Showing papers on "Mutant published in 2021"
TL;DR: In this paper, the authors describe the identification of an antibody highly specific to the most common TP53 mutation (R175H, in which arginine at position 175 is replaced with histidine) in complex with a common human leukocyte antigen-A (HLA-A) allele on the cell surface.
Abstract: TP53 (tumor protein p53) is the most commonly mutated cancer driver gene, but drugs that target mutant tumor suppressor genes, such as TP53, are not yet available. Here, we describe the identification of an antibody highly specific to the most common TP53 mutation (R175H, in which arginine at position 175 is replaced with histidine) in complex with a common human leukocyte antigen–A (HLA-A) allele on the cell surface. We describe the structural basis of this specificity and its conversion into an immunotherapeutic agent: a bispecific single-chain diabody. Despite the extremely low p53 peptide-HLA complex density on the cancer cell surface, the bispecific antibody effectively activated T cells to lyse cancer cells that presented the neoantigen in vitro and in mice. This approach could in theory be used to target cancers containing mutations that are difficult to target in conventional ways.
146 citations
02 Mar 2021
TL;DR: In this article, a new strain is reported in the UK that includes a mutation (N501Y) in the receptor binding domain (RBD), which is possibly increasing the infection rate.
Abstract: SARS-CoV-2 is a global challenge due to its ability to spread much faster than the SARS-CoV, which was attributed to the mutations in the receptor binding domain (RBD). These mutations enhanced the electrostatic interactions. Recently, a new strain is reported in the UK that includes a mutation (N501Y) in the RBD, that is possibly increasing the infection rate. Here, using Molecular Dynamics simulations (MD) and Monte Carlo (MC) sampling, we show that the N501 mutation enhanced the electrostatic interactions due to the formation of a strong hydrogen bond between SARS-CoV-2-T500 and ACE2-D355 near the mutation site. In addition, we observed that the electrostatic interactions between the SARS-CoV-2 and ACE2 in the wild type and the mutant are dominated by salt-bridges formed between SARS-CoV-2-K417 and ACE2-D30, SARS-CoV-2-K458, ACE2-E23, and SARS-CoV-2-R403 and ACE2-E37. These interactions contributed more than 40% of the total binding energies.
137 citations
TL;DR: A review of treatments for cancers with mutant p53 that focus on directly targeting mutant P53, restoring wild-type functions, and exploiting synthetic lethal interactions with mutantP53 is presented in this paper.
Abstract: TP53 is a critical tumor-suppressor gene that is mutated in more than half of all human cancers. Mutations in TP53 not only impair its antitumor activity, but also confer mutant p53 protein oncogenic properties. The p53-targeted therapy approach began with the identification of compounds capable of restoring/reactivating wild-type p53 functions or eliminating mutant p53. Treatments that directly target mutant p53 are extremely structure and drug-species-dependent. Due to the mutation of wild-type p53, multiple survival pathways that are normally maintained by wild-type p53 are disrupted, necessitating the activation of compensatory genes or pathways to promote cancer cell survival. Additionally, because the oncogenic functions of mutant p53 contribute to cancer proliferation and metastasis, targeting the signaling pathways altered by p53 mutation appears to be an attractive strategy. Synthetic lethality implies that while disruption of either gene alone is permissible among two genes with synthetic lethal interactions, complete disruption of both genes results in cell death. Thus, rather than directly targeting p53, exploiting mutant p53 synthetic lethal genes may provide additional therapeutic benefits. Additionally, research progress on the functions of noncoding RNAs has made it clear that disrupting noncoding RNA networks has a favorable antitumor effect, supporting the hypothesis that targeting noncoding RNAs may have potential synthetic lethal effects in cancers with p53 mutations. The purpose of this review is to discuss treatments for cancers with mutant p53 that focus on directly targeting mutant p53, restoring wild-type functions, and exploiting synthetic lethal interactions with mutant p53. Additionally, the possibility of noncoding RNAs acting as synthetic lethal targets for mutant p53 will be discussed.
114 citations
TL;DR: The Spike glycoprotein receptor-binding domain (RBD) of SARS-CoV-2 mediates the viral particle's binding to the angiotensin-converting enzyme 2 (ACE2) receptor on the surface of human cells as mentioned in this paper.
Abstract: The Spike glycoprotein receptor-binding domain (RBD) of SARS-CoV-2 mediates the viral particle’s binding to the angiotensin-converting enzyme 2 (ACE2) receptor on the surface of human cells. Therefore, Spike-ACE2 interaction is a crucial determining factor for viral infectivity. A new phylogenetic group of SARS-CoV-2 (lineage B.1.1.7) has been recently identified in the COVID-19 Genomics UK Consortium dataset, which features an amino acid substitution in the Spike RBD (N501Y mutation). Infections with the SARS-CoV-2 lineage B.1.1.7 have been overgrowing in recent weeks in the United Kingdom, indicating an even greater spread capacity than that seen with previous strains of the novel coronavirus. We hypothesized that this rapid spreading/infectivity of the B.1.1.7 lineage might be due to changes in the interaction force between the mutant Spike RBD and ACE2. This study employed in silico methods involving mutagenesis (N501Y mutation) and interface analysis focusing on the Spike RDB-ACE2 interaction. The results showed that the SARS-CoV-2 N501Y mutant (lineage B.1.1.7) establishes a more significant number of interactions relating to the mutant residue Y501 (Spike RDB) with residues Y41 and K353 (ACE2). This finding shows that the increased infectivity of SARS-CoV-2 lineage B.1.1.7 is associated with the interaction force between the Spike RBD Y501 mutant residue with the ACE2 receptor, which in this strain is increased.
82 citations
TL;DR: In this article, a PCR-based, bacterium-free method was proposed to generate SARS-CoV-2 infectious clones. But the method was limited by the lack of convenient mutagenesis methods.
65 citations
TL;DR: Most mutations were caused by ultraviolet (UV) light, but mutational signature analysis suggested differences in DNA repair processes between sites, and analysis of hair follicles showed mutations in the upper follicle resembled adjacent skin, but the lower follicle was sparsely mutated.
Abstract: Skin cancer risk varies substantially across the body, yet how this relates to the mutations found in normal skin is unknown. Here we mapped mutant clones in skin from high- and low-risk sites. The density of mutations varied by location. The prevalence of NOTCH1 and FAT1 mutations in forearm, trunk, and leg skin was similar to that in keratinocyte cancers. Most mutations were caused by ultraviolet light, but mutational signature analysis suggested differences in DNA-repair processes between sites. Eleven mutant genes were under positive selection, with TP53 preferentially selected in the head and FAT1 in the leg. Fine-scale mapping revealed 10% of clones had copy-number alterations. Analysis of hair follicles showed mutations in the upper follicle resembled adjacent skin, but the lower follicle was sparsely mutated. Normal skin is a dense patchwork of mutant clones arising from competitive selection that varies by location. Significance: Mapping mutant clones across the body reveals normal skin is a dense patchwork of mutant cells. The variation in cancer risk between sites substantially exceeds that in mutant clone density. More generally, mutant genes cannot be assigned as cancer drivers until their prevalence in normal tissue is known. See related commentary by De Dominici and DeGregori, p. 227. This article is highlighted in the In This Issue feature, p. 211
52 citations
08 Mar 2021
TL;DR: To optimize the efficiency of the Cas9 nuclease in generating mutations in target genes in Arabidopsis thaliana, several features of its nucleotide and/or amino acid sequence are investigated, including the codon usage, the number of nuclear localization signals (NLSs), and the presence or absence of introns.
Abstract: The recent discovery of the mode of action of the CRISPR/Cas9 system has provided biologists with a useful tool for generating site-specific mutations in genes of interest. In plants, site-targeted mutations are usually obtained by the stable transformation of a Cas9 expression construct into the plant genome. The efficiency of introducing mutations in genes of interest can vary considerably depending on the specific features of the constructs, including the source and nature of the promoters and terminators used for the expression of the Cas9 gene and the guide RNA, and the sequence of the Cas9 nuclease itself. To optimize the efficiency of the Cas9 nuclease in generating mutations in target genes in Arabidopsis thaliana, we investigated several features of its nucleotide and/or amino acid sequence, including the codon usage, the number of nuclear localization signals (NLSs), and the presence or absence of introns. We found that the Cas9 gene codon usage had some effect on its activity and that two NLSs worked better than one. However, the highest efficiency of the constructs was achieved by the addition of 13 introns into the Cas9 coding sequence, which dramatically improved the editing efficiency of the constructs. None of the primary transformants obtained with a Cas9 gene lacking introns displayed a knockout mutant phenotype, whereas between 70% and 100% of the primary transformants generated with the intronized Cas9 gene displayed mutant phenotypes. The intronized Cas9 gene was also found to be effective in other plants such as Nicotiana benthamiana and Catharanthus roseus.
51 citations
TL;DR: It is suggested that OsTTG1 is an important gene resource for the genetic engineering of anthocyanin biosynthesis in rice and other plants and directional selection has driven the evolutionary divergence of the indica and japonica Os TTG1 alleles.
Abstract: Anthocyanins play an important role in the growth of plants, and are beneficial to human health. In plants, the MYB-bHLH-WD40 (MBW) complex activates the genes for anthocyanin biosynthesis. However, in rice, the WD40 regulators remain to be conclusively identified. Here, a crucial anthocyanin biosynthesis gene was fine mapped to a 43.4-kb genomic region on chromosome 2, and a WD40 gene OsTTG1 (Oryza sativa TRANSPARENT TESTA GLABRA1) was identified as ideal candidate gene. Subsequently, a homozygous mutant (osttg1) generated by CRISPR/Cas9 showed significantly decreased anthocyanin accumulation in various rice organs. OsTTG1 was highly expressed in various rice tissues after germination, and it was affected by light and temperature. OsTTG1 protein was localized to the nucleus, and can physically interact with Kala4, OsC1, OsDFR and Rc. Furthermore, a total of 59 hub transcription factor genes might affect rice anthocyanin biosynthesis, and LOC_Os01g28680 and LOC_Os02g32430 could have functional redundancy with OsTTG1. Phylogenetic analysis indicated that directional selection has driven the evolutionary divergence of the indica and japonica OsTTG1 alleles. Our results suggest that OsTTG1 is a vital regulator of anthocyanin biosynthesis, and an important gene resource for the genetic engineering of anthocyanin biosynthesis in rice and other plants.
48 citations
TL;DR: This article showed that mutations that limit ABA synthesis or signaling reduce the extent of secondary cell-wall thickness and lignification in Arabidopsis thaliana through the core ABA-signaling pathway involving SnRK2 kinases.
Abstract: Plant secondary cell-wall (SCW) deposition and lignification are affected by both seasonal factors and abiotic stress, and these responses may involve the hormone abscisic acid (ABA). However, the mechanisms involved are not clear. Here we show that mutations that limit ABA synthesis or signaling reduce the extent of SCW thickness and lignification in Arabidopsis thaliana through the core ABA-signaling pathway involving SnRK2 kinases. SnRK2.2. 3 and 6 physically interact with the SCW regulator NAC SECONDARY WALL THICKENING PROMOTING FACTOR 1 (NST1), a NAC family transcription factor that orchestrates the transcriptional activation of a suite of downstream SCW biosynthesis genes, some of which are involved in the biosynthesis of cellulose and lignin. This interaction leads to phosphorylation of NST1 at Ser316, a residue that is highly conserved among NST1 proteins from dicots, but not monocots, and is required for transcriptional activation of downstream SCW-related gene promoters. Loss of function of NST1 in the snd1 mutant background results in lack of SCWs in the interfascicular fiber region of the stem, and the Ser316Ala mutant of NST1 fails to complement this phenotype and ABA-induced lignin pathway gene expression. The discovery of NST1 as a key substrate for phosphorylation by SnRK2 suggests that the ABA-mediated core-signaling cascade provided land plants with a hormone-modulated, competitive desiccation-tolerance strategy allowing them to differentiate water-conducting and supporting tissues built of cells with thicker cell walls.
47 citations
TL;DR: In this article, a combination of genetic and metabolite analysis was used to identify the role of myb21 in regulating flavonol biosynthesis through direct binding to the GARE cis-element in the FLS1 promoter.
Abstract: Commonly found flavonols in plants are synthesized from dihydroflavonols by flavonol synthase (FLS). The genome of Arabidopsis thaliana contains six FLS genes, among which FLS1 encodes a functional enzyme. Previous work has demonstrated that the R2R3-MYB subgroup 7 transcription factors MYB11, MYB12, and MYB111 redundantly regulate flavonol biosynthesis. However, flavonol accumulation in pollen grains was unaffected in the myb11myb12myb111 triple mutant. Here we show that MYB21 and its homologs MYB24 and MYB57, which belong to subgroup 19, promote flavonol biosynthesis through regulation of FLS1 gene expression. We used a combination of genetic and metabolite analysis to identify the role of MYB21 in regulating flavonol biosynthesis through direct binding to the GARE cis-element in the FLS1 promoter. Treatment with kaempferol or overexpression of FLS1 rescued stamen defects in the myb21 mutant. We also observed that excess reactive oxygen species (ROS) accumulated in the myb21 stamen, and that treatment with the ROS inhibitor diphenyleneiodonium chloride partly rescued the reduced fertility of the myb21 mutant. Furthermore, drought increased ROS abundance and impaired fertility in myb21, myb21myb24myb57, and chs, but not in the wild type or myb11myb12myb111, suggesting that pollen-specific flavonol accumulation contributes to drought-induced male fertility by ROS scavenging in Arabidopsis.
47 citations
TL;DR: This work identified a rice LMM, early lesion leaf 1 (ell1), cloned the causal gene by map-based cloning, and verified this by complementation, revealing that ELL1 affects chloroplast development or function, and that loss of E LL1 function induces ROS accumulation and lesion formation in rice.
Abstract: Lesion-mimic mutants (LMMs) provide a valuable tool to reveal the molecular mechanisms determining programmed cell death (PCD) in plants. Despite intensive research, the mechanisms behind PCD and the formation of lesions in various LMMs still remain to be elucidated. Here, we identified a rice (Oryza sativa) LMM, early lesion leaf 1 (ell1), cloned the causal gene by map-based cloning, and verified this by complementation. ELL1 encodes a cytochrome P450 monooxygenase, and the ELL1 protein was located in the endoplasmic reticulum. The ell1 mutant exhibited decreased chlorophyll contents, serious chloroplast degradation, upregulated expression of chloroplast degradation-related genes, and attenuated photosynthetic protein activity, indicating that ELL1 is involved in chloroplast development. RNA sequencing analysis showed that genes related to oxygen binding were differentially expressed in ell1 and wild-type plants; histochemistry and paraffin sectioning results indicated that hydrogen peroxide (H2 O2 ) and callose accumulated in the ell1 leaves, and the cell structure around the lesions was severely damaged, which indicated that reactive oxygen species (ROS) accumulated and cell death occurred in the mutant. TUNEL staining and comet experiments revealed that severe DNA degradation and abnormal PCD occurred in the ell1 mutants, which implied that excessive ROS accumulation may induce DNA damage and ROS-mediated cell death in the mutant. Additionally, lesion initiation in the ell1 mutant was light dependent and temperature sensitive. Our findings revealed that ELL1 affects chloroplast development or function, and that loss of ELL1 function induces ROS accumulation and lesion formation in rice.
TL;DR: In this paper, the authors identify a disease-linked mutation for hereditary angioedema (HAE), which is associated with a genetic deficiency of functional C1 inhibitors or with normal C1 inhibitor (HAEnCI).
Abstract: Background Hereditary angioedema (HAE) is a potentially fatal disorder resulting in recurrent attacks of severe swelling. It may be associated with a genetic deficiency of functional C1 inhibitor or with normal C1 inhibitor (HAEnCI). In families with HAEnCI, HAE-linked mutations in the F12, PLG, KNG1, ANGPT1, or MYOF genes have been identified. In many families with HAEnCI the genetic cause of the disease is currently unknown. Objective The aim of this study was to identify a novel disease-linked mutation for HAEnCI. Methods The study methods comprised whole exome sequencing, Sanger sequencing analysis, pedigree analysis, bioinformatic analysis of the mutation, and biochemical analysis of parameters of the kallikrein-kinin (contact) system. Results By performing whole exome sequencing on a multigenerational family with HAEnCI we were able to identify the heparan sulfate (HS)-glucosamine 3-O-sulfotransferase 6 (HS3ST6) mutation c.430A>T (p.Thr144Ser) in all 3 affected family members who were sequenced. This gene encodes HS-glucosamine 3-O-sulfotransferase 6 (3-OST-6), which is involved in the last step of HS biosynthesis. The p.Thr144Ser mutation is likely to affect the interaction between 2 β-sheets stabilizing the active center of the 3-OST-6 protein. Conclusions We conclude that mutant 3-OST-6 fails to transfer sulfo groups to the 3-OH position of HS, resulting in incomplete HS biosynthesis. This likely affects cell surface interactions of key players in angioedema formation and is a novel mechanism for disease development.
TL;DR: In this article, the small molecule REV1-Polζ was used to treat BRCA1/2 mutant tumor cells and showed that it is preferentially toxic toward HR-deficient cancer cells.
TL;DR: In this article, the authors used a protein mimetic to abrogate mutant p53 aggregation and rescue p53 function, which inhibits cancer cell proliferation in vitro and halts tumor growth in vivo.
Abstract: Missense mutations in p53 are severely deleterious and occur in over 50% of all human cancers. The majority of these mutations are located in the inherently unstable DNA-binding domain (DBD), many of which destabilize the domain further and expose its aggregation-prone hydrophobic core, prompting self-assembly of mutant p53 into inactive cytosolic amyloid-like aggregates. Screening an oligopyridylamide library, previously shown to inhibit amyloid formation associated with Alzheimer’s disease and type II diabetes, identified a tripyridylamide, ADH-6, that abrogates self-assembly of the aggregation-nucleating subdomain of mutant p53 DBD. Moreover, ADH-6 targets and dissociates mutant p53 aggregates in human cancer cells, which restores p53’s transcriptional activity, leading to cell cycle arrest and apoptosis. Notably, ADH-6 treatment effectively shrinks xenografts harboring mutant p53, while exhibiting no toxicity to healthy tissue, thereby substantially prolonging survival. This study demonstrates the successful application of a bona fide small-molecule amyloid inhibitor as a potent anticancer agent. Amyloid aggregation of mutant p53 contributes to its loss of tumor suppressor function and oncogenic gain-of-function. Here, the authors use a protein mimetic to abrogate mutant p53 aggregation and rescue p53 function, which inhibits cancer cell proliferation in vitro and halts tumor growth in vivo.
TL;DR: In this article, the direct impact of epigenetic modifications and temperature stress on mitotic mutation rates in a fungal pathogen using a mutation accumulation approach was determined, and it was shown that epigenetic modification and environmental conditions significantly increased the mutation rate.
Abstract: Mutations are the source of genetic variation and the substrate for evolution. Genome-wide mutation rates appear to be affected by selection and are probably adaptive. Mutation rates are also known to vary along genomes, possibly in response to epigenetic modifications, but causality is only assumed. In this study we determine the direct impact of epigenetic modifications and temperature stress on mitotic mutation rates in a fungal pathogen using a mutation accumulation approach. Deletion mutants lacking epigenetic modifications confirm that histone mark H3K27me3 increases whereas H3K9me3 decreases the mutation rate. Furthermore, cytosine methylation in transposable elements (TE) increases the mutation rate 15-fold resulting in significantly less TE mobilization. Also accessory chromosomes have significantly higher mutation rates. Finally, we find that temperature stress substantially elevates the mutation rate. Taken together, we find that epigenetic modifications and environmental conditions modify the rate and the location of spontaneous mutations in the genome and alter its evolutionary trajectory. While a correlation between epigenetic modifications and mutation rates has been observed, experimental evidence of causality is limited. Here the authors measure the mutation rate in fungal mutants lacking histone modifications and confirm experimentally a causal effect of epigenetic modifications on mutation rates.
TL;DR: In this paper, a comprehensive genome-wide analysis of the LACS gene family across green plants was performed, followed by phylogenetic clustering analysis, gene structure determination, detection of conserved motifs, gene expression in tissues and subcellular localization.
TL;DR: In this paper, the authors showed that IDH1/2 mutant-derived 2-hydroxyglutarate is synthetically lethal to TET-dioxygenase deficient cells.
Abstract: TET2 is frequently mutated in myeloid neoplasms. Genetic TET2 deficiency leads to skewed myeloid differentiation and clonal expansion, but minimal residual TET activity is critical for survival of neoplastic progenitor and stem cells. Consistent with mutual exclusivity of TET2 and neomorphic IDH1/2 mutations, here we report that IDH1/2 mutant-derived 2-hydroxyglutarate is synthetically lethal to TET-dioxygenase deficient cells. In addition, a TET-selective small molecule inhibitor decreased cytosine hydroxymethylation and restricted clonal outgrowth of TET2 mutant, but not normal hematopoietic precursor cells in vitro and in vivo. While TET-inhibitor phenocopied somatic TET2 mutations, its pharmacologic effects on normal stem cells were, unlike mutations, reversible. Treatment with TET inhibitor suppressed the clonal evolution of TET2 mutant cells in murine models and TET2-mutated human leukemia xenografts. These results suggest that TET inhibitors may constitute a new class of targeted agents in TET2 mutant neoplasia.
French Institute of Health and Medical Research1, École nationale vétérinaire de Toulouse2, Institut national de la recherche agronomique3, University of Texas MD Anderson Cancer Center4, University of Pennsylvania5, University of Montpellier6, Agios Pharmaceuticals7, Barcelona Supercomputing Center8
TL;DR: In this article, the authors showed that patients with acute myeloid leukemia (AML) harboring an IDH mutation displayed an enhanced mitochondrial oxidative metabolism, along with an increase in TCA cycle intermediates, this AML-specific metabolic behavior mechanistically occurred through the increase in electron transport chain complex I activity, mitochondrial respiration, and methylation-driven CEBPα-induced fatty acid β-oxidation of IDH1 mutant cells.
Abstract: Mutations in IDH induce epigenetic and transcriptional reprogramming, differentiation bias, and susceptibility to mitochondrial inhibitors in cancer cells. Here, we first show that cell lines, PDXs, and patients with acute myeloid leukemia (AML) harboring an IDH mutation displayed an enhanced mitochondrial oxidative metabolism. Along with an increase in TCA cycle intermediates, this AML-specific metabolic behavior mechanistically occurred through the increase in electron transport chain complex I activity, mitochondrial respiration, and methylation-driven CEBPα-induced fatty acid β-oxidation of IDH1 mutant cells. While IDH1 mutant inhibitor reduced 2-HG oncometabolite and CEBPα methylation, it failed to reverse FAO and OxPHOS. These mitochondrial activities were maintained through the inhibition of Akt and enhanced activation of peroxisome proliferator-activated receptor-γ coactivator-1 PGC1α upon IDH1 mutant inhibitor. Accordingly, OxPHOS inhibitors improved anti-AML efficacy of IDH mutant inhibitors in vivo. This work provides a scientific rationale for combinatory mitochondrial-targeted therapies to treat IDH mutant AML patients, especially those unresponsive to or relapsing from IDH mutant inhibitors.
TL;DR: Significant modifications to cell wall composition in the psl1 mutant as compared to the wild type plants were identified and enhanced drought tolerance of the mutant plants by reducing water loss under osmotic stress and drought conditions.
Abstract: The biosynthesis and modification of cell wall composition and structure are controlled by hundreds of enzymes and have a direct consequence on plant growth and development. However, the majority of these enzymes has not been functionally characterised. Rice mutants with leaf-rolling phenotypes were screened in a field. Phenotypic analysis under controlled conditions was performed for the selected mutant and the relevant gene was identified by map-based cloning. Cell wall composition was analysed by glycome profiling assay. We identified a photo-sensitive leaf rolling 1 (psl1) mutant with 'napping' (midday depression of photosynthesis) phenotype and reduced growth. The PSL1 gene encodes a cell wall-localised polygalacturonase (PG), a pectin-degrading enzyme. psl1 with a 260-bp deletion in its gene displayed leaf rolling in response to high light intensity and/or low humidity. Biochemical assays revealed PG activity of recombinant PSL1 protein. Significant modifications to cell wall composition in the psl1 mutant compared with the wild-type plants were identified. Such modifications enhanced drought tolerance of the mutant plants by reducing water loss under osmotic stress and drought conditions. Taken together, PSL1 functions as a PG that modifies cell wall biosynthesis, plant development and drought tolerance in rice.
TL;DR: In this paper, the miR164-GhCUC2 (CUP-SHAPED COTYLEDON2) module regulates lateral shoot development in cotton and Arabidopsis.
Abstract: Branching determines cotton architecture and production, but the underlying regulatory mechanisms remain unclear. Here, we report that the miR164-GhCUC2 (CUP-SHAPED COTYLEDON2) module regulates lateral shoot development in cotton and Arabidopsis. We generated OE-GhCUC2m (overexpression GhCUC2m) and STTM164 (short tandem target mimic RNA of miR164) lines in cotton and heterologous expression lines for gh-miR164, GhCUC2 and GhCUC2m in Arabidopsis to study the mechanisms controlling lateral branching. GhCUC2m overexpression resulted in a short-branch phenotype similar to STTM164. In addition, heterologous expression of GhCUC2m led to decreased number and length of branches compared with wild type, opposite to the effects of the OE-gh-pre164 line in Arabidopsis. GhCUC2 interacted with GhBRC1 and exhibited similar negative regulation of branching. Overexpression of GhBRC1 in the brc1-2 mutant partially rescued the mutant phenotype and decreased branch number. GhBRC1 directly bound to the NCED1 promoter and activated its transcription, leading to local abscisic acid (ABA) accumulation and response. Mutation of the NCED1 promoter disrupted activation by GhBRC1. This finding demonstrates a direct relationship between BRC1 and ABA signalling and places ABA downstream of BRC1 in the control of branching development. The miR164-GhCUC2-GhBRC1-GhNCED1 module provides a clear regulatory axis for ABA signalling to control plant architecture.
TL;DR: In this paper, small molecule inhibitors targeting the PI3K p110α catalytic subunit have entered clinical trials, with early-phase GDC-0077 (Inavolisib) studies showing anti-tumor activity and a manageable safety profile in patients with PIK3CA-mutant, hormone receptor-positive breast cancer as a single agent or in combination therapy.
Abstract: PIK3CA is one of the most frequently mutated oncogenes; the p110α protein it encodes plays a central role in tumor cell proliferation and survival. Small molecule inhibitors targeting the PI3K p110α catalytic subunit have entered clinical trials, with early-phase GDC-0077 (Inavolisib) studies showing anti-tumor activity and a manageable safety profile in patients with PIK3CA-mutant, hormone receptor-positive breast cancer as a single agent or in combination therapy. Despite this, preclinical studies have shown that PI3K pathway inhibition releases negative feedback and activates receptor tyrosine kinase signaling, reengaging the pathway and attenuating drug activity. Here we discover that GDC-0077 and taselisib more potently inhibit mutant PI3K pathway signaling and cell viability through unique HER2-dependent degradation. Both are more effective than other PI3K inhibitors at maintaining prolonged pathway suppression, resulting in enhanced apoptosis and greater efficacy. This unique mechanism against mutant p110α reveals a new strategy for creating inhibitors that specifically target mutant tumors with selective degradation of the mutant oncoprotein and also provide a strong rationale for pursuing PI3Kα degraders in patients with HER2-positive breast cancer.
TL;DR: In this article, the authors described rice YUCCA (YUC) flavin-containing monooxygenase encoding gene OsYUC11 as a key contributor to auxin biosynthesis in rice (Oryza sativa) endosperm.
Abstract: Auxin is a phytohormone essential for plant development. However, our understanding of auxin-regulated endosperm development remains limited. Here, we described rice YUCCA (YUC) flavin-containing monooxygenase encoding gene OsYUC11 as a key contributor to auxin biosynthesis in rice (Oryza sativa) endosperm. Grain filling or storage product accumulation was halted by mutation of OsYUC11, but the deficiencies could be recovered by the exogenous application of auxin. A rice transcription factor (TF) yeast library was screened, and 41 TFs that potentially bind to the OsYUC11 promoter were identified, of which OsNF-YB1, a member of the nuclear factor Y family, is predominantly expressed in the endosperm. Both osyuc11 and osnf-yb1 mutants exhibited reduced seed size and increased chalkiness, accompanied by a reduction in indole-3-acetic acid biosynthesis. OsNF-YB1 can bind the OsYUC11 promoter to induce gene expression in vivo. We also found that OsYUC11 was a dynamically imprinted gene that predominantly expressed the paternal allele in the endosperm up to 10 d after fertilization (DAF) but then became a non-imprinted gene at 15 DAF. A functional maternal allele of OsYUC11 was able to recover the paternal defects of this gene. Overall, the findings indicate that OsYUC11-mediated auxin biosynthesis is essential for endosperm development in rice.
TL;DR: In this article, the role of SOD1 in cancer is not fully understood, however, the generation of an inducible Sod1 knockout in KRAS-driven NSCLC mouse model is described.
Abstract: SOD1 is known as the major cytoplasmic superoxide dismutase and an anticancer target. However, the role of SOD1 in cancer is not fully understood. Herein we describe the generation of an inducible Sod1 knockout in KRAS-driven NSCLC mouse model. Sod1 knockout markedly reduces tumor burden in vivo and blocks growth of KRAS mutant NSCLC cells in vitro. Intriguingly, SOD1 is enriched in the nucleus and notably in the nucleolus of NSCLC cells. The nuclear and nucleolar, not cytoplasmic, form of SOD1 is essential for lung cancer cell proliferation. Moreover, SOD1 interacts with PeBoW complex and controls its assembly necessary for pre-60S ribosomal subunit maturation. Mechanistically, SOD1 regulates co-localization of PeBoW with and processing of pre-rRNA, and maturation of cytoplasmic 60S ribosomal subunits in KRAS mutant lung cancer cells. Collectively, our study unravels a nuclear SOD1 function essential for ribosome biogenesis and proliferation in KRAS-driven lung cancer.
TL;DR: In this article, the authors identified and characterized the narrow leaf21 (nal21) mutant in rice (Oryza sativa), showing a significant reduction in leaf width, leaf length and plant height, and increased tiller number.
Abstract: Leaf morphology influences photosynthesis, transpiration, and ultimately crop yield. However, the molecular mechanism of leaf development is still not fully understood. Here, we identified and characterized the narrow leaf21 (nal21) mutant in rice (Oryza sativa), showing a significant reduction in leaf width, leaf length and plant height, and increased tiller number. Microscopic observation revealed defects in the vascular system and reduced epidermal cell size and number in the nal21 leaf blade. Map-based cloning revealed that NAL21 encodes a ribosomal small subunit protein RPS3A. Ribosome-targeting antibiotics resistance assay and ribosome profiling showed a significant reduction in the free 40S ribosome subunit in the nal21 mutant. The nal21 mutant showed aberrant auxin responses in which multiple auxin response factors (ARFs) harboring upstream open-reading frames (uORFs) in their 5'-untranslated region were repressed at the translational level. The WUSCHEL-related homeobox 3A (OsWOX3A) gene, a key transcription factor involved in leaf blade lateral outgrowth, is also under the translational regulation by RPS3A. Transformation with modified OsARF11, OsARF16, and OsWOX3A genomic DNA (gDNA) lacking uORFs rescued the narrow leaf phenotype of nal21 to a better extent than transformation with their native gDNA, implying that RPS3A could regulate translation of ARFs and WOX3A through uORFs. Our results demonstrate that proper translational regulation of key factors involved in leaf development is essential to maintain normal leaf morphology.
TL;DR: In this paper, the role of mutant p53 proteins in regulating different autophagic pathways and further discuss the available evidence suggesting a possible autophagy stimulatory role of mutants.
Abstract: One of the most notoriously altered genes in human cancer is the tumor-suppressor TP53, which is mutated with high frequency in more cancers than any other tumor suppressor gene Beyond the loss of wild-type p53 functions, mutations in the TP53 gene often lead to the expression of full-length proteins with new malignant properties Among the defined oncogenic functions of mutant p53 is its effect on cell metabolism and autophagy Due to the importance of autophagy as a stress adaptive response, it is frequently dysfunctional in human cancers However, the role of p53 is enigmatic in autophagy regulation While the complex action of the wild-type p53 on autophagy has extensively been described in literature, in this review, we focus on the conceivable role of distinct mutant p53 proteins in regulating different autophagic pathways and further discuss the available evidence suggesting a possible autophagy stimulatory role of mutant p53 Moreover, we describe the involvement of different autophagic pathways in targeting and degrading mutant p53 proteins, exploring the potential strategies of targeting mutant p53 in cancer by autophagy
TL;DR: The findings indicate that mutant ER causes several consistent effects on gene expression, both indirectly and through constant activity, which is still unclear regarding the effect of mutant ER beyond its estrogen independence.
Abstract: While breast cancer patients with tumors that express estrogen receptor α (ER) generally respond well to hormone therapies that block ER activity, a significant number of patients relapse. Approximately 30% of these recurrences harbor activating mutations in the ligand binding domain (LBD) of ER, which have been shown to confer ligand-independent function. However, much is still unclear regarding the effect of mutant ER beyond its estrogen independence. To investigate the molecular effects of mutant ER, we developed multiple isogenic ER-mutant cell lines for the most common LBD mutations, Y537S and D538G. These mutations induced differential expression of thousands of genes, the majority of which were mutant allele specific and were not observed upon estrogen treatment of wild-type (WT) cells. These mutant-specific genes showed consistent differential expression across ER-mutant lines developed in other laboratories. WT cells with long-term estrogen exposure only exhibited some of these transcriptional changes, suggesting that mutant ER causes novel regulatory effects that are not simply due to constant activity. While ER mutations exhibited minor effects on ER genomic binding, with the exception of ligand independence, ER mutations conferred substantial differences in chromatin accessibility. Mutant ER was bound to approximately a quarter of mutant-enriched accessible regions that were enriched for other DNA binding factors, including FOXA1, CTCF, and OCT1. Overall, our findings indicate that mutant ER causes several consistent effects on gene expression, both indirectly and through constant activity. Significance: This study demonstrates the multiple roles of mutant ER in breast cancer progression, including constant ER activity and secondary regulatory effects on gene expression and chromatin accessibility. Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/81/3/539/F1.large.jpg. See related commentary by Hermida-Prado and Jeselsohn, p. 537 See related article by Williams and colleagues, p. 732
TL;DR: Results showed that ZmRPP13-LK3, a new AC, could catalyse ATP for producing cAMP and may be involved in ABA-regulated heat resistance.
Abstract: In plants, 3´,5´-cyclic adenosine monophosphate (cAMP) is an important second messenger with varied functions; however, only a few adenylyl cyclases (ACs) that synthesize cAMP have been identified. Moreover, the biological roles of ACs/cAMP in response to stress remain largely unclear. In this study, we used quantitative proteomics techniques to identify a maize heat-induced putative disease-resistance RPP13-like protein 3 (ZmRPP13-LK3), which has three conserved catalytic AC centres. The AC activity of ZmRPP13-LK3 was confirmed by in vitro enzyme activity analysis, in vivo RNAi experiments, and functional complementation in the E. coli cyaA mutant. ZmRPP13-LK3 is located in the mitochondria. The results of in vitro and in vivo experiments indicated that ZmRPP13-LK3 interacts with ZmABC2, a possible cAMP exporter. Under heat stress, the concentrations of ZmRPP13-LK3 and cAMP in the ABA-deficient mutant vp5 were significantly less than those in the wild-type, and treatment with ABA and an ABA inhibitor affected ZmRPP13-LK3 expression in the wild-type. Application of 8-Br-cAMP, a cAMP analogue, increased heat-induced expression of heat-shock proteins in wild-type plants and alleviated heat-activated oxidative stress. Taken together, our results indicate that ZmRPP13-LK3, a new AC, can catalyse ATP for the production of cAMP and may be involved in ABA-regulated heat resistance.
TL;DR: In this article, a highly variable region at residues 203-205 in SARS-CoV-2 nucleocapsid protein was investigated and it was shown that the R203K/G204R mutation is sufficient to enhance replication, fitness, and pathogenesis.
Abstract: While SARS-CoV-2 continues to adapt for human infection and transmission, genetic variation outside of the spike gene remains largely unexplored. This study investigates a highly variable region at residues 203-205 in SARS-CoV-2 nucleocapsid protein. Recreating the alpha variant mutation in an early pandemic (WA-1) background, we found that the R203K/G204R mutation is sufficient to enhance replication, fitness, and pathogenesis of SARS-CoV-2. Importantly, the R203K/G204R mutation increases nucleocapsid phosphorylation, providing a molecular basis for these phenotypes. Notably, an analogous alanine substitution mutant also increases SARS-CoV-2 fitness and phosphorylation, suggesting that infection is enhanced through ablation of the ancestral 'RG' motif. Overall, these results demonstrate that variant mutations outside spike are also key components in SARS-CoV-2's continued adaptation to human infection. One-sentence summary A mutation in the nucleocapsid gene of the SARS-CoV-2 alpha variant is found to enhance replication, fitness, and pathogenesis.
TL;DR: In this paper, the authors found that ClNAC68 is a transcriptional repressor and that the repression domain is located in the C-terminus of the NAM gene.
Abstract: NAC (NAM, ATAF1/2, and CUC2) transcription factors play important roles in fruit ripening and quality. The watermelon genome encodes 80 NAC genes, and 21 of these NAC genes are highly expressed in both the flesh and vascular tissues. Among these genes, ClNAC68 expression was significantly higher in flesh than in rind. However, the intrinsic regulatory mechanism of ClNAC68 in fruit ripening and quality is still unknown. In this study, we found that ClNAC68 is a transcriptional repressor and that the repression domain is located in the C-terminus. Knockout of ClNAC68 by the CRISPR-Cas9 system decreased the soluble solid content and sucrose accumulation in mutant flesh. Development was delayed, germination was inhibited, and the IAA content was significantly decreased in mutant seeds. Transcriptome analysis showed that the invertase gene ClINV was the only gene involved in sucrose metabolism that was upregulated in mutant flesh, and expression of the indole-3-acetic acid-amido synthetase gene ClGH3.6 in the IAA signaling pathway was also induced in mutant seeds. EMSA and dual-luciferase assays showed that ClNAC68 directly bound to the promoters of ClINV and ClGH3.6 to repress their expression. These results indicated that ClNAC68 positively regulated sugar and IAA accumulation by repressing ClINV and ClGH3.6. Our findings provide new insights into the regulatory mechanisms by which NAC transcription factors affect fruit quality and seed development.
TL;DR: In this article, a genetically encoded, H3K27me3-specific intracellular antibody or H4K20me1-mintbody was developed to follow X chromosome inactivation (XCI) dynamics in living cells.
Abstract: During X chromosome inactivation (XCI), in female placental mammals, gene silencing is initiated by the Xist long non-coding RNA. Xist accumulation at the X leads to enrichment of specific chromatin marks, including PRC2-dependent H3K27me3 and SETD8-dependent H4K20me1. However, the dynamics of this process in relation to Xist RNA accumulation remains unknown as is the involvement of H4K20me1 in initiating gene silencing. To follow XCI dynamics in living cells, we developed a genetically encoded, H3K27me3-specific intracellular antibody or H3K27me3-mintbody. By combining live-cell imaging of H3K27me3, H4K20me1, the X chromosome and Xist RNA, with ChIP-seq analysis we uncover concurrent accumulation of both marks during XCI, albeit with distinct genomic distributions. Furthermore, using a Xist B and C repeat mutant, which still shows gene silencing on the X but not H3K27me3 deposition, we also find a complete lack of H4K20me1 enrichment. This demonstrates that H4K20me1 is dispensable for the initiation of gene silencing, although it may have a role in the chromatin compaction that characterises facultative heterochromatin.