Browsing by Subject "Transcriptomics"
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- PublicationEmbargoA rational approach to improving the biotechnological production of taxanes in plant cell cultures of Taxus spp.(Elsevier, 2014-03-27) Cusido, Rosa M.; Onrubia, Miriam; Sabater Jara, Ana Belén; Moyano, Elisabeth; Bonfill, Mercedes; Goossens, Alain; Pedreño, María Angeles; Palazón, Javier; Biología VegetalTaxol is a complex diterpene alkaloid scarcely produced in nature and with a high anticancer activity. Biotechnological systems for taxol production based on cell cultures of Taxus spp. have been developed, but the growing commercial demand for taxol and its precursors requires the optimization of these procedures. In order to increase the biotechnological production of taxol and related taxanes in Taxus spp. cell cultures, it is necessary not only to take an empirical approach that strives to optimize in-put factors (cell line selection, culture conditions, elicitation, up-scaling, etc.) and out-put factors (growth, production, yields, etc.), but also to carry out molecular biological studies. The latter can provide valuable insight into how the enhancement of taxane biosynthesis and accumulation affects metabolic profiles and gene expression in Taxus spp. cell cultures. Several rational approaches have focused on studying the transcriptomic profiles of key genes in the taxol biosynthetic pathway in Taxus spp. cell cultures treated with elicitors such as methyl jasmonate, coronatine and cyclodextrins in relation with the taxane pattern, production and excretion to the culture medium. These studies have provided new insights into the taxol biosynthetic pathway and its regulation. Additionally, identifying genes with low levels of expression even in the presence of elicitors, together with metabolomics studies, has shed light on the limiting steps in taxol biosynthesis and could help define suitable metabolic targets for engineering with the main aim of obtaining highly productive Taxus cultured cells. In this review, we have summarized the latest endeavors to enhance the molecular understanding of the action mechanism of elicitors in Taxus spp. cell cultures. Developments in the ongoing search for new and more effective elicitation treatments and the application of metabolic engineering to design new transgenic cell lines of Taxus with an improved capacity for taxane production are described.
- PublicationOpen AccessCommensal Bacteria Regulate Gene Expression and Differentiation in Vertebrate Olfactory Systems Through Transcription Factor REST(Oxford University Press, 2019-08-14) Casadei, Elisa; Tacchi, Luca; Lickwar, Colin R.; Espenschied, Scott T.; Davison, James M.; Muñoz, Pilar; Rawls, John F.; Salinas, Irene; Sanidad AnimalSensory systems such as the olfactory system detect chemical stimuli and thereby determine the relationships between the animal and its surroundings. Olfaction is one of the most conserved and ancient sensory systems in vertebrates. The vertebrate olfactory epithelium is colonized by complex microbial communities, but microbial contribution to host olfactory gene expression remains unknown. In this study, we show that colonization of germ-free zebrafish and mice with microbiota leads to widespread transcriptional responses in olfactory organs as measured in bulk tissue transcriptomics and RT-qPCR. Germ-free zebrafish olfactory epithelium showed defects in pseudostratification; however, the size of the olfactory pit and the length of the cilia were not different from that of colonized zebrafish. One of the mechanisms by which microbiota control host transcriptional programs is by differential expression and activity of specific transcription factors (TFs). REST (RE1 silencing transcription factor, also called NRSF) is a zinc finger TF that binds to the conserved motif repressor element 1 found in the promoter regions of many neuronal genes with functions in neuronal development and differentiation. Colonized zebrafish and mice showed increased nasal expression of REST, and genes with reduced expression in colonized animals were strongly enriched in REST-binding motifs. Nasal commensal bacteria promoted in vitro differentiation of Odora cells by regulating the kinetics of REST expression. REST knockdown resulted in decreased Odora cell differentiation in vitro. Our results identify a conserved mechanism by which microbiota regulate vertebrate olfactory transcriptional programs and reveal a new role for REST in sensory organs.
- PublicationOpen AccessExtensive dynamic changes in the human transcriptome and its circadian organization during prolonged bed rest(Cell Press, 2024-03-15) Archer, Simon N.; Möller-Levet, Carla; Bonmatí-Carrión, María Ángeles; Laing, Emma E.; Derk-Jan, Dijk; Anatomía Humana y PsicobiologíaPhysiological and molecular processes including the transcriptome change across the 24-h day, driven by molecular circadian clocks and behavioral and systemic factors. It is not known how the temporal organization of the human transcriptome responds to a long-lasting challenge. This may, however, provide insights into adaptation, disease, and recovery. We investigated the human 24-h time series transcriptome in 20 individuals during a 90-day constant bed rest protocol. We show that the protocol affected 91% of the transcriptome with 76% of the transcriptome still affected after 10 days of recovery. Dimensionality-reduction approaches revealed that many affected transcripts were associated with mRNA translation and immune function. The number, amplitude, and phase of rhythmic transcripts, including clock genes, varied significantly across the challenge. These findings of long-lasting changes in the temporal organization of the transcriptome have implications for understanding the mechanisms underlying health consequences of conditions such as microgravity and bed rest.
- PublicationOpen AccessHistological and transcriptomic analysis of paravaginal and central defects in anterior vaginal wall prolapse: Insights from DeLancey's pelvic floor theory(Universidad de Murcia, Departamento de Histología e Histopatología, 2025) Zhang Chun; Tang Qingxia; Zhou Yan; Fu Xuemei; Hu Pan; Liu Lubin; Biología Celular e HistologíaBackground. This study aimed to pre-liminarily explore the differences between paravaginal and central defect types of anterior vaginal wall prolapse based on DeLancey's pelvic floor theory. Methods. Seventy-eight patients with normal, paravaginal, or central defect vaginal wall tissues were collected and stained using hematoxylin and eosin (HE) and immunofluorescence staining to analyze and identify the expression of vimentin and phosphohistone H3 (PH3). Ribonucleic acid from fresh tissues was extracted for transcriptome sequencing to analyze differences between paravaginal and central defect types of anterior vaginal wall prolapse. Results. Significant differences were found in age, menopausal status, body mass index, pregnancy, and delivery among the control, paravaginal, and central defect groups. Histological analysis revealed that the distribution of interstitium in the normal HE staining group was compact and continuous. In the paravaginal defect interstitium, fiber morphology was altered, while central defect interstitial fibers were fragmented. PH3 expression was significantly lower in the central defect type than in the normal and paravaginal defect groups, suggesting degenerative lesions in the vaginal mucosa with central defects. Vimentin distribution in the normal group was tightly packed and continuous, whereas, in the paravaginal defect interstitium, vimentin filaments were fragmented into small spots and micro-aggregates. In the central defect interstitium, vimentin micro-aggregates exhibited altered coalescence and cell shape, appearing punctate. These findings indicated degenerative lesions in the anterior vaginal interstitium of both paravaginal and central defect types. KEGG enrichment analysis of differential genes revealed their involvement in proteinaceous extracellular matrix (ECM)-related signaling pathways, with increased expression of matrix metalloproteinase 13 (MMP13), MMP3, MMP12, and MMP7 in the paravaginal defect type compared with the central defect type. Conclusion. The differences between paravaginal and central defect types of anterior vaginal wall prolapse may be related to the expression of MMP-related proteins; KEGG enrichment analysis of differential genes indicated that they were closely related to the protein ECM pathway. Moreover, delineative lesions appeared in the paravaginal defect interstitium, and degenerative lesions appeared in the central defect mucosa and interstitium, which further enriched the DeLancey three-level theory
- PublicationOpen AccessIdentification of transcriptomic and associated DNA methylation changes Induced by the oocyte In vitro maturation in pigAbril-Parreño, Laura; Lopes, Jordana S.; Galvao, Antonio; Kelsey, Gavin; Coy, Pilar; FisiologíaIn humans, oocyte in vitro maturation (IVM) is an assisted reproductive technique used for patients with a high antral follicle count, polycystic ovary syndrome, and fertility preservation in cancer. In other cases, controlled ovarian hyperstimulation allowing in vivo maturation is the preferred method since it yields higher success rates. However, increasing evidence has revealed that hyperstimulation can lead to perturbed genomic imprinting and DNA methylation, which may impair offspring health. Therefore, IVM should be considered an alternative technique, but it is yet to be determined if similar epigenetic alterations also occur. Here we used the pig as a model to investigate the potential impacts of IVM on gene expression and DNA methylation in oocytes. To account for such variation, we used single-cell bisulphite sequencing and single-cell RNA sequencing techniques. The study was performed on 20 porcine oocytes obtained after IVM and 20 in vivo matured oocytes obtained by ovum pick-up from ex vivo ovaries. Differentially expressed genes (DEGs: P < 0.05, FC > 1.5) and differentially methylated regions (DMRs, P < 0.05, FC > 0.1) were determined and functionally annotated using Bioconductor packages in R. For the integration of both omics datasets, the single-cell aggregation and integration tool was used and a P < 0.05 and FC > 0.25 was applied to consider the interaction significant. Using the in vivo group as a reference, 1297 and 476 DEGs were down-regulated and up-regulated, respectively, in the IVM group. The up-regulated DEGs in IVM oocytes were mainly associated with the regulation of organelle organization, DNA methylation, and cell cycle processes; down-regulated genes were mainly enriched in ribosomal RNA processing, protein synthesis, oxidative phosphorylation and metabolomic processes such as glycosyl and aldehyde compound pathways. The global percentage of methylation was similar between the IVM and in vivo groups, but 321, 344, and 843 DMRs were detected (P < 0.05) in CpG islands, promoters and coding regions, respectively. Notably, the germline differentially regions of imprinted genes were appropriately methylated irrespective of IVM. Integrative analysis of DNA methylation and RNA-seq data identified the main methylated regions and genes that define each group. A total of 236 loci and 296 genes defined the IVM group, while 856 loci and 688 genes were related to the in vivo group. In addition, in the IVM group, we found a higher number of negative correlations between gene expression and DNA methylation, while the in vivo group showed higher number and stronger positive correlations. Taken together, these results indicate a discrete effect of IVM on the DNA methylation landscape in mature porcine oocytes, but these changes seem to have a greater impact on gene expression regulation.
- PublicationOpen AccessLaser capture microdissection: Big data from small samples(Universidad de Murcia. Departamento de Biología Celular e Histología, 2015) Datta, Soma; Malhotra, Lavina; Dickerson, Ryan; Chaffee, Scott; Sen, Chandan K.; Roy, SashwatiAny tissue is made up of a heterogeneous mix of spatially distributed cell types. In response to any (patho) physiological cue, responses of each cell type in any given tissue may be unique and cannot be homogenized across cell-types and spatial co-ordinates. For example, in response to myocardial infarction, on one hand myocytes and fibroblasts of the heart tissue respond differently. On the other hand, myocytes in the infarct core respond differently compared to those in the peri-infarct zone. Therefore, isolation of pure targeted cells is an important and essential step for the molecular analysis of cells involved in the progression of disease. Laser capture microdissection (LCM) is powerful to obtain a pure targeted cell subgroup, or even a single cell, quickly and precisely under the microscope, successfully tackling the problem of tissue heterogeneity in molecular analysis. This review presents an overview of LCM technology, the principles, advantages and limitations and its down-stream applications in the fields of proteomics, genomics and transcriptomics. With powerful technologies and appropriate applications, this technique provides unprecedented insights into cell biology from cells grown in their natural tissue habitat as opposed to those cultured in artificial petri dish conditions.
- PublicationOpen AccessThe transcriptome of pig spermatozoa, and its role in fertility(MDPI, 2020-02-25) Álvarez Rodríguez, Manuel; Martínez, Cristina; Wright, Dominic; Barranco, Isabel; Roca, Jordi; Rodríguez Martínez, Heriberto; Medicina y Cirugía AnimalIn the study presented here we identified transcriptomic markers for fertility in the cargo of pig ejaculated spermatozoa using porcine-specific micro-arrays (GeneChip® miRNA 4.0 and GeneChip® Porcine Gene 1.0 ST). We report (i) the relative abundance of the ssc-miR-1285, miR-16, miR-4332, miR-92a, miR-671-5p, miR-4334-5p, miR-425-5p, miR-191, miR-92b-5p and miR-15b miRNAs, and (ii) the presence of 347 up-regulated and 174 down-regulated RNA transcripts in high-fertility breeding boars, based on differences of farrowing rate (FS) and litter size (LS), relative to low-fertility boars in the (Artificial Insemination) AI program. An overrepresentation analysis of the protein class (PANTHER) identified significant fold-increases for C-C chemokine binding (GO:0019957): CCR7, which activates B- and T-lymphocytes, 8-fold increase), XCR1 and CXCR4 (with ubiquitin as a natural ligand, 1.24-fold increase), cytokine receptor activity (GO:0005126): IL23R receptor of the IL23 protein, associated to JAK2 and STAT3, 3.4-fold increase), the TGF-receptor (PC00035) genes ACVR1C and ACVR2B (12-fold increase). Moreover, two micro-RNAs (miR-221 and mir-621) were down- and up-regulated, respectively, in high-fertility males. In conclusion, boars with different fertility performance possess a wide variety of differentially expressed RNA present in spermatozoa that would be attractive targets as non-invasive molecular markers for predicting fertility.