Accurate identification of tumor-derived somatic variants in plasma circulating cell-free DNA (cfDNA) requires understanding the various biologic compartments contributing to the cfDNA pool. We sought to define the technical feasibility of a high-intensity sequencing assay of cfDNA and matched white-blood cell (WBC) DNA covering a large genomic region (508 genes, 2Mb, >60,000X raw-depth) in a prospective study of 124 metastatic cancer patients, with contemporaneous matched tumor tissue biopsies, and 47 non-cancer controls. The assay displayed a high sensitivity and specificity, allowing for de novo detection of tumor-derived mutations and inference of tumor mutational burden, microsatellite instability, mutational signatures and sources of somatic mutations identified in cfDNA. The vast majority of cfDNA mutations (81.6% in controls and 53.2% in cancer patients) had features consistent with clonal hematopoiesis (CH). This cfDNA sequencing approach revealed that CH constitutes a pervasive biological phenomenon emphasizing the importance of matched cfDNA-WBC sequencing for accurate variant interpretation.
Villin/gelsolin/fragmin superfamily proteins have been shown to function in tip-growing plant cells. However, genes encoding gelsolin/fragmin do not exist in the Arabidopsis thaliana and rice (Oryza sativa) databases, and it is possible that these proteins are encoded by villin mRNA splicing variants. We cloned a 1006-bp full-length cDNA from Lilium longiflorum that encodes a 263-amino acid predicted protein sharing 100% identity with the N terminus of 135-ABP (Lilium villin) except for six C-terminal amino acids. The deduced 29-kD protein, Lilium ACTIN BINDING PROTEIN29 (ABP29), contains only the G1 and G2 domains and is the smallest identified member of the villin/gelsolin/fragmin superfamily. The purified recombinant ABP29 accelerates actin nucleation, blocks barbed ends, and severs actin filaments in a Ca 2þ -and/or phosphatidylinositol 4,5-bisphosphate-regulated manner in vitro. Microinjection of the protein into stamen hair cells disrupted transvacuolar strands whose backbone is mainly actin filament bundles. Transient expression of ABP29 by microprojectile bombardment of lily pollen resulted in actin filament fragmentation and inhibited pollen germination and tube growth. Our results suggest that ABP29 is a splicing variant of Lilium villin and a member of the villin/gelsolin/fragmin superfamily, which plays important roles in rearrangement of the actin cytoskeleton during pollen germination and tube growth.
Pleiotropic developmental regulators have been repeatedly linked to the evolution of anatomical novelties. Known mechanisms include cis-regulatory DNA changes that alter regulator transcription patterns or modify target-gene linkages. Here, we examine the role of another form of regulation, translational control, in the repeated evolution of self-fertile hermaphroditism in Caenorhabditis nematodes. Caenorhabditis elegans hermaphrodites initiate spermatogenesis in an otherwise female body through translational repression of the gene tra-2. This repression is mediated by GLD-1, an RNA-binding protein also required for oocyte meiosis and differentiation. By contrast, we show that in the convergently hermaphroditic Caenorhabditis briggsae, GLD-1 acts to promote oogenesis. The opposite functions of gld-1 in these species are not gene-intrinsic, but instead result from the unique contexts for its action that evolved in each. In C. elegans, GLD-1 became essential for promoting XX spermatogenesis via changes in the tra-2 mRNA and evolution of the species-specific protein FOG-2. C. briggsae GLD-1 became an essential repressor of sperm-promoting genes, including Cbr-puf-8, and did not evolve a strong association with tra-2. Despite its variable roles in sex determination, the function of gld-1 in female meiotic progression is ancient and conserved. This conserved role may explain why gld-1 is repeatedly recruited to regulate hermaphroditism. We conclude that, as with transcription factors, spatially localized translational regulators play important roles in the evolution of anatomical novelties.germ cells | translation | breeding systems | mutant
Esophageal squamous cell carcinoma (ESCC) is one of the most common malignant tumors with poor survival. It is urgent to search for new efficient drugs with good stability and safety for clinical therapy. This study aims to identify potential anticancer drugs from a compound library consisting of 429 natural products. Echinatin, a compound isolated from the Chinese herb Glycyrrhiza uralensis Fisch, was found to markedly induce apoptosis and inhibit proliferation and colony-formation ability in ESCC. Confocal fluorescence microscopy data showed that echinatin significantly induced autophagy in ESCC cells, and autophagy inhibitor bafilomycinA1 attenuated the suppressive effects of echinatin on cell viability and apoptosis. Mechanistically, RNA sequencing coupled with bioinformatics analysis and a series of functional assays revealed that echinatin induced apoptosis and autophagy through inactivation of AKT/mTOR signaling pathway, whereas constitutive activation of AKT significantly abrogated these effects. Furthermore, we demonstrated that echinatin had a significant antitumor effect in the tumor xenograft model and markedly suppressed cell migration and invasion abilities of ESCC cells in a dose-dependent manner. Our findings provide the first evidence that echinatin could be a novel therapeutic strategy for treating ESCC.
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