Biology subjects

Ragoussis, I.

Publications and source records attributed to Ragoussis, I..

15 recordsLinked to original sources

bioRxiv2026

LINC00536 regulates transcriptional repressor TRPS1 in breast cancer

Metastatic breast cancer with complex molecular mechanisms of progression accounts for most cancer related deaths in women. To improve diagnosis and drug development, it is important to identify novel biomarkers and critical molecular pathways involved in tumor initiation and progression. Here, we profiled and analyzed the expression of long non-coding RNAs (lncRNAs) from three distinct stages of tumor initiation and progression (hyperplasia, adenoma, and carcinoma). We performed RNAseq on tumor and mammary epithelial cells derived from ROSAmT/mG tumor and non-tumor mice. We identified 1913 differentially expressed protein coding genes and 324 lncRNAs in breast cancer cells of all stages compared with normal mammary epithelial cells. Pearson correlation analysis correlated 93 differentially expressed lncRNAs with protein coding genes, providing a comprehensive lncRNA-protein coding genes co-expression network. Among them, we focused on Gm19303 which was paired with the differentially expressed protein coding gene, transcriptional repressor GATA binding 1 (Trps1), and identified its human counterpart as LINC00536. Both LINC00536 and TRPS1 are only overexpressed in breast cancer and correlate with poor prognosis of patient from the TCGA and GTEx databases. Single cell RNAseq data from the Atlas of Human breast cancers further confirmed that TRPS1 is upregulated in human breast cancer compared to normal human mammary tissue with highest expression in ER+ subgroup. In summary, our study explored the potential role of lncRNAs in breast cancer initiation and progression. *Implications statement: Our findings imply that human LINC00536/TRPS1 serves as a novel and early biomarker of cancer progression and a potential therapeutic target for breast cancer.

cancer biology↗

bioRxiv2025

Long-read RNA-seq delineates temporal transcriptional dynamics in multiplexed and sexed single medfly embryos

Long-read RNA sequencing has great potential to improve genomic characterization of non-model organisms due to its ability to yield full-length genes. Coupled with absolute gene expression quantification, dynamics of development orchestrated at transcript level can be elucidated with high precision. The resolution of this precision can be further improved by studying organisms as close as possible to their basic entities, single cells for example or single embryos. Here, we collected developing embryos of the Mediterranean fruit fly (medfly, Ceratitis capitata) at hourly time-points for the first 15 hours of development. The medfly is an organism of huge economic importance in agriculture due to its wide host range including apples, pear, citrus, olives, etc. We simultaneously isolated total RNA and genomic DNA from single embryos and sexed the embryos using Y-specific PCR assays. The RNA, spiked with external ERCC standards to aid in absolute quantification, was used to perform Nanopore long-read RNA-seq. We developed a genome-guided transcriptome assembly based on full-length transcripts and identified a total of 22,875 transcripts comprising 3879 novel genes, missed in the current NCBI predicted gene models. We show that, indeed, the absolute quantification of gene expression performs superiorly to relative quantification in highly dynamic systems such as developing embryos. Further, we used unsupervised clustering and lineage tracing algorithms to group and accurately place embryos along a pseudo-temporal development trajectory. We show that medfly embryos undergo successive waves of zygotic genome activation. We discover a dramatic reorganization of maternally deposited mRNA occurring within the first 3 hours of egg laying followed by maternal-to-zygotic transition. We finally identify modules of temporal synexpression and elucidate the biological role of these modules. Together, these results provide the first detailed look at early embryo development in the medfly and should aid in future control efforts of this pest.

genomics↗

bioRxiv2025

Genome assembly of five Tephritid species for the enhancement of the Sterile Insect Technique

Tephritidae insect pests account for extensive crop damage and yield losses globally. Modern, sustainable pest management approaches are species-specific and, therefore, high-quality genome assemblies are required for their application. Here, we present chromosome-level assemblies for five members of the Tephritidae family: Anastrepha fraterculus, Anastrepha ludens, Bactrocera dorsalis, Bactrocera zonata and Zeugodacus cucurbitae. The assemblies used long read sequencing polished with short read sequencing and scaffolded using Hi-C (chromatin conformation capture) sequencing. Prior to scaffolding the assembly deduplication was performed to separate a primary assembly and an alternate assembly, and each was then scaffolded independently. The scaffolded assemblies reached N50 length in the range of 60Mb to 120Mb. The scaffolded assemblies were verified with BUSCO and completeness was in the range 97% to 98.5% and had very low duplicated, fragmented and missing orthologs.

ecology↗

bioRxiv2025

A fibroblast-centric network drives cold fibrosis in the tumor microenvironment of lung squamous cell carcinoma

The tumor microenvironment (TME) of chronic inflammation-associated cancers (CIACs) is shaped by cycles of injury and maladaptive repair, yet the principles organizing fibrotic stroma in these tumors remain unclear. Here, we applied the concept of hot versus cold fibrosis, originally credentialed in non-cancerous fibrosis of heart and kidney, to lung squamous cell carcinoma (LUSC), a prototypical CIAC. Single-cell transcriptomics of matched tumor and adjacent-normal tissue from 16 treatment-naive LUSC patients identified a cold fibrotic architecture in the LUSC TME: cancer-associated fibroblasts (CAFs) expanded and adopted myofibroblast and stress-response states, while macrophages were depleted. This macrophage-poor, CAF-rich stroma was maintained by CAF autocrine growth factor loops, including TIMP1, INHBA, TGFB1, and GMFB. In parallel, the immune compartment exhibited a hot tumor phenotype with abundant T and B cells, forming spatially distinct but molecularly engaged networks with CAFs. CAF gene programs typifying cold fibrosis in LUSC were conserved in other CIACs, including esophageal and gastric adenocarcinomas. These results redefine desmoplastic regions of tumors through the lens of a non-cancer fibrosis model, demonstrating that conserved stromal circuits constitute therapeutic vulnerabilities in CIACs.

cancer biology↗

bioRxiv2025

A Fibroblast State Choreographs an Epithelial YAP-dependent Regenerative Program Essential to (Pre)malignancy via ECM-mediated Mechanotransduction

Chronic lung injury generates metaplasia which occasionally, but ominously, progresses to squamous dysplasia and squamous lung cancer. To identify mechanisms through which disrupted tissue homeostasis contributes to malignant initiation and progression, we used in vivo and in vitro heterotypic recombinant models of human bronchial epithelial cells (hBECs) and fibroblasts. We demonstrate that injury-associated TGF-{beta} signaling creates a fibroblast state dependent upon HSP47 upregulation. These fibroblasts accumulated collagen, thus elevating tissue stiffness and activating mechanosignaling that sustained YAP-dependent embryonic-like, pro-malignant activities in adjacent hBECs. This Stress/Tension-Instructive Fibroblast (STIF) state, exhibited by stressed fibroblasts in premalignant and malignant lesions across multiple cancer types, was sufficient to reprogram disease-free hBECs to metaplasia and to drive hBECs with compromised tumor suppressor function to dysplasia, yet could be inhibited and reversed. STIFs suffice to activate epithelial phenotypes reminiscent of oncogene-mediated cell transformation and induce (pre)malignancy via increased force transmission, providing novel targets for prevention. Statement of significanceTissue injury creates a regenerative pro-tumorigenic Stress/Tension-Instructive Fibroblast (STIF) state which is sufficient to activate a YAP-dependent, pre-malignant program to induce or unmask pre-cancerous phenotypes in epithelial cells through mechanotransduction. Inhibition of STIF activity or mechanosignaling prevents metaplasia and progression to dysplasia. HighlightsO_LITissue injury creates a pro-tumorigenic Stress/Tension-Instructive Fibroblast (STIF) state in multiple organs that precedes and persists through cancer C_LIO_LISTIF signaling alone, working through fibroblasts and not epithelial cells, is sufficient to activate embryonic-like plasticity and induce epithelial pre-cancerous metaplastic lesions C_LIO_LISTIFs program (pre)malignant phenotypes in adjacent epithelial cells through mechanosignaling by activating YAP prior to tumor formation C_LIO_LIInhibiting STIFs or mechanosignaling prevents/reverts metaplasia and prevents progression to dysplasia C_LI

cancer biology↗

bioRxiv2025

The genome sequence of the hemlock woolly adelgid, Adelges tsugae Annand 1924

The genome of the hemlock woolly adelgid (Adelges tsugae) is presented in 10 chromosomal pseudomolecules along with 132 unplaced scaffolds for a combined length of 216.56 Mb. The genome is highly contiguous, with a benchmarking universal single-copy orthologs (BUSCO) completion score of 98.25%, an N50 of 20.54 Mb, and the longest scaffold 37.41 Mb in length. On the basis of chromosomal synteny with other aphid genomes, the hemlock woolly adelgids genome is composed of 8 autosomes and two sex (X) chromosomes. Annotation of the assembly identified 11,800 coding genes, 1,930 non-coding genes, and 20,403 mRNA transcripts. The mitogenome is also presented in a single, annotated, circular contig 25,980 bases in length. Species taxonomyEukaryota; Metazoa; Arthropoda; Hexapoda; Insecta; Pterygota; Neoptera; Paraneoptera; Hemiptera; Sternorrhyncha; Aphidomorpha; Adelgoidea; Adelgidae; Adelges; Annandina; Adelges (Annandina) tsugae Annand, 1924 (NCBI:txid357502)

genomics↗

bioRxiv2023

Concerted epithelial and stromal changes during progression of Barrett's Esophagus to invasive adenocarcinoma exposed by multi-scale, multi-omics analysis

Esophageal adenocarcinoma arises from Barretts esophagus, a precancerous metaplastic replacement of squamous by columnar epithelium in response to chronic inflammation. Multi-omics profiling, integrating single-cell transcriptomics, extracellular matrix proteomics, tissue-mechanics and spatial proteomics of 64 samples from 12 patients paths of progression from squamous epithelium through metaplasia, dysplasia to adenocarcinoma, revealed shared and patient-specific progression characteristics. The classic metaplastic replacement of epithelial cells was paralleled by metaplastic changes in stromal cells, ECM and tissue stiffness. Strikingly, this change in tissue state at metaplasia was already accompanied by appearance of fibroblasts with characteristics of carcinoma-associated fibroblasts and of an NK cell-associated immunosuppressive microenvironment. Thus, Barretts esophagus progresses as a coordinated multi-component system, supporting treatment paradigms that go beyond targeting cancerous cells to incorporating stromal reprogramming. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=187 SRC="FIGDIR/small/544265v1_ufig1.gif" ALT="Figure 1"> View larger version (62K): org.highwire.dtl.DTLVardef@64c375org.highwire.dtl.DTLVardef@15c6ef2org.highwire.dtl.DTLVardef@177dee0org.highwire.dtl.DTLVardef@c945e7_HPS_FORMAT_FIGEXP M_FIG Graphical AbstractTo obtain a comprehensive picture of the coordinated changes in epithelial, stromal and immune compartments during development of Barretts-associated esophageal adenocarcinoma, patient-matched samples corresponding to various phases of disease progression were collected from 12 patients, each of which had at a given time point lesions at multiple stages progression (matched-normal, metaplasia, dysplasia, and carcinoma). Matched "normal" gastric tissues were also collected. These sample were analyzed by single cell RNA-sequencing (scRNAseq) for single-cell resolution transcriptomics and Copy Number Variant (CNV), by proteomics for extracellular matrix (ECM) proteins, by Atomic Force Microscopy (AFM for tissue stiffness and by CODEX spatial proteomics imaging The integrative multi-omics analysis exposed drastic alterations in cell type composition and shifts in cell states in all three compartments. A large subpopulation of fibroblasts absent in the normal esophagus and characteristic of dysplasia and adenocarcinoma sample, that based on markers would indeed be considered cancer associated fibroblasts (CAF), appeared already in the metaplastic phase. This fibroblast subpopulation had transcriptomes virtually indistinguishable with fibroblasts of the cancer free gastric epithelium in these patients C_FIG

cancer biology↗

bioRxiv2023

Natural evolution of SARS-CoV-2 variants in K18-ACE2 mice gives rise to more virulent virus and variant alleles associated with treatment resistance.

The persistence of COVID-19 is partly due to viral evolution reducing vaccine and treatment efficacy. Serial infections of Wuhan-like SARS-CoV-2 in Balb/c mice yielded mouse-adapted strains with greater infectivity and mortality. We investigated if passaging unmodified B.1.351 (Beta) and B.1.617.2 (Delta) 20 times in K18-ACE2 mice, expressing human ACE2 receptor, in a BSL-3 laboratory without selective pressures, would drive human health-relevant evolution and if evolution was lineage-dependent. Late-passage virus caused more severe disease, at organism and lung tissue scales, with late-passage Delta demonstrating antibody resistance and interferon suppression. This resistance co-occurred with a de novo spike S371F mutation, linked with both traits. S371F, an Omicron-characteristic mutation, was co-inherited at times with spike E1182G per Nanopore sequencing, existing in different quasi-species at others. Both are linked to mammalian GOLGA7 and ZDHHC5 interactions, which mediate viral-cell entry and antiviral response. This study demonstrates SARS-CoV-2s tendency to evolve with phenotypic consequences, its evolution varying by lineage, and suggests non-dominant quasi-species contribute.

genetics↗

bioRxiv2023

Inhibition of Type I PRMTs Reforms Muscle Stem Cell Identity Enhancing their Therapeutic Capacity

In skeletal muscle, muscle stem cells (MuSC) are the main cells responsible for regeneration upon injury. In diseased skeletal muscle, it would be therapeutically advantageous to replace defective MuSCs, or rejuvenate them with drugs to enhance their self-renewal and ensure long-term regenerative potential. One limitation of the replacement approach has been the inability to efficiently expand MuSCs ex vivo, while maintaining their stemness and engraftment abilities. Herein, we show that inhibition of type I protein arginine methyltransferases (PRMTs) with MS023 increases the proliferative capacity of ex vivo cultured MuSCs. Single cell RNA sequencing (scRNAseq) of ex vivo cultured MuSCs revealed the emergence of subpopulations in MS023-treated cells which are defined by elevated Pax7 expression and markers of MuSC quiescence, both features of enhanced self-renewal. Furthermore, the scRNAseq identified MS023-specific subpopulations to be metabolically altered with upregulated glycolysis and oxidative phosphorylation (OxPhos). Transplantation of MuSCs treated with MS023 had a better ability to repopulate the MuSC niche and contributed efficiently to muscle regeneration following injury. Interestingly, the preclinical mouse model of Duchenne muscular dystrophy had increased grip strength with MS023 treatment. Our findings show that inhibition of type I PRMTs increased the proliferation capabilities of MuSCs with altered cellular metabolism, while maintaining their stem-like properties such as self-renewal and engraftment potential.

molecular biology↗

bioRxiv2021

A compendium of novel genomics technologies provides a chromosome-scale assembly and insights into the sex determining system of the Greenland Halibut

Despite the commercial importance of Greenland Halibut (Reinhardtius hippoglossoides), important gaps still persist in our knowledge of this species, including its reproductive biology and sex determination mechanism. In this study, we combined single molecule sequencing of long reads (Pacific Sciences) with Chromatin Conformation Capture sequencing (Hi-C) data to provide the first chromosome-level genome reference for this species. The high-quality assembly encompassed more than 598 Megabases (Mb) assigned to 1 594 scaffolds (scaffold N50 = 25 Mb) with 96 % of its total length distributed among 24 chromosomes. The investigation of its syntenic relationships with other economically important flatfish species revealed a high conservation of synteny blocks among members of this phylogenetic clade. Sex determination analysis revealed that flatfishes do not escape the rule applied to other teleost fish and exhibit a high level of plasticity and turnover in sex-determination mechanisms. A whole-genome sequence analysis of 198 individuals allowed us to draw a full picture of the molecular sex determination (SD) system for Greenland Halibut, revealing that this species possesses a very nascent male heterogametic XY system, with a putative major effect of the sox2 gene, also described as the main SD driver in two other flatfishes. Interestingly, our study also suggested for the first time in flatfishes that a putative Y-autosomal fusion could be associated with a reduction of recombination typical of early steps of sex chromosome evolution.

genomics↗

bioRxiv2020

Single-cell RNA sequencing reveals time- and sex-specific responses of spinal cord microglia to peripheral nerve injury and links ApoE to neuropathic pain

Activation of microglia in the spinal cord following peripheral nerve injury is critical for the development of long-lasting pain hypersensitivity. However, it remains unknown whether distinct microglia subpopulations or states contribute to different stages of pain development and maintenance. We show, using single-cell RNA-sequencing, that nerve injury induces the generation of a male-specific inflammatory microglia subtype, and demonstrate increased proliferation of microglia in males as compared to females. We also show time- and sex-specific transcriptional changes in different microglial subpopulations following injury. Apolipoprotein E (Apoe) is the top upregulated gene in microglia at chronic time points after nerve injury in mice and polymorphisms in the APOE gene in humans are associated with chronic pain. Single-cell analysis of human spinal cord microglia reveals a subpopulation with a disease-related transcriptional signature. Our data provide a detailed analysis of transcriptional states of mouse and human spinal cord microglia, and identify a previously unrecognized role for ApoE in neuropathic pain.

neuroscience↗

bioRxiv2020

Oviduct epithelial cells constitute two developmentally distinct lineages that are spatially separated along the distal-proximal axis

Owing to technical advances in single cell biology, the appreciation of cellular heterogeneity has increased, which has aided our understanding of organ function, homeostasis and disease progression. The oviduct (also known as the fallopian tube in humans) is the distal-most portion of the female reproductive tract. It is essential for reproduction and the proposed origin of high grade serous ovarian carcinoma (HGSOC). In mammals, the oviduct is morphologically segmented along the ovary-uterus axis into four evolutionally conserved regions. It is unknown however if there is a diversification of epithelial cell characteristics between these regions. In this study, we identified transcriptionally distinct populations of secretory and multiciliated cells restricted to the distal and proximal regions of the oviduct. We demonstrated that these distal and proximal populations are distinct lineages specified early in Mullerian duct development and are maintained separately. These results aid our understanding of epithelial development, homeostasis and initiation of disease from the oviduct.

developmental biology↗

bioRxiv2020

White pupae genes in the Tephritids Ceratitis capitata, Bactrocera dorsalis and Zeugodacus cucurbitae: a story of parallel mutations

Mass releases of sterilized male insects, in the frame of sterile insect technique programs, have helped suppress insect pest populations since the 1950s. In the major horticultural pests Bactrocera dorsalis, Ceratitis capitata, and Zeugodacus cucurbitae, a key phenotype white pupae (wp) has been used for decades to selectively remove females before releases, yet the gene responsible remained unknown. Here we use classical and modern genetic approaches to identify and functionally characterize causal wp− mutations in these distantly related fruit fly species. We find that the wp phenotype is produced by parallel mutations in a single, conserved gene. CRISPR/Cas9-mediated knockout of the wp gene leads to the rapid generation of novel white pupae strains in C. capitata and B. tryoni. The conserved phenotype and independent nature of the wp− mutations suggest that this technique can provide a generic approach to produce sexing strains in other major medical and agricultural insect pests.Competing Interest StatementThe authors have declared no competing interest.View Full Text

genetics↗

bioRxiv2019

Single-cell analysis of childhood leukemia reveals a link between developmental states and ribosomal protein expression as a source of intra-individual heterogeneity

Childhood acute lymphoblastic leukemia (cALL) is the most common pediatric cancer. It is characterized by bone marrow lymphoid precursors that acquire genetic alterations, resulting in disrupted maturation and uncontrollable proliferation. More than a dozen molecular subtypes of variable severity can be used to classify cALL cases. Modern therapy protocols currently cure 85-90% of cases, but other patients are refractory or will relapse and eventually succumb to their disease. To better understand these difficult cases, we investigated the nature and extent of intra-individual transcriptional heterogeneity of cALL at the cellular level by sequencing the transcriptomes of 39,375 individual cells in eight patients (six pre-B and two pre-T) and three healthy pediatric controls. We observed intra-individual transcriptional clusters in five out of the eight patients. Using pseudotime maturation trajectories of healthy B and T cells, we obtained the predicted developmental state of each leukemia cell and observed distribution shifts within patients. We showed that the predicted developmental states of these cancer cells are inversely correlated with ribosomal protein expression levels, which could be a common contributor to intra-individual heterogeneity in cALL patients.

cancer biology↗

bioRxiv2018

Analysis of head and neck carcinoma progression reveals novel and relevant stage-specific genetic changes associated with immortalisation and malignancy.

Head and neck squamous cell carcinoma (HNSCC) is a widely prevalent cancer globally with high mortality and morbidity. We report here changes in the genomic landscape in the development of HNSCC from potentially premalignant lesions (PPOLS) to malignancy and lymph node metastases. Frequent likely pathological mutations are restricted to a relatively small set of genes including TP53, CDKN2A, FBXW7, FAT1, NOTCH1 and KMT2D; these arise early in tumour progression and are present in PPOLs with NOTCH1 mutations restricted to cell lines from lesions that subsequently progressed to HNSCC. The most frequent genetic changes are of consistent somatic copy number alterations (SCNA). The earliest SCNAs involved deletions of CSMD1 (8p23.2), FHIT (3p14.2) and CDKN2A (9p21.3) together with gains of chromosome 20. CSMD1 deletions or promoter hypermethylation were present in all of the immortal PPOLs and occurred at high frequency in the immortal HNSCC cell lines (promoter hypermethylation ~63%, hemizygous deletions ~75%, homozygous deletions ~18%). Forced expression of CSMD1 in the HNSCC cell line H103 showed significant suppression of proliferation (p=0.0053) and invasion in vitro (p=5.98X10-5) supporting a role for CSMD1 inactivation in early head and neck carcinogenesis. In addition, knockdown of CSMD1 in the CSMD1-expressing BICR16 cell line showed significant stimulation of invasion in vitro (p=1.82 x 10-5) but not cell proliferation (p=0.239). HNSCC with and without nodal metastases showed some clear differences including high copy number gains of CCND1, hsa-miR-548k and TP63 in the metastases group. GISTIC peak SCNA regions showed significant enrichment (adj P<0.01) of genes in multiple KEGG cancer pathways at all stages with disruption of an increasing number of these involved in the progression to lymph node metastases. Sixty-seven genes from regions with statistically significant differences in SCNA/LOH frequency between immortal PPOL and HNSCC cell lines showed correlation with expression including 5 known cancer drivers.\n\nLay SummaryCancers affecting the head and neck region are relatively common. A large percentage of these are of one particular type; these are generally detected late and are associated with poor prognosis. Early detection and treatment dramatically improve survival and reduces the damage associated with the cancer and its treatment. Cancers arise and progress because of changes in the genetic material of the cells. This study focused on identifying such changes in these cancers particularly in the early stages of development, which are not fully known. Identification of these changes is important in developing new treatments as well as markers of behaviour of cancers and also the early or premalignant lesions. We used a well-characterised panel of cell lines generated from premalignant lesions as well as cancers, to identify mutations in genes, and an increase or decrease in number of copies of genes. We mapped new and previously identified changes in these cancers to specific stages in the development of these cancers and their spread. We additionally report here for the first time, alterations in CSMD1 gene in early premalignant lesions; we further show that this is likely to result in increased ability of the cells to spread and possibly, multiply faster as well.

cancer biology↗