Demonstration of targeted crossovers in hybrid maize using CRISPR technology

Naturally occurring chromosomal crossovers (CO) during meiosis are a key driver of genetic diversity. The ability to target CO at specific allelic loci in hybrid plants would provide an advantage to the plant breeding process by facilitating trait introgression, and potentially increasing the rate of genetic gain. We present the first demonstration of targeted CO in hybrid maize utilizing the CRISPR Cas12a system. Our experiments showed that stable and heritable targeted CO can be produced in F1 somatic cells using Cas12a at a significantly higher rate than the natural CO in the same interval. Molecular characterization of the recombinant plants demonstrated that the targeted CO were driven by the non-homologous end joining (NHEJ) or HDR repair pathways, presumably during the mitotic cell cycle. These results are a step towards the use of RNA-guided nuclease technology to simplify the creation of targeted genome combinations in progeny and accelerate breeding.

Single-cell monitoring of dry mass and dry density reveals exocytosis of cellular dry contents in mitosis

Cell mass and composition change with cell cycle progression. Our previous work characterized buoyant mass accumulation dynamics in mitosis (Miettinen et al., 2019), but how dry mass and cell composition change in mitosis has remained unclear. To better understand mitotic cell growth and compositional changes, we develop a single-cell approach for monitoring dry mass and the density of that dry mass every ~75 seconds with 1.3% and 0.3% measurement precision, respectively. We find that suspension grown mammalian cells lose dry mass and increase dry density following mitotic entry. These changes display large, non-genetic cell-to-cell variability, and the changes are reversed at metaphase-anaphase transition, after which dry mass continues accumulating. The change in dry density causes buoyant and dry mass to differ specifically in early mitosis, thus reconciling existing literature on mitotic cell growth. Mechanistically, the dry composition changes do not require mitotic cell swelling or elongation. Instead, cells in early mitosis increase lysosomal exocytosis, and inhibition of exocytosis prevents the dry composition from changing. Overall, our work provides a new approach for monitoring single-cell dry mass and composition and reveals that mitosis is coupled to extensive exocytosis-mediated secretion of cellular contents.

Combined effect of cell geometry and polarity domains determines the orientation of unequal division

Cell division orientation is thought to result from a competition between cell geometry and polarity domains controlling the position of the mitotic spindle during mitosis. Depending on the level of cell shape anisotropy or the strength of the polarity domain, one dominates the other and determines the orientation of the spindle. Whether and how such competition is also at work to determine unequal cell division (UCD), producing daughter cells of different size, remains unclear. Here, we show that cell geometry and polarity domains cooperate, rather than compete, in positioning the cleavage plane during UCDs in early ascidian embryos. We found that the UCDs and their orientation at the ascidian third cleavage rely on the spindle tilting in an anisotropic cell shape, and cortical polarity domains exerting different effects on spindle astral microtubules. By systematically varying mitotic cell shape, we could modulate the effect of attractive and repulsive polarity domains and consequently generate predicted daughter cell size asymmetries and position. We therefore propose that the spindle position during UCD is set by the combined activities of cell geometry and polarity domains, where cell geometry modulates the effect of cortical polarity domain(s).

XDeathDB: a visualization platform for cell death molecular interactions

Lots of cell death initiator and effector molecules, signalling pathways and subcellular sites have been identified as key mediators in both cell death processes in cancer. The XDeathDB visualization platform provides a comprehensive cell death and their crosstalk resource for deciphering the signaling network organization of interactions among different cell death modes associated with 1461 cancer types and COVID-19, with an aim to understand the molecular mechanisms of physiological cell death in disease and facilitate systems-oriented novel drug discovery in inducing cell deaths properly. Apoptosis, autosis, efferocytosis, ferroptosis, immunogenic cell death, intrinsic apoptosis, lysosomal cell death, mitotic cell death, mitochondrial permeability transition, necroptosis, parthanatos, and pyroptosis related to 12 cell deaths and their crosstalk can be observed systematically by the platform. Big data for cell death gene-disease associations, gene-cell death pathway associations, pathway-cell death mode associations, and cell death-cell death associations is collected by literature review articles and public database from iRefIndex, STRING, BioGRID, Reactom, Pathway’s commons, DisGeNET, DrugBank, and Therapeutic Target Database (TTD). An interactive webtool, XDeathDB, is built by web applications with R-Shiny, JavaScript (JS) and Shiny Server Iso. With this platform, users can search specific interactions from vast interdependent networks that occur in the realm of cell death. A multilayer spectral graph clustering method that performs convex layer aggregation to identify crosstalk function among cell death modes for a specific cancer. 147 hallmark genes of cell death could be observed in detail in these networks. These potential druggable targets are displayed systematically and tailoring networks to visualize specified relations is available to fulfil user-specific needs. Users can access XDeathDB for free at https://pcm2019.shinyapps.io/XDeathDB/.

A Case Report of a Bleeding Duodenal Gastro-Intestinal Stromal Tumor and its Emergent Management

A gastrointestinal stromal tumor (GIST) arising in the duodenum is a rare subtype of mesenchymal tumor. GISTs have a reported incidence of 11.9–19.6 per million population and duodenal GISTs make up just 5% of these tumors. Common presentation of duodenal GISTs is through an upper gastrointestinal bleed, of which, they are responsible for less than 1% of all gastrointestinal bleeding. In an elective setting, surgical management remains the mainstay of treatment. In this Case Report, the emergent management of a localized duodenal resection was performed by an acute care surgeon, in an unstable patient. The risk factors for malignancy include tumor size and a high mitotic cell index. Tumor recurrence is determined by tumor size, tumor rupture, high mitotic cell index, a non-gastric location and gastrointestinal bleeding.

Mitotic Outcomes in Fibrous Environments

During mitosis, cells round up and generate outward forces to create space and orient the mitotic spindles. Here, using suspended ECM-mimicking nanofiber networks, we recapitulate in vivo adhesion organization and confinement to interrogate mitotic outcomes for various interphase cell shapes. Elongated cells attached to single fibers through two focal adhesion clusters (FACs) at their extremities result in perfect spherical mitotic cell bodies that undergo large 3D displacement while being held by retraction fibers. Increasing the number of parallel fibers increases cellular extremity FACs and retraction fiber-driven stability, leading to reduced 3D cell-body movement, metaphase plate rotations, and significantly faster division times. Interestingly, interphase kite shapes on a crosshatch pattern of four fibers undergo mitosis resembling single-fiber outcomes due to rounded bodies being primarily held in position by retraction fibers from two perpendicular suspended fibers. We develop a cortex-astral microtubule analytical friction and force model to capture retraction-fiber-driven stability of the metaphase plate rotations. We report that reduced orientational stability results in increased monopolar mitotic defects. In the case of cells attached to two parallel fibers, rounded mitotic cells can get confined between the suspended fibers, allowing estimation of the mitotic forces through measurement of the outward deflection of the fibers. Interestingly, confinement causes rotated mitotic spindles similar to those reported in dense tissues. Overall, we establish dynamics of mitosis in fibrous environments governed by fiber arrangement and architecture-driven differences in interphase cell shapes, adhesion geometries, and varying levels of mechanical confinement.

Bioinformatics Analysis Identifies Precision Treatment with Paclitaxel for Hepatocellular Carcinoma Patients Harboring Mutant TP53 or Wild-Type CTNNB1 Gene

Hepatocellular carcinoma (HCC) is an aggressive and chemoresistant cancer type. The development of novel therapeutic strategies is still urgently needed. Personalized or precision medicine is a new trend in cancer therapy, which treats cancer patients with specific genetic alterations. In this study, a gene signature was identified from the transcriptome of HCC patients, which was correlated with the patients’ poorer prognoses. This gene signature is functionally related to mitotic cell cycle regulation, and its higher or lower expression is linked to the mutation in tumor protein p53 (TP53) or catenin beta 1 (CTNNB1), respectively. Gene–drug association analysis indicated that the taxanes, such as the clinically approved anticancer drug paclitaxel, are potential drugs targeting this mitotic gene signature. Accordingly, HCC cell lines harboring mutant TP53 or wild-type CTNNB1 genes are more sensitive to paclitaxel treatment. Therefore, our results imply that HCC patients with mutant TP53 or wild-type CTNNB1 genes may benefit from the paclitaxel therapy.

Multiple Omics Analysis Reveals E2F5 Predict Prognostic Biomarker in Diffuse Large B-Cell Lymphoma

Diffuse large B-cell lymphoma (DLBCL) is highly aggressive and fatal hematological malignancy. There are few biomarkers that can be used to predict the survival of DLBCL patients. Therefore, there is an urgent need to find new biological targets to improve the predictive value and sensitive diagnosis of DLBCL.E2F family play an essential role in tumorigenesis, however, remains obscure in DLBCL.E2F transcription factor family(E2Fs) mRNA expression between DLBCL and nonmalignant samples were screened by GEPIA,CCLE and EMBL-EBI. The associated regulation pathway in DLBLC was established using the GeneMANIA,Metascape ,SMATAPP database. Transcription analysis indicated E2F1/4/5/8 mRNA expression was significantly higher in patients and the cell lines.What’s more, the high E2F5/8 expression had significantly lower survival rate. Further functional analysis showed that E2F1/3/5 were hypomethylated in DLBCL,which may associated with patient chemo-resistance. Subsequently,these genes with their co-expression genes mainly formed transcription factor complex, regulated G1/S transition of mitotic cell cycle and through TGF-beta signaling pathway to participate DLBCL tumorigenesis. This results demonstrate that E2F5 were potential prognostic biomarkers for better survival of DLBCL patients.

iASPP contributes to cell cortex rigidity, mitotic cell rounding, and spindle positioning

iASPP is a protein mostly known as an inhibitor of p53 pro-apoptotic activity and a predicted regulatory subunit of the PP1 phosphatase, which is often overexpressed in tumors. We report that iASPP associates with the microtubule plus-end binding protein EB1, a central regulator of microtubule dynamics, via an SxIP motif. iASPP silencing or mutation of the SxIP motif led to defective microtubule capture at the cortex of mitotic cells, leading to abnormal positioning of the mitotic spindle. These effects were recapitulated by the knockdown of the membrane-to-cortex linker Myosin-Ic (Myo1c), which we identified as a novel partner of iASPP. Moreover, iASPP or Myo1c knockdown cells failed to round up upon mitosis because of defective cortical stiffness. We propose that by increasing cortical rigidity, iASPP helps cancer cells maintain a spherical geometry suitable for proper mitotic spindle positioning and chromosome partitioning.

Functional Genomics of PRUNE1 in Neurodevelopmental Disorders (NDDs) Tied to Medulloblastoma (MB) and Other Tumors

We analyze the fundamental functions of Prune_1 in brain pathophysiology. We discuss the importance and maintenance of the function of Prune_1 and how its perturbation influences both brain pathological conditions, neurodevelopmental disorder with microcephaly, hypotonia, and variable brain anomalies (NMIHBA; OMIM: 617481), and tumorigenesis of medulloblastoma (MB) with functional correlations to other tumors. A therapeutic view underlying recent discoveries identified small molecules and cell penetrating peptides to impair the interaction of Prune_1 with protein partners (e.g., Nm23-H1), thus further impairing intracellular and extracellular signaling (i.e., canonical Wnt and TGF-β pathways). Identifying the mechanism of action of Prune_1 as responsible for neurodevelopmental disorders (NDDs), we have recognized other genes which are found overexpressed in brain tumors (e.g., MB) with functional implications in neurodevelopmental processes, as mainly linked to changes in mitotic cell cycle processes. Thus, with Prune_1 being a significant target in NDDs, we discuss how its network of action can be dysregulated during brain development, thus generating cancer and metastatic dissemination.