Biology subjects

Miller, K. E.

Publications and source records attributed to Miller, K. E..

23 records · Page 2Linked to original sources

bioRxiv2021

Discovery of Clinically Relevant Fusions in Pediatric Cancer

AO_SCPLOWBSTRACTC_SCPLOWO_ST_ABSBackgroundC_ST_ABSPediatric cancers typically have a distinct genomic landscape when compared to adult cancers and frequently carry somatic gene fusion events that alter gene expression and drive tumorigenesis. Sensitive and specific detection of gene fusions through the analysis of next-generation-based RNA sequencing (RNA-Seq) data is computationally challenging and may be confounded by low tumor cellularity or underlying genomic complexity. Furthermore, numerous computational tools are available to identify fusions from supporting RNA-Seq reads, yet each algorithm demonstrates unique variability in sensitivity and precision, and no clearly superior approach currently exists. To overcome these challenges, we have developed an ensemble fusion calling approach to increase the accuracy of identifying fusions. ResultsOur ensemble fusion detection approach utilizes seven fusion calling algorithms: Arriba, CICERO, FusionMap, FusionCatcher, JAFFA, MapSplice, and STAR-Fusion, which are packaged as a fully automated pipeline using Docker and AWS serverless technology. This method uses paired end RNA-Seq sequence reads as input, and the output from each algorithm is examined to identify fusions detected by a consensus of at least three algorithms. These consensus fusion results are filtered by comparison to an internal database to remove likely artifactual fusions occurring at high frequencies in our internal cohort, while a "known fusion list" prevents failure to report known pathogenic events. We have employed the ensemble fusion-calling pipeline on RNA-Seq data from 229 patients with pediatric cancer or blood disorders studied under an IRB-approved protocol. The samples consist of 138 central nervous system tumors, 73 solid tumors, and 18 hematologic malignancies or disorders. The combination of an ensemble fusion-calling pipeline and a knowledge-based filtering strategy identified 67 clinically relevant fusions among our cohort (diagnostic yield of 29.3%), including RBPMS-MET, BCAN-NTRK1, and TRIM22-BRAF fusions. Following clinical confirmation and reporting in the patients medical record, both known and novel fusions provided medically meaningful information. ConclusionsOur ensemble fusion detection pipeline offers a streamlined approach to discover fusions in cancer, at higher levels of sensitivity and accuracy than single algorithm methods. Furthermore, this method accurately identifies driver fusions in pediatric cancer, providing clinical impact by contributing evidence to diagnosis and, when appropriate, indicating targeted therapies.

genomics↗

bioRxiv2020

Sequestration of the exocytic SNARE Psy1 into multiprotein nodes reinforces polarized morphogenesis in fission yeast

Polarized morphogenesis is achieved by targeting or inhibiting growth at distinct regions. Rod-shaped fission yeast cells grow exclusively at their ends by restricting exocytosis and secretion to these sites. This growth pattern implies the existence of mechanisms that prevent exocytosis and growth along non-growing cell sides. We previously identified a set of 50-100 megadalton-sized node structures along the sides of fission yeast cells that contain the interacting proteins Skb1 and Slf1. Here, we show that Skb1-Slf1 nodes contain the syntaxin-like SNARE Psy1, which mediates exocytosis in fission yeast. Psy1 localizes in a diffuse pattern at cell tips where it likely promotes exocytosis and growth, but Psy1 is sequestered in Skb1-Slf1 nodes at cell sides where growth does not occur. Mutations that prevent node assembly or inhibit Psy1 localization to nodes lead to aberrant exocytosis at cell sides and increased cell width. Genetic results indicate that this Psy1 node mechanism acts in parallel to actin cables and Cdc42 regulation. Our work suggests that sequestration of syntaxin-like Psy1 at non-growing regions of the cell cortex reinforces cell morphology by restricting exocytosis to proper sites of polarized growth.

cell biology↗

bioRxiv2020

A Critical Role of Hepatic GABA in The Metabolic Dysfunction and Hyperphagia of Obesity

Hepatic lipid accumulation is a hallmark of type II diabetes (T2D) and associated with hyperinsulinemia, insulin resistance, and hyperphagia. Hepatic synthesis of GABA, catalyzed by GABA-transaminase (GABA-T), is upregulated in obese mice. To assess the role of hepatic GABA production in obesity-induced metabolic and energy dysregulation, we treated mice with two pharmacologic GABA-T inhibitors and knocked down hepatic GABA-T expression using an antisense oligonucleotide. Hepatic GABA-T inhibition and knockdown decreased basal hyperinsulinemia and hyperglycemia, and improved glucose intolerance. GABA-T knockdown improved insulin sensitivity assessed by hyperinsulinemic-euglycemic clamps in obese mice. Hepatic GABA-T knockdown also decreased food intake and induced weight loss without altering energy expenditure in obese mice. Data from people with obesity support the notion that hepatic GABA production and transport are associated with serum insulin, HOMA-IR, T2D, and BMI. These results support a key role for hepatocyte GABA production in the dysfunctional glucoregulation and feeding behavior associated with obesity.

physiology↗

bioRxiv2019

Temporal regulation of cell polarity via the interaction of the Ras GTPase Rsr1 and the scaffold protein Bem1

Establishing cell polarity is critical for growth and development of most organisms. The Cdc42 GTPase plays a central role in polarity development in species ranging from yeast to humans. In budding yeast, a specific growth site (i.e. bud site) is selected in the G1 phase, which determines the axis of cell polarization. Rsr1, a Ras GTPase, interacts with Cdc42 and its associated proteins to promote polarized growth at the proper bud site. Yet the mechanism underlying spatial cue-directed cell polarization is not fully understood. Here, we show that Rsr1 associates with Bem1, a scaffold protein, preferentially in its GDP-bound state in early G1. This interaction involves a part of the Bem1 Phox homology (PX) domain, which overlaps with a region previously shown to interact with Exo70, an exocyst component. Furthermore, overexpression of the constitutively GDP-bound Rsr1 interferes with Bem1s association with Exo70 and inhibits Bem1-dependent Exo70 polarization. We propose that Rsr1 plays a delicate role in coordination of spatial and temporal regulation of polarity establishment via its GTP- and GDP-bound states.

cell biology↗

bioRxiv2018

The contribution of mitochondrial metagenomics to largescale data mining and phylogenetic analysis of Coleoptera

A phylogenetic tree at the species level is still far off for highly diverse insect orders, including the Coleoptera, but the taxonomic breadth of public sequence databases is growing. In addition, new types of data may contribute to increasing taxon coverage, such as metagenomic shotgun sequencing for assembly of mitogenomes from bulk specimen samples. The current study explores the application of these techniques for large-scale efforts to build the tree of Coleoptera. We used shotgun data from 17 different ecological and taxonomic datasets (5 unpublished) to assemble a total of 1942 mitogenome contigs of >3000 bp. These sequences were combined into a single dataset together with all mitochondrial data available at GenBank, in addition to nuclear markers widely used in molecular phylogenetics. The resulting matrix of nearly 16000 species with two or more loci produced trees (RAxML) showing overall congruence with the Linnaean taxonomy at hierarchical levels from suborders to genera. We tested the role of full-length mitogenomes in stabilizing the tree from GenBank data, as mitogenomes might link terminals with non-overlapping gene representation. However, the mitogenome data were only partly useful in this respect, presumably because of the purely automated approach to assembly and gene delimitation, but improvements in future may be possible by using multiple assemblers and manual curation. In conclusion, the combination of data mining and metagenomic sequencing of bulk samples provided the largest phylogenetic tree of Coleoptera to date, which represents a summary of existing phylogenetic knowledge and a defensible tree of great utility, in particular for studies at the intra-familial level, despite some shortcomings for resolving basal nodes.

evolutionary biology↗