Efficient and generalizable prediction of molecular alterations in multiple cancer cohorts using H&E whole slide images.

Molecular testing of tumor samples for targetable biomarkers is restricted by a lack of standardization, turnaround-time, cost, and tissue availability across cancer types. Additionally, targetable alterations of low prevalence may not be tested in routine workflows. Algorithms that predict DNA alterations from routinely generated hematoxylin and eosin (H&E)-stained images could prioritize samples for confirmatory molecular testing.

Fetal fraction amplification within prenatal cfDNA screening enables detection of genome-wide copy-number variants at enhanced resolution.

Purpose: Clinically significant copy-number variants (CNVs) occur in 1% to 2% of pregnancies and are difficult to detect via prenatal cell-free DNA (cfDNA) screening because of the low fraction of fetal-derived cfDNA in maternal plasma. Here, we use fetal fraction amplification (FFA) and improved computational algorithms to enhance the resolution and sensitivity of CNV detection.

Methods: We implemented and characterized the performance of a hidden Markov model that identifies fetal CNVs.

Validation of an Expanded Carrier Screen that Optimizes Sensitivity via Full-Exon Sequencing and Panel-wide Copy Number Variant Identification.

Background: By identifying pathogenic variants across hundreds of genes, expanded carrier screening (ECS) enables prospective parents to assess the risk of transmitting an autosomal recessive or X-linked condition. Detection of at-risk couples depends on the number of conditions tested, the prevalence of the respective diseases, and the screen's analytical sensitivity for identifying disease-causing variants. Disease-level analytical sensitivity is often <100% in ECS tests because copy number variants (CNVs) are typically not interrogated because of their technical complexity.

Machine-Learning-Based Analysis in Genome-Edited Cells Reveals the Efficiency of Clathrin-Mediated Endocytosis.

Cells internalize various molecules through clathrin-mediated endocytosis (CME). Previous live-cell imaging studies suggested that CME is inefficient, with about half of the events terminated. These CME efficiency estimates may have been confounded by overexpression of fluorescently tagged proteins and inability to filter out false CME sites.

Centrosomes. Regulated assembly of a supramolecular centrosome scaffold in vitro.

The centrosome organizes microtubule arrays within animal cells and comprises two centrioles surrounded by an amorphous protein mass called the pericentriolar material (PCM). Despite the importance of centrosomes as microtubule-organizing centers, the mechanism and regulation of PCM assembly are not well understood. In Caenorhabditis elegans, PCM assembly requires the coiled-coil protein SPD-5.

Actin and dynamin2 dynamics and interplay during clathrin-mediated endocytosis.

Clathrin-mediated endocytosis (CME) involves the recruitment of numerous proteins to sites on the plasma membrane with prescribed timing to mediate specific stages of the process. However, how choreographed recruitment and function of specific proteins during CME is achieved remains unclear. Using genome editing to express fluorescent fusion proteins at native levels and live-cell imaging with single-molecule sensitivity, we explored dynamin2 stoichiometry, dynamics, and functional interdependency with actin.

Caulobacter chromosome in vivo configuration matches model predictions for a supercoiled polymer in a cell-like confinement.

We measured the distance between fluorescent-labeled DNA loci of various interloci contour lengths in Caulobacter crescentus swarmer cells to determine the in vivo configuration of the chromosome. For DNA segments less than about 300 kb, the mean interloci distances, , scale as n(0.22), where n is the contour length, and cell-to-cell distribution of the interloci distance r is a universal function of r/n(0.

Compaction and transport properties of newly replicated Caulobacter crescentus DNA.

Upon initiating replication of the Caulobacter chromosome, one copy of the parS centromere remains at the stalked pole; the other moves to the distal pole. We identified the segregation dynamics and compaction characteristics of newly replicated Caulobacter DNA during transport (highly variable from cell to cell) using time-lapse fluorescence microscopy. The parS centromere and a length (also highly variable) of parS proximal DNA on each arm of the chromosome are segregated with the same relatively slow transport pattern as the parS locus.

The three-dimensional architecture of a bacterial genome and its alteration by genetic perturbation.

We have determined the three-dimensional (3D) architecture of the Caulobacter crescentus genome by combining genome-wide chromatin interaction detection, live-cell imaging, and computational modeling. Using chromosome conformation capture carbon copy (5C), we derive ~13 kb resolution 3D models of the Caulobacter genome. The resulting models illustrate that the genome is ellipsoidal with periodically arranged arms.

Assembly of the Caulobacter cell division machine.

Cytokinesis in Gram-negative bacteria is mediated by a multiprotein machine (the divisome) that invaginates and remodels the inner membrane, peptidoglycan and outer membrane. Understanding the order of divisome assembly would inform models of the interactions among its components and their respective functions. We leveraged the ability to isolate synchronous populations of Caulobacter crescentus cells to investigate assembly of the divisome and place the arrival of each component into functional context.

High-throughput identification of protein localization dependency networks.

Bacterial cells are highly organized with many protein complexes and DNA loci dynamically positioned to distinct subcellular sites over the course of a cell cycle. Such dynamic protein localization is essential for polar organelle development, establishment of asymmetry, and chromosome replication during the Caulobacter crescentus cell cycle. We used a fluorescence microscopy screen optimized for high-throughput to find strains with anomalous temporal or spatial protein localization patterns in transposon-generated mutant libraries.

Caulobacter PopZ forms a polar subdomain dictating sequential changes in pole composition and function.

The bacterium Caulobacter crescentus has morphologically and functionally distinct cell poles that undergo sequential changes during the cell cycle. We show that the PopZ oligomeric network forms polar ribosome exclusion zones that change function during cell cycle progression. The parS/ParB chromosomal centromere is tethered to PopZ at one pole prior to the initiation of DNA replication.

Caulobacter requires a dedicated mechanism to initiate chromosome segregation.

Chromosome segregation in bacteria is rapid and directed, but the mechanisms responsible for this movement are still unclear. We show that Caulobacter crescentus makes use of and requires a dedicated mechanism to initiate chromosome segregation. Caulobacter has a single circular chromosome whose origin of replication is positioned at one cell pole.