Ionophore-stimulation promotes re-organization of the invasion machinery of Toxoplasma gondii

Host cell invasion by intracellular, eukaryotic parasites, like the many important species within the phylum Apicomplexa, is a remarkable and active process involving the coordinated action of many apical organelles and other structures. To date, capturing how these various structures interact during invasion has been difficult to observe in detail. Here, we used cryogenic electron tomography to generate images of the apical complex of Toxoplasma gondii tachyzoites under conditions that mimic resting parasites and those primed to invade through addition of a calcium ionophore. Using AI-based image-processing we were able to annotate 48 tomograms to identify and extract densities of the relevant subcellular organelles and accurately analyze features in 3D. We describe an interaction between an anteriorly located apical vesicle and a rhoptry tip that occurs only in the ionophore-stimulated parasites and that is associated with dramatic changes in the vesicle's shape in what appears to be a stalled fusion event. We also present information to support the presumption that this vesicle originates from the well-described vesicles that parallel the intraconoidal microtubules and that the latter two structures are linked by a novel tether. Lastly, we show that a previously described rosette is found associated with more than just the anterior-most apical vesicle, indicating that multiple such vesicles are primed to enable rhoptry secretion.

Natural Chromosome-Chromid Fusion across rRNA Operons in a Burkholderiaceae Bacterium

A bacterial chromosome that was naturally fused with the secondary chromosome, or “chromid,” and presented as an unexpectedly large single replicon was discovered in the genome of Cupriavidus necator strain KK10, a biotechnologically useful member of the family Burkholderiaceae . Although Burkholderiaceae is a well-documented group that conserves chromids in their genomes, this chromosomal fusion event has not been previously reported for this family.

A functionally conserved STORR gene fusion in Papaver species that diverged 16.8 million years ago

The STORR gene fusion event is considered a key step in the evolution of benzylisoquinoline alkaloid (BIA) metabolism in opium poppy as the resulting bi-modular protein performs the isomerization of (S)- to (R)- reticuline which is required for morphinan biosynthesis. Our previous analysis of the opium poppy genome suggested the STORR gene fusion event occurred before a whole genome duplication event 7.2 million years ago. Here we use a combination of phylogenetic, transcriptomic, metabolomic, biochemical and genomic analysis to investigate the origin of the STORR gene fusion across the Papaveraceae family. The pro-morphinan/morphinan subclass of BIAs was present in a subset of 10 Papaver species including P. somniferum (opium poppy) and this correlated with the presence of the STORR gene fusion with one important exception. P. californicum does not produce morphinans but it does contain a STORR gene fusion that epimerizes (S)- to (R)- reticuline when heterologously expressed in yeast. The high similarity of the amino acid sequence linking the two modules of STORR along with phylogenetic gene tree analysis strongly suggests the gene fusion occurred only once and between 17-25 million years ago before the separation of P. californicum from the other Papaver species. We discovered that the most abundant BIA in P. californicum is (R)- glaucine, a member of the aporphine subclass of BIAs. Only the (S) isomer of this compound has previously been reported from nature. These results lead us to conclude that the function of the STORR gene fusion is not exclusive to morphinan production in the Papaveraceae.

Full-length transcriptome analysis and identification of transcript structures in Eimeria necatrix from different developmental stages by single-molecule real-time sequencing

Background Eimeria necatrix is one of the most pathogenic parasites, causing high mortality in chickens. Although its genome sequence has been published, the sequences and complete structures of its mRNA transcripts remain unclear, limiting exploration of novel biomarkers, drug targets and genetic functions in E. necatrix. Methods Second-generation merozoites (MZ-2) of E. necatrix were collected using Percoll density gradients, and high-quality RNA was extracted from them. Single-molecule real-time (SMRT) sequencing and Illumina sequencing were combined to generate the transcripts of MZ-2. Combined with the SMRT sequencing data of sporozoites (SZ) collected in our previous study, the transcriptome and transcript structures of E. necatrix were studied. Results SMRT sequencing yielded 21,923 consensus isoforms in MZ-2. A total of 17,151 novel isoforms of known genes and 3918 isoforms of novel genes were successfully identified. We also identified 2752 (SZ) and 3255 (MZ-2) alternative splicing (AS) events, 1705 (SZ) and 1874 (MZ-2) genes with alternative polyadenylation (APA) sites, 4019 (SZ) and 2588 (MZ-2) fusion transcripts, 159 (SZ) and 84 (MZ-2) putative transcription factors (TFs) and 3581 (SZ) and 2039 (MZ-2) long non-coding RNAs (lncRNAs). To validate fusion transcripts, reverse transcription-PCR was performed on 16 candidates, with an accuracy reaching up to 87.5%. Sanger sequencing of the PCR products further confirmed the authenticity of chimeric transcripts. Comparative analysis of transcript structures revealed a total of 3710 consensus isoforms, 815 AS events, 1139 genes with APA sites, 20 putative TFs and 352 lncRNAs in both SZ and MZ-2. Conclusions We obtained many long-read isoforms in E. necatrix SZ and MZ-2, from which a series of lncRNAs, AS events, APA events and fusion transcripts were identified. Information on TFs will improve understanding of transcriptional regulation, and fusion event data will greatly improve draft versions of gene models in E. necatrix. This information offers insights into the mechanisms governing the development of E. necatrix and will aid in the development of novel strategies for coccidiosis control. Graphical Abstract

A high-quality chromosome-scale assembly of the centipedegrass [Eremochloa ophiuroides (Munro) Hack.] genome provides insights into chromosomal structural evolution and prostrate growth habit

Centipedegrass [Eremochloa ophiuroides (Munro) Hack.], a member of the Panicoideae subfamily, is one of the most important warm-season turfgrasses originating from China. This grass has an extremely developed prostrate growth habit and has been widely used in transitional and warm climatic regions. To better understand the genetic basis of important biological characteristics, such as prostrate growth and seed yield, in warm-season turfgrasses, we present a high-quality reference genome for centipedegrass and use PacBio, BioNano, and Hi-C technologies to anchor the 867.43 Mb genome assembly into nine pseudochromosomes, with a scaffold N50 of 86.05 Mb and 36,572 annotated genes. Centipedegrass was most closely related to sorghum and diverged from their common ancestor ~16.8 Mya. We detected a novel chromosome reshuffling event in centipedegrass, namely, the nest chromosome fusion event in which fusion of chromosomes 8 and 10 of sorghum into chromosome 3 of centipedegrass likely occurred after the divergence of centipedegrass from sorghum. The typical prostrate growth trait in centipedegrass may be linked to the expansion of candidate PROSTRATE GROWTH 1 (PROG1) genes on chromosome 2. Two orthologous genes of OsPROG1, EoPROG1, and EoPROG2, were confirmed to increase the stem number and decrease the stem angle in Arabidopsis. Collectively, our assembled reference genome of centipedegrass offers new knowledge and resources to dissect the genome evolution of Panicoideae and accelerate genome-assisted breeding and improvement of plant architecture in turf plants.

KDM5A suppresses PML-RARα target gene expression and APL differentiation through repressing H3K4me2

Epigenetic abnormalities are frequently involved in the initiation and progression of cancers, including acute myeloid leukemia (AML). A subtype of AML, acute promyelocytic leukemia (APL), is mainly driven by a specific oncogenic fusion event of promyelocytic leukemia–RA receptor fusion oncoprotein (PML-RARα). PML-RARα was reported as a transcription repressor through the interaction with nuclear receptor corepressor and histone deacetylase complexes leading to the mis-suppression of its target genes and differentiation blockage. Although previous studies were mainly focused on the connection of histone acetylation, it is still largely unknown whether alternative epigenetics mechanisms are involved in APL progression. KDM5A is a demethylase of histone H3 lysine 4 di- and tri-methylations (H3K4me2/3) and a transcription corepressor. Here, we found that the loss of KDM5A led to APL NB4 cell differentiation and retarded growth. Mechanistically, through epigenomics and transcriptomics analyses, KDM5A binding was detected in 1889 genes, with the majority of the binding events at promoter regions. KDM5A suppressed the expression of 621 genes, including 42 PML-RARα target genes, primarily by controlling the H3K4me2 in the promoters and 5′ end intragenic regions. In addition, a recently reported pan-KDM5 inhibitor, CPI-455, on its own could phenocopy the differentiation effects as KDM5A loss in NB4 cells. CPI-455 treatment or KDM5A knockout could greatly sensitize NB4 cells to all-trans retinoic acid–induced differentiation. Our findings indicate that KDM5A contributed to the differentiation blockage in the APL cell line NB4, and inhibition of KDM5A could greatly potentiate NB4 differentiation.

Lipid bilayer degradation induced by SARS-CoV-2 spike protein as revealed by neutron reflectometry

SARS-CoV-2 spike proteins are responsible for the membrane fusion event, which allows the virus to enter the host cell and cause infection. This process starts with the binding of the spike extramembrane domain to the angiotensin-converting enzyme 2 (ACE2), a membrane receptor highly abundant in the lungs. In this study, the extramembrane domain of SARS-CoV-2 Spike (sSpike) was injected on model membranes formed by supported lipid bilayers in presence and absence of the soluble part of receptor ACE2 (sACE2), and the structural features were studied at sub-nanometer level by neutron reflection. In all cases the presence of the protein produced a remarkable degradation of the lipid bilayer. Indeed, both for membranes from synthetic and natural lipids, a significant reduction of the surface coverage was observed. Quartz crystal microbalance measurements showed that lipid extraction starts immediately after sSpike protein injection. All measurements indicate that the presence of proteins induces the removal of membrane lipids, both in the presence and in the absence of ACE2, suggesting that sSpike molecules strongly associate with lipids, and strip them away from the bilayer, via a non-specific interaction. A cooperative effect of sACE2 and sSpike on lipid extraction was also observed.

Second Report of PDE10A-BRAF Fusion in Pediatric Spindle Cell Sarcoma With Infantile Fibrosarcoma-Like Morphology Suggesting PDE10A-BRAF Fusion Is a Recurrent Event

Infantile/congenital fibrosarcoma (IFS) is the most common soft tissue tumor in children less than one year of age. The most common anatomic site of IFS is in the extremities or trunk, and rarely in the abdomen or retroperitoneum. Approximately 70-90% of cases are characterized by a distinct t(12;15)(p13;q25) translocation resulting in an ETV6-NTRK3 gene fusion. As such, TRK inhibitors are considered frontline therapy in TRK-fusion positive IFS. The ETV6-NTRK3 fusion is also detected in congenital mesoblastic nephroma (CMN) and less frequently in myeloid leukemias, secretory breast carcinoma, and mammary-type secretory carcinoma of the skin and salivary glands. Infrequently, cases of tumors with IFS-like morphology without the characteristic ETV6-NTRK3 gene fusion have been identified. Herein, an ETV6-NTRK3 fusion negative spindle cell sarcoma with IFS-like morphology subjected to genomic profiling revealed a PDE10A-BRAF fusion, a fusion event that has been detected previously in an isolated case of undifferentiated infantile sarcoma.

Novel nitrite reductase domain structure suggests a chimeric denitrification repertoire in Phylum Chloroflexi

Denitrification plays a central role in the global nitrogen cycle, reducing and removing nitrogen from marine and terrestrial ecosystems. The flux of nitrogen species through this pathway has a widespread impact, affecting ecological carrying capacity, agriculture, and climate. Nitrite reductase (Nir) and nitric oxide reductase (NOR) are the two central enzymes in this pathway. Here we present a previously unreported Nir domain architecture in members of Phylum Chloroflexi. Phylogenetic analyses of protein domains within Nir indicate that an ancestral horizontal transfer and fusion event produced this chimeric domain architecture. We also identify an expanded genomic diversity of a rarely reported nitric oxide reductase subtype, eNOR. Together, these results suggest a greater diversity of denitrification enzyme arrangements exist than have been previously reported.