (PI4KIIIβ) regulates both endocytic kinetics and receptor signaling in breast cancer cells. PI4KIIIβ generates phosphatidylinositol 4-phosphate from phosphatidylinositol and is highly expressed in a subset of breast cancers. However, the molecular mechanism by which PI4KIIIβ promotes breast cancer is unclear. We demonstrate that ectopic PI4KIIIβ expression increases the rates of both endocytic internalization and recycling. PI4KIIIβ deletion reduces endocytic kinetics accompanied by a concomitant decrease in activity of the Rab11a GTPase, a protein required for endocytic function. Finally, we find that PI4KIIIβ activates IGF-IRβ signaling dependent on endosome function. Regulation of endocytic function by PI4KIIIβ is independent of its kinase activity but requires interaction with the Rab11a. This suggests that PI4KIIIβ controls endosomal kinetics and signaling by directly modulating Rab11a function. Our work suggests a novel regulatory role for PI4KIIIβ in endosome function and plasma membrane receptor signaling.

Plasma membrane receptor-like kinases and transporters are associated with 2,4-D resistance in wild radish

Background and Aims Resistance to the synthetic auxin 2,4-dichlorophenoxyacetic acid (2,4-D) in wild radish (Raphanus raphanistrum) appears to be due to a complex, multifaceted mechanism possibly involving enhanced constitutive plant defence and alterations in auxin signalling. Based on a previous gene expression analysis highlighting the plasma membrane as being important for 2,4-D resistance, this study aimed to identify the components of the leaf plasma membrane proteome that contribute to resistance. Methods Isobaric tagging of peptides was used to compare the plasma membrane proteomes of a 2,4-D-susceptible and a 2,4-D-resistant wild radish population under control and 2,4-D-treated conditions. Eight differentially abundant proteins were then targeted for quantification in the plasma membranes of 13 wild radish populations (two susceptible, 11 resistant) using multiple reaction monitoring. Key Results Two receptor-like kinases of unknown function (L-type lectin domain-containing receptor kinase IV.1-like and At1g51820-like) and the ATP-binding cassette transporter ABCB19, an auxin efflux transporter, were identified as being associated with auxinic herbicide resistance. The variability between wild radish populations suggests that the relative contributions of these candidates are different in the different populations. Conclusions To date, no receptor-like kinases have been reported to play a role in 2,4-D resistance. The lectin-domain-containing kinase may be involved in perception of 2,4-D at the plasma membrane, but its ability to bind 2,4-D and the identity of its signalling partner(s) need to be confirmed experimentally. ABCB19 is known to export auxinic compounds, but its role in 2,4-D resistance in wild radish appears to be relatively minor.

Iron Depletion Reduces Abce1 Transcripts While Inducing the Mitophagy Factors Pink1 and Parkin

Lifespan extension was recently achieved in Caenorhabditis elegans nematodes by mitochondrial stress and mitophagy, triggered via iron depletion. Conversely in man, deficient mitophagy due to Pink1/Parkin mutations triggers iron accumulation in patient brain and limits survival. We now aimed to identify murine fibroblast factors, which adapt their mRNA expression to acute iron manipulation, relate to mitochondrial dysfunction and may influence survival. After iron depletion, expression of the plasma membrane receptor Tfrc with its activator Ireb2, the mitochondrial membrane transporter Abcb10, the heme-release factor Pgrmc1, the heme-degradation enzyme Hmox1, the heme-binding cholesterol metabolizer Cyp46a1, as well as the mitophagy regulators Pink1 and Parkin showed a negative correlation to iron levels. After iron overload, these factors did not change expression. Conversely, a positive correlation of mRNA levels with both conditions of iron availability was observed for the endosomal factors Slc11a2 and Steap2, as well as for the iron-sulfur-cluster (ISC)-containing factors Ppat, Bdh2 and Nthl1. Positive correlation only after iron depletion was observed for the iron export factor Slc40a1, mitochondrial iron transporters Slc25a28, Abcb7 and Abcb8, mitochondrial ISC-containing factors Glrx5, Nfu1, Bola1 and Abce1, cytosolic Aco1 and Tyw5, as well as nuclear Dna2, Elp3, Pold1 and Prim2. The latter are regulators of nucleotide synthesis and DNA quality control, which have known importance for growth and lifespan. The only Pink1-/- triggered transcript modulation was the reduced expression of the ISC-containing ribosomal factor Abce1. These mammalian findings support previous fly data that Pink1 influences co-translational quality control via Abce1, as well as mitophagy. Our findings provide the first systematic survey how iron dosage triggers homeostatic transcriptional regulations and elucidate how iron deprivation results in mitophagy.