A machine learning one-class logistic regression model to predict stemness for single cell transcriptomics and spatial omics.

Cell annotation is a crucial methodological component to interpreting single cell and spatial omics data. These approaches were developed for single cell analysis but are often biased, manually curated and yet unproven in spatial omics. Here we apply a stemness model for assessing oncogenic states to single cell and spatial omic cancer datasets.

Multiplatform Analysis of Intratumoral PTEN Heterogeneity in Melanoma.

Loss of protein expression of the tumor suppressor PTEN is associated with increased cancer aggressiveness, decreased tumor immune infiltration, and resistance to immune and targeted therapies in melanoma. We assessed a unique cohort of eight melanoma samples with focal loss of PTEN protein expression to understand the features and mechanisms of PTEN loss in this disease. We compared the PTEN-negative (PTEN[-]) areas to their adjacent PTEN-positive (PTEN[+]) areas using DNA sequencing, DNA methylation, RNA expression, digital spatial profiling, and immunohistochemical platforms.

High-plex imaging of RNA and proteins at subcellular resolution in fixed tissue by spatial molecular imaging.

Resolving the spatial distribution of RNA and protein in tissues at subcellular resolution is a challenge in the field of spatial biology. We describe spatial molecular imaging, a system that measures RNAs and proteins in intact biological samples at subcellular resolution by performing multiple cycles of nucleic acid hybridization of fluorescent molecular barcodes. We demonstrate that spatial molecular imaging has high sensitivity (one or two copies per cell) and very low error rate (0.

Multiplex digital spatial profiling of proteins and RNA in fixed tissue.

Digital Spatial Profiling (DSP) is a method for highly multiplex spatial profiling of proteins or RNAs suitable for use on formalin-fixed, paraffin-embedded (FFPE) samples. The approach relies on (1) multiplexed readout of proteins or RNAs using oligonucleotide tags; (2) oligonucleotide tags attached to affinity reagents (antibodies or RNA probes) through a photocleavable (PC) linker; and (3) photocleaving light projected onto the tissue sample to release PC oligonucleotides in any spatial pattern across a region of interest (ROI) covering 1 to ~5,000 cells. DSP is capable of single-cell sensitivity within an ROI using the antibody readout, with RNA detection feasible down to ~600 individual mRNA transcripts.

High-Plex Predictive Marker Discovery for Melanoma Immunotherapy-Treated Patients Using Digital Spatial Profiling.

Purpose: Protein expression in formalin-fixed, paraffin-embedded tissue is routinely measured by IHC or quantitative fluorescence (QIF) on a handful of markers on a single section. Digital spatial profiling (DSP) allows spatially informed simultaneous assessment of multiple biomarkers. Here we demonstrate the DSP technology using a 44-plex antibody cocktail to find protein expression that could potentially be used to predict response to immune therapy in melanoma.

A polyploid admixed origin of beer yeasts derived from European and Asian wine populations.

Strains of Saccharomyces cerevisiae used to make beer, bread, and wine are genetically and phenotypically distinct from wild populations associated with trees. The origins of these domesticated populations are not always clear; human-associated migration and admixture with wild populations have had a strong impact on S. cerevisiae population structure.

High-Throughput Identification of Adaptive Mutations in Experimentally Evolved Yeast Populations.

High-throughput sequencing has enabled genetic screens that can rapidly identify mutations that occur during experimental evolution. The presence of a mutation in an evolved lineage does not, however, constitute proof that the mutation is adaptive, given the well-known and widespread phenomenon of genetic hitchhiking, in which a non-adaptive or even detrimental mutation can co-occur in a genome with a beneficial mutation and the combined genotype is carried to high frequency by selection. We approximated the spectrum of possible beneficial mutations in Saccharomyces cerevisiae using sets of single-gene deletions and amplifications of almost all the genes in the S.

Comprehensive Analysis of the SUL1 Promoter of Saccharomyces cerevisiae.

In the yeast Saccharomyces cerevisiae, beneficial mutations selected during sulfate-limited growth are typically amplifications of the SUL1 gene, which encodes the high-affinity sulfate transporter, resulting in fitness increases of >35% . Cis-regulatory mutations have not been observed at this locus; however, it is not clear whether this absence is due to a low mutation rate such that these mutations do not arise, or they arise but have limited fitness effects relative to those of amplification. To address this question directly, we assayed the fitness effects of nearly all possible point mutations in a 493-base segment of the gene's promoter through mutagenesis and selection.

Aneuploidy shortens replicative lifespan in Saccharomyces cerevisiae.

Aneuploidy and aging are correlated; however, a causal link between these two phenomena has remained elusive. Here, we show that yeast disomic for a single native yeast chromosome generally have a decreased replicative lifespan. In addition, the extent of this lifespan deficit correlates with the size of the extra chromosome.

Origin-Dependent Inverted-Repeat Amplification: Tests of a Model for Inverted DNA Amplification.

DNA replication errors are a major driver of evolution--from single nucleotide polymorphisms to large-scale copy number variations (CNVs). Here we test a specific replication-based model to explain the generation of interstitial, inverted triplications. While no genetic information is lost, the novel inversion junctions and increased copy number of the included sequences create the potential for adaptive phenotypes.

The fitness consequences of aneuploidy are driven by condition-dependent gene effects.

Aneuploidy is a hallmark of tumor cells, and yet the precise relationship between aneuploidy and a cell's proliferative ability, or cellular fitness, has remained elusive. In this study, we have combined a detailed analysis of aneuploid clones isolated from laboratory-evolved populations of Saccharomyces cerevisiae with a systematic, genome-wide screen for the fitness effects of telomeric amplifications to address the relationship between aneuploidy and cellular fitness. We found that aneuploid clones rise to high population frequencies in nutrient-limited evolution experiments and show increased fitness relative to wild type.

The dynamics of diverse segmental amplifications in populations of Saccharomyces cerevisiae adapting to strong selection.

Population adaptation to strong selection can occur through the sequential or parallel accumulation of competing beneficial mutations. The dynamics, diversity, and rate of fixation of beneficial mutations within and between populations are still poorly understood. To study how the mutational landscape varies across populations during adaptation, we performed experimental evolution on seven parallel populations of Saccharomyces cerevisiae continuously cultured in limiting sulfate medium.