Cell Chem Biol. 2024 Nov 16:S2451-9456(24)00452-5. doi: 10.1016/j.chembiol.2024.10.013. Online ahead of print.

ABSTRACT

Engineered probiotics are an emerging platform for in situ delivery of therapeutics to the gut. Herein, we developed an orally administered, yeast-based therapeutic delivery system to deliver next-generation immune checkpoint inhibitor (ICI) proteins directly to gastrointestinal tumors. We engineered Saccharomyces cerevisiae var. boulardii (Sb), a probiotic yeast with high genetic tractability and innate anticancer activity, to secrete "miniature" antibody variants that target programmed death ligand 1 (Sb_haPD-1). When tested in an ICI-refractory colorectal cancer (CRC) mouse model, Sb_haPD-1 significantly reduced intestinal tumor burden and resulted in significant shifts to the immune cell profile and microbiome composition. This oral therapeutic platform is modular and highly customizable, opening new avenues of targeted drug delivery that can be applied to treat a myriad of gastrointestinal malignancies.

PMID:39571582 | DOI:10.1016/j.chembiol.2024.10.013

Plant Physiol Biochem. 2024 Oct 6;218:109180. doi: 10.1016/j.plaphy.2024.109180. Online ahead of print.

ABSTRACT

Soybean (Glycine max) and common bean (Phaseolus vulgaris) diverged approximately 19 million years ago. While these species share a whole-genome duplication (WGD), the Glycine lineage experienced a second, independent WGD. Despite the significance of these WGDs, their impact on gene families related to oil-traits remains poorly understood. Here, we report an in-depth investigation of oil-related gene families in soybean, common bean, and twenty-eight other legume species. We adopted a systematic approach that included 605 RNAseq samples for transcriptome and co-expression analyses, identification of orthologous groups, gene duplication modes and evolutionary rates, and family expansions and contractions. We curated a list of oil candidate genes and found that 91.5% of the families containing these genes expanded in soybean in comparison to common bean. Notably, we observed an expansion of triacylglycerol (TAG) biosynthesis (∼3:1) and an erosion of TAG degradation (∼1.4:1) families in soybean in comparison to common bean. In addition, TAG degradation genes were two-fold more expressed in common bean than in soybean, suggesting that oil degradation is also important for the sharply contrasting seed oil contents in these species. We found 17 transcription factor hub genes that are likely regulators of lipid metabolism. Finally, we inferred expanded and contracted families and correlated these patterns with oil content found in different legume species. In summary, our results do not only shed light on the evolution of oil metabolism genes in soybean, but also present multifactorial evidence supporting the prioritization of promising candidate genes that, if experimentally validated, could accelerate the development of high-oil soybean varieties.

PMID:39571454 | DOI:10.1016/j.plaphy.2024.109180

Blood Adv. 2024 Nov 21:bloodadvances.2024014592. doi: 10.1182/bloodadvances.2024014592. Online ahead of print.

ABSTRACT

Natural killer (NK) cells play an integral role in immunosurveillance against myeloid malignancies, with their mature phenotype and abundance linked to prolonged treatment-free remission in chronic myeloid leukemia (CML). However, NK cell function is suppressed during the disease, and the orchestrators of this impairment are not fully understood. Using a chimeric BCR::ABL1+ CML mouse model, we characterized the impact of the leukemic microenvironment on NK cell function. We showed that NK cells have reduced counts, immature phenotype, poor cytotoxicity, and altered expression of activating and inhibitory receptors in CML mice, which revert to a steady state upon BCR::ABL1 inhibition. Single-cell RNA sequencing revealed an inflammatory cytokine response in CML-exposed NK cells, highlighted by the tumor necrosis factor-a (TNFa)-induced gene signature, upregulation of TNFa receptor TNFR2, and enrichment of SOCS family genes such as Cish, the critical NK cell checkpoint. Ex vivo exposure of healthy NK cells to leukemic soluble factors compromised target-specific NK cell degranulation, which was partially rescued by targeting Cish or TNFa. In alignment with these findings, NK cells from healthy donors displayed suppressed cytotoxicity when exposed to plasma from untreated CML patients, with a partial restoration upon Cish or TNFa inhibition. Furthermore, NK cells from newly diagnosed CML patients pre-destined for blast crisis showed an enrichment of the TNFa-induced pro-inflammatory gene signature identified in CML mice. These results suggest that targeting inflammatory signaling could enhance NK cell-based immunotherapies for CML.

PMID:39571169 | DOI:10.1182/bloodadvances.2024014592

Diabetes Care. 2024 Nov 21:dc241754. doi: 10.2337/dc24-1754. Online ahead of print.

ABSTRACT

OBJECTIVE: Insulin resistance (IR) may be a risk factor for lung disease, but objective evidence is limited. We sought to define the relationship of longitudinal IR with radiographic imaging outcomes and examiner-identified incident lung disease in the Framingham Offspring Study.

RESEARCH DESIGN AND METHODS: Participants without baseline lung disease underwent repeated measurements of fasting insulin and glucose levels over an average period of 13.6 years, from which time-weighted average HOMA-IR was calculated. Each participant then underwent a cardiac gated whole-lung computed tomography scan, which was analyzed for the presence of emphysema, interstitial lung abnormalities (ILAs), and quantitative airway features. Incident lung disease was determined by a study examiner. The relationship of HOMA-IR to these outcomes was estimated in models adjusted for demographics, BMI, and lifetime smoking.

RESULTS: A total of 875 participants with longitudinal IR data and outcomes were identified. Their mean age was 51.5 years, and BMI was 26.7 kg/m2. HOMA-IR was temporally unstable, with a within-person SD approximately two-thirds of the between-person SD. In adjusted models, a 1 SD increase in log(HOMA-IR) z score was associated with higher odds of qualitative emphysema (odds ratio [OR] 1.33; 95% CI 1.04-1.70), ILAs (OR 1.35; 95% CI 1.05-1.74), and modest increases in airway wall thickness and wall area percentage. These radiographic findings were corroborated by a positive association of HOMA-IR with incident lung disease.

CONCLUSIONS: IR is associated with radiographic lung abnormalities and incident lung disease. Deeper phenotyping is necessary to define mechanisms of IR-associated lung injury.

PMID:39571139 | DOI:10.2337/dc24-1754

PLoS One. 2024 Nov 21;19(11):e0311379. doi: 10.1371/journal.pone.0311379. eCollection 2024.

ABSTRACT

Multiple accidents in nuclear power plants and the growing concerns about the misuse of radiation exposure in warfare have called for the rapid determination of absorbed radiation doses (RDs). The latest findings about circulating microRNA (miRNAs) using several animal models revealed considerable promises, although translating this knowledge to clinics remains a major challenge. To address this issue, we randomly divided 36 nonhuman primates (NHPs) into six groups and exposed these groups to six different radiation doses ranging from 6.0-8.5 Gy in increments of 0.5 Gy. Serum samples were collected pre-irradiation as well as three post-irradiation timepoints, namely 1, 2 and 6 days post-total body irradiation (TBI). Generated from a deep sequencing platform, the miRNA reads were multi-variate analyzed to find the differentially expressed putative biomarkers that were linked to RDs, time since irradiation (TSI) and sex. To increase these biomarkers' translational potential, we aligned the NHP-miRNAs' sequences and their functional responses to humans following an in-silico routine. Those miRNAs, which were sequentially and functionally conserved between NHPs and humans, were down selected for further analysis. A linear regression model identified miRNA markers that were consistently regulated with increasing RD but independent TSI. Likewise, a set of potential TSI-markers were identified that consistently shifted with increasing TSI, but independent of RD. Additional molecular analysis found a considerable gender bias in the low-ranges of doses when the risk to radiation-induced fatality was low. Bionetworks linked to cell quantity and cell invasion were significantly altered between the survivors and decedents. Using these biomarkers, an assay could be developed to retrospectively determine the RD and TSI with high translational potential. Ultimately, this knowledge can lead to precise and personalized medicine.

PMID:39570918 | DOI:10.1371/journal.pone.0311379

Plant Cell. 2024 Nov 21:koae310. doi: 10.1093/plcell/koae310. Online ahead of print.

ABSTRACT

Protein activity, abundance, and stability can be regulated by posttranslational modification including ubiquitination. Ubiquitination is conserved among eukaryotes and plays a central role in modulating cellular function, yet we lack comprehensive catalogs of proteins that are modified by ubiquitin in plants. In this study, we describe an antibody-based approach to enrich ubiquitinated peptides coupled with isobaric labeling to enable quantification of up to 18-multiplexed samples. This approach identified 17,940 ubiquitinated lysine sites arising from 6,453 proteins from Arabidopsis (Arabidopsis thaliana) primary roots, seedlings, and rosette leaves. Gene ontology analysis indicated that ubiquitinated proteins are associated with numerous biological processes including hormone signaling, plant defense, protein homeostasis, and metabolism. We determined ubiquitinated lysine residues that directly regulate the stability of three transcription factors, CRYPTOCHROME-INTERACTING BASIC-HELIX-LOOP-HELIX 1 (CIB1), CIB1 LIKE PROTEIN 2 (CIL2), and SENSITIVE TO PROTON RHIZOTOXICITY1 (STOP1) using in vivo degradation assays. Furthermore, codon mutation of CIB1 to create a K166R conversion to prevent ubiquitination, via CRISPR/Cas9-derived adenosine base editing, led to an early flowering phenotype and increased expression of FLOWERING LOCUS T (FT). These comprehensive site-level ubiquitinome profiles provide a wealth of data for future functional studies related to modulation of biological processes mediated by this posttranslational modification in plants.

PMID:39570863 | DOI:10.1093/plcell/koae310

Plant Physiol. 2024 Nov 21:kiae621. doi: 10.1093/plphys/kiae621. Online ahead of print.

NO ABSTRACT

PMID:39570744 | DOI:10.1093/plphys/kiae621

Proc Natl Acad Sci U S A. 2024 Nov 26;121(48):e2413743121. doi: 10.1073/pnas.2413743121. Epub 2024 Nov 21.

ABSTRACT

Delineating a protein's essential and dispensable domains provides critical insight into how it carries out its function. Here, we developed a high-throughput method to synthesize and test the functionality of all possible in-frame and continuous deletions in a gene of interest, enabling rapid and unbiased determination of protein domain importance. Our approach generates precise deletions using a CRISPR library framework that is free from constraints of gRNA target site availability and efficacy. We applied our method to AcrIIA4, a phage-encoded anti-CRISPR protein that robustly inhibits SpCas9. Extensive structural characterization has shown that AcrIIA4 physically occupies the DNA-binding interfaces of several SpCas9 domains; nonetheless, the importance of each AcrIIA4 interaction for SpCas9 inhibition is unknown. We used our approach to determine the essential and dispensable regions of AcrIIA4. Surprisingly, not all contacts with SpCas9 were required, and in particular, we found that the AcrIIA4 loop that inserts into SpCas9's RuvC catalytic domain can be deleted. Our results show that AcrIIA4 inhibits SpCas9 primarily by blocking PAM binding and that its interaction with the SpCas9 catalytic domain is inessential.

PMID:39570312 | DOI:10.1073/pnas.2413743121

Curr Protoc. 2024 Nov;4(11):e70059. doi: 10.1002/cpz1.70059.

ABSTRACT

The yeast one-hybrid system (Y1H) is used extensively to identify DNA-protein interactions. The generation of large collections of open reading frames (ORFs) to be used as prey in screenings is not a bottleneck nowadays and can be carried out in-house or offered as a service by companies. However, the straightforward use of full gene promoters as baits to identify interacting proteins undermines the accuracy and sensitivity of the assay, especially in the case of multicellular eukaryotes. Therefore, it is paramount to implement procedures for efficient identification of suitable promoter fragments compatible with the Y1H assay. Here, we describe a workflow to identify biologically relevant conserved promoter fragments of Arabidopsis thaliana through simple and robust phylogenetic analyses. Additionally, we describe a manual method and its automated robotized version for rapid and efficient high-throughput Y1H screenings of arrayed ORF libraries with the identified DNA fragments. Moreover, this method can be scaled up or down and used for yeast two-hybrid screenings to search for possible interactors of proteins identified by the Y1H approach or any other protein of interest, altogether underscoring its suitability to build gene regulatory networks. © 2024 The Author(s). Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Selection of DNA baits for Y1H screenings Basic Protocol 2: Y1H screenings with arrayed gene libraries Alternate Protocol: Automated screening with a liquid-handling robot.

PMID:39570200 | DOI:10.1002/cpz1.70059

R Soc Open Sci. 2024 Nov 20;11(11):240094. doi: 10.1098/rsos.240094. eCollection 2024 Nov.

ABSTRACT

In this study, we investigate the impact of demographic characteristics on Middle East respiratory syndrome coronavirus (MERS-CoV) cases in Saudi Arabia, specifically focusing on the time intervals between symptom onset and key events such as hospitalization, case confirmation, reporting and death. We estimate these intervals using data from 2196 cases occurring between June 2012 and January 2020, partitioning the data into four age groups (0-24 years, 25-49 years, 50-74 years and 75-100 years). The duration from symptom onset to hospitalization varies between age cohorts, ranging from 4.03 to 4.75 days, with the 75-100 age group experiencing the longest delay. The interval from symptom onset to case confirmation spans 5.83-8.24 days, and again, the 75-100 age group faces the lengthiest delay. The interval from symptom onset and case reporting ranges from 7.0 to 9.8 days, with the 75-100 age group experiencing the longest delay. The period from symptom onset to death varies across age groups (12.3-16.1 days), with elevated mortality rates during outbreaks. Importantly, we observe age-based differences in the risk of hospitalization and other measures of infection severity, including the probability of death conditional on hospitalization. Careful quantification of epidemiological characteristics, including inference of key epidemiological periods and assessments of differences between cases of different ages, plays a crucial role in understanding the progression of MERS-CoV outbreaks and formulating effective public health strategies to mitigate their impact.

PMID:39569349 | PMC:PMC11576103 | DOI:10.1098/rsos.240094

Bioinform Adv. 2024 Nov 14;4(1):vbae175. doi: 10.1093/bioadv/vbae175. eCollection 2024.

ABSTRACT

MOTIVATION: Analysis of gene and isoform expression levels is becoming critical for the detailed understanding of biochemical mechanisms. In addition, integrating RNA-seq data with other omics data types, such as proteomics and metabolomics, provides a strong approach for consolidating our understanding of biological processes across various organizational tiers, thus promoting the identification of potential therapeutic targets.

RESULTS: We present our pipeline, called MultiOmicsIntegrator (MOI), an inclusive pipeline for comprehensive omics analyses. MOI represents a unified approach that performs in-depth individual analyses of diverse omics. Specifically, exhaustive analysis of RNA-seq data at the level of genes, isoforms of genes, as well as miRNA is offered, coupled with functional annotation and structure prediction of these transcripts. Additionally, proteomics and metabolomics data are supported providing a holistic view of biological systems. Finally, MOI has tools to integrate simultaneously multiple and diverse omics datasets, with both data- and function-driven approaches, fostering a deeper understanding of intricate biological interactions.

AVAILABILITY AND IMPLEMENTATION: MOI and ReadTheDocs.

PMID:39569320 | PMC:PMC11576358 | DOI:10.1093/bioadv/vbae175

Chem Sci. 2024 Nov 12. doi: 10.1039/d4sc05459h. Online ahead of print.

ABSTRACT

Chemical probes have gained importance in the elucidation of signal transduction in biology. Insufficient selectivity and potency, lack of cellular activity and inappropriate use of chemical probes has major consequences on interpretation of biological results. The catalytic subunit of phosphoinositide 3-kinase α (PI3Kα) is one of the most frequently mutated genes in cancer, but fast-acting, high-quality probes to define PI3Kα's specific function to clearly separate it from other class I PI3K isoforms, are not available. Here, we present a series of novel covalent PI3Kα-targeting probes with optimized intracellular target access and kinetic parameters. On-target TR-FRET and off-target assays provided relevant kinetic parameters (k chem, k inact and K i) to validate our chemical probes. Additional intracellular nanoBRET tracer displacement measurements showed rapid diffusion across the cell membrane and extremely fast target engagement, while investigations of signaling downstream of PI3Kα via protein kinase B (PKB/Akt) and forkhead box O (FOXO) revealed blunted pathway activity in cancer cell lines with constitutively activated PI3Kα lasting for several days. In contrast, persistent PI3Kα inhibition was rapidly bypassed by other class I PI3K isoforms in cells lacking functional phosphatase and tensin homolog (PTEN). Comparing the rapidly-diffusing, fast target-engaging chemical probe 9 to clinical reversible PI3Kα-selective inhibitors alpelisib, inavolisib and 9r, a reversible analogue of 9, revealed 9's superior potency to inhibit growth (up to 600-fold) associated with sustained suppression of PI3Kα signaling in breast cancer cell lines. Finally, using a simple washout protocol, the utility of the highly-selective covalent PI3Kα probe 9 was demonstrated by the quantification of the coupling of insulin, EGF and CXCL12 receptors to distinct PI3K isoforms for signal transduction in response to ligand-dependent activation. Collectively, these findings along with the novel covalent chemical probes against PI3Kα provide insights into isoform-specific functions in cancer cells and highlight opportunities to achieve improved selectivity and long-lasting efficacy.

PMID:39568927 | PMC:PMC11575505 | DOI:10.1039/d4sc05459h

Front Genet. 2024 Nov 6;15:1451679. doi: 10.3389/fgene.2024.1451679. eCollection 2024.

ABSTRACT

INTRODUCTION: Polygenic Risk Scores (PRS) are an emerging tool for predicting an individual's genetic risk to a complex trait. Several methods have been proposed to construct and calculate these scores. Here, we develop a biologically driven PRS using the UK BioBank cohort through validated protein interactions (PPI) and network construction for psoriasis, incorporating variants mapped to the interacting genes of 14 psoriasis susceptibility (PSORS) loci, as identified from previous genetic linkage studies.

METHODS: We constructed the PPI network via the implementation of two major meta-databases of protein interactions, and identified variants mapped to the identified PSORS-interacting genes. We selected only European unrelated participants including individuals with psoriasis and randomly selected healthy controls using an at least 1:4 ratio to maximize statistical power. We next compared our PPI-PRS model to (i) clinical risk models and (ii) conventional PRS calculations through p-value thresholding.

RESULTS: Our PPI-PRS model provides comparable results to both clinical risk models and conventional approaches, despite the incorporation of a limited number of variants which have not necessarily reached genome-wide significance (GWS). Exclusion of variants mapped to the HLA-C locus, an established risk locus for psoriasis resulted in highly similar associations compared to our primary model, indicating the contribution of the genetic variability mapped to non-GWS variants in PRS computations.

DISCUSSION: Our findings support the implementation of biologically driven approaches in PRS calculations in psoriasis, highlighting their potential clinical utility in risk assessment and treatment management.

PMID:39568675 | PMC:PMC11576467 | DOI:10.3389/fgene.2024.1451679

Patterns (N Y). 2024 Oct 31;5(11):101093. doi: 10.1016/j.patter.2024.101093. eCollection 2024 Nov 8.

ABSTRACT

Computational analyses of transcriptomic data have dramatically improved our understanding of complex diseases. However, such approaches are limited by small sample sets of disease-affected material. We asked if a variational autoencoder trained on large groups of healthy human RNA sequencing (RNA-seq) data can capture the fundamental gene regulation system and generalize to unseen disease changes. Importantly, we found this model to successfully compress unseen transcriptomic changes from 25 independent disease datasets. We decoded disease-specific signals from the latent space and found them to contain more disease-specific genes than the corresponding differential expression analysis in 20 of 25 cases. Finally, we matched these disease signals with known drug targets and extracted sets of known and potential pharmaceutical candidates. In summary, our study demonstrates how data-driven representation learning enables the arithmetic deconstruction of the latent space, facilitating the dissection of disease mechanisms and drug targets.

PMID:39568475 | PMC:PMC11573900 | DOI:10.1016/j.patter.2024.101093

BMC Genomics. 2024 Nov 20;25(1):1119. doi: 10.1186/s12864-024-11025-3.

ABSTRACT

BACKGROUND: Over their evolutionary history, corals have adapted to sea level rise and increasing ocean temperatures, however, it is unclear how quickly they may respond to rapid change. Genome structure and genetic diversity contained within may highlight their adaptive potential.

RESULTS: We present chromosome-scale genome assemblies and linkage maps of the critically endangered Atlantic acroporids, Acropora palmata and A. cervicornis. Both assemblies and linkage maps were resolved into 14 chromosomes with their gene content and colinearity. Repeats and chromosome arrangements were largely preserved between the species. The family Acroporidae and the genus Acropora exhibited many phylogenetically significant gene family expansions. Macrosynteny decreased with phylogenetic distance. Nevertheless, scleractinians shared six of the 21 cnidarian ancestral linkage groups as well as numerous fission and fusion events compared to other distantly related cnidarians. Genetic linkage maps were constructed from one A. palmata family and 16 A. cervicornis families using a genotyping array. The consensus maps span 1,013.42 cM and 927.36 cM for A. palmata and A. cervicornis, respectively. Both species exhibited high genome-wide recombination rates (3.04 to 3.53 cM/Mb) and pronounced sex-based differences, known as heterochiasmy, with 2 to 2.5X higher recombination rates estimated in the female maps.

CONCLUSIONS: Together, the chromosome-scale assemblies and genetic maps we present here are the first detailed look at the genomic landscapes of the critically endangered Atlantic acroporids. These data sets revealed that adaptive capacity of Atlantic acroporids is not limited by their recombination rates. The sister species maintain macrosynteny with few genes with high sequence divergence that may act as reproductive barriers between them. In the Atlantic Acropora, hybridization between the two sister species yields an F1 hybrid with limited fertility despite the high levels of macrosynteny and gene colinearity of their genomes. Together, these resources now enable genome-wide association studies and discovery of quantitative trait loci, two tools that can aid in the conservation of these species.

PMID:39567907 | DOI:10.1186/s12864-024-11025-3

Nature. 2024 Nov;635(8039):699-707. doi: 10.1038/s41586-024-07571-1. Epub 2024 Nov 20.

ABSTRACT

The gastrointestinal tract is a multi-organ system crucial for efficient nutrient uptake and barrier immunity. Advances in genomics and a surge in gastrointestinal diseases1,2 has fuelled efforts to catalogue cells constituting gastrointestinal tissues in health and disease3. Here we present systematic integration of 25 single-cell RNA sequencing datasets spanning the entire healthy gastrointestinal tract in development and in adulthood. We uniformly processed 385 samples from 189 healthy controls using a newly developed automated quality control approach (scAutoQC), leading to a healthy reference atlas with approximately 1.1 million cells and 136 fine-grained cell states. We anchor 12 gastrointestinal disease datasets spanning gastrointestinal cancers, coeliac disease, ulcerative colitis and Crohn's disease to this reference. Utilizing this 1.6 million cell resource (gutcellatlas.org), we discover epithelial cell metaplasia originating from stem cells in intestinal inflammatory diseases with transcriptional similarity to cells found in pyloric and Brunner's glands. Although previously linked to mucosal healing4, we now implicate pyloric gland metaplastic cells in inflammation through recruitment of immune cells including T cells and neutrophils. Overall, we describe inflammation-induced changes in stem cells that alter mucosal tissue architecture and promote further inflammation, a concept applicable to other tissues and diseases.

PMID:39567783 | DOI:10.1038/s41586-024-07571-1

Nature. 2024 Nov 20. doi: 10.1038/s41586-024-08240-z. Online ahead of print.

ABSTRACT

Predicting elemental cycles and maintaining water quality under increasing anthropogenic influence requires knowledge of the spatial drivers of river microbiomes. However, understanding of the core microbial processes governing river biogeochemistry is hindered by a lack of genome-resolved functional insights and sampling across multiple rivers. Here we used a community science effort to accelerate the sampling, sequencing and genome-resolved analyses of river microbiomes to create the Genome Resolved Open Watersheds database (GROWdb). GROWdb profiles the identity, distribution, function and expression of microbial genomes across river surface waters covering 90% of United States watersheds. Specifically, GROWdb encompasses microbial lineages from 27 phyla, including novel members from 10 families and 128 genera, and defines the core river microbiome at the genome level. GROWdb analyses coupled to extensive geospatial information reveals local and regional drivers of microbial community structuring, while also presenting foundational hypotheses about ecosystem function. Building on the previously conceived River Continuum Concept1, we layer on microbial functional trait expression, which suggests that the structure and function of river microbiomes is predictable. We make GROWdb available through various collaborative cyberinfrastructures2,3, so that it can be widely accessed across disciplines for watershed predictive modelling and microbiome-based management practices.

PMID:39567690 | DOI:10.1038/s41586-024-08240-z

Nature. 2024 Nov 20. doi: 10.1038/s41586-024-08129-x. Online ahead of print.

ABSTRACT

Aneuploid epithelial cells are common in breast cancer1,2; however, their presence in normal breast tissues is not well understood. To address this question, we applied single-cell DNA sequencing to profile copy number alterations in 83,206 epithelial cells from the breast tissues of 49 healthy women, and we applied single-cell DNA and assay for transposase-accessible chromatin sequencing co-assays to the samples of 19 women. Our data show that all women harboured rare aneuploid epithelial cells (median 3.19%) that increased with age. Many aneuploid epithelial cells (median 82.22%) in normal breast tissues underwent clonal expansions and harboured copy number alterations reminiscent of invasive breast cancers (gains of 1q; losses of 10q, 16q and 22q). Co-assay profiling showed that the aneuploid cells were mainly associated with the two luminal epithelial lineages, and spatial mapping showed that they localized in ductal and lobular structures with normal histopathology. Collectively, these data show that even healthy women have clonal expansions of rare aneuploid epithelial cells in their breast tissues.

PMID:39567687 | DOI:10.1038/s41586-024-08129-x

Nat Microbiol. 2024 Nov 20. doi: 10.1038/s41564-024-01869-6. Online ahead of print.

NO ABSTRACT

PMID:39567660 | DOI:10.1038/s41564-024-01869-6

Genome Res. 2024 Nov 20;34(11):1735-1746. doi: 10.1101/gr.279640.124.

ABSTRACT

RNA isoform diversity, produced via alternative splicing, and alternative usage of transcription start and poly(A) sites, results in varied transcripts being derived from the same gene. Distinct isoforms can play important biological roles, including by changing the sequences or expression levels of protein products. The first single-cell approaches to RNA sequencing-and later, spatial approaches-which are now widely used for the identification of differentially expressed genes, rely on short reads and offer the ability to transcriptomically compare different cell types but are limited in their ability to measure differential isoform expression. More recently, long-read sequencing methods have been combined with single-cell and spatial technologies in order to characterize isoform expression. In this review, we provide an overview of the emergence of single-cell and spatial long-read sequencing and discuss the challenges associated with the implementation of these technologies and interpretation of these data. We discuss the opportunities they offer for understanding the relationships between the distinct variable elements of transcript molecules and highlight some of the ways in which they have been used to characterize isoforms' roles in development and pathology. Single-nucleus long-read sequencing, a special case of the single-cell approach, is also discussed. We attempt to cover both the limitations of these technologies and their significant potential for expanding our still-limited understanding of the biological roles of RNA isoforms.

PMID:39567235 | DOI:10.1101/gr.279640.124