| 1. |
- Klionsky, Daniel J., et al.
(författare)
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Guidelines for the use and interpretation of assays for monitoring autophagy
- 2012
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Ingår i: Autophagy. - : Informa UK Limited. - 1554-8635 .- 1554-8627. ; 8:4, s. 445-544
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Forskningsöversikt (refereegranskat)abstract
- In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field.
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| 2. |
- Acevedo, Nathalie, et al.
(författare)
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DNA Methylation Levels in Mononuclear Leukocytes from the Mother and Her Child Are Associated with IgE Sensitization to Allergens in Early Life
- 2021
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Ingår i: International Journal of Molecular Sciences. - : MDPI AG. - 1661-6596 .- 1422-0067. ; 22:2
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Tidskriftsartikel (refereegranskat)abstract
- DNA methylation changes may predispose becoming IgE-sensitized to allergens. We analyzed whether DNA methylation in peripheral blood mononuclear cells (PBMC) is associated with IgE sensitization at 5 years of age (5Y). DNA methylation was measured in 288 PBMC samples from 74 mother/child pairs from the birth cohort ALADDIN (Assessment of Lifestyle and Allergic Disease During INfancy) using the HumanMethylation450BeadChip (Illumina). PBMCs were obtained from the mothers during pregnancy and from their children in cord blood, at 2 years and 5Y. DNA methylation levels at each time point were compared between children with and without IgE sensitization to allergens at 5Y. For replication, CpG sites associated with IgE sensitization in ALADDIN were evaluated in whole blood DNA of 256 children, 4 years old, from the BAMSE (Swedish abbreviation for Children, Allergy, Milieu, Stockholm, Epidemiology) cohort. We found 34 differentially methylated regions (DMRs) associated with IgE sensitization to airborne allergens and 38 DMRs associated with sensitization to food allergens in children at 5Y (Sidak p <= 0.05). Genes associated with airborne sensitization were enriched in the pathway of endocytosis, while genes associated with food sensitization were enriched in focal adhesion, the bacterial invasion of epithelial cells, and leukocyte migration. Furthermore, 25 DMRs in maternal PBMCs were associated with IgE sensitization to airborne allergens in their children at 5Y, which were functionally annotated to the mTOR (mammalian Target of Rapamycin) signaling pathway. This study supports that DNA methylation is associated with IgE sensitization early in life and revealed new candidate genes for atopy. Moreover, our study provides evidence that maternal DNA methylation levels are associated with IgE sensitization in the child supporting early in utero effects on atopy predisposition.
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| 3. |
- Acevedo, Nathalie, et al.
(författare)
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Epigenetic alterations in skin homing CD4(+)CLA(+) T cells of atopic dermatitis patients
- 2020
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 10:1
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Tidskriftsartikel (refereegranskat)abstract
- T cells expressing the cutaneous lymphocyte antigen (CLA) mediate pathogenic inflammation in atopic dermatitis (AD). The molecular alterations contributing to their dysregulation remain unclear. With the aim to elucidate putative altered pathways in AD we profiled DNA methylation levels and miRNA expression in sorted T cell populations -(CD4(+), -CD4(+)CD45RA(+) naive, -CD4(+)CLA(+), and -CD8(+)) from adult AD patients and healthy controls (HC). Skin homing -CD4(+)CLA(+) T cells from AD patients showed significant differences in DNA methylation in 40 genes compared to HC (p < 0.05). Reduced DNA methylation levels in the upstream region of the interleukin-13 gene (IL13) in -CD4(+)CLA(+) T cells from AD patients correlated with increased IL13 mRNA expression in these cells. Sixteen miRNAs showed differential expression in -CD4(+)CLA(+) T cells from AD patients targeting genes in 202 biological processes (p < 0.05). An integrated network analysis of miRNAs and CpG sites identified two communities of strongly interconnected regulatory elements with strong antagonistic behaviours that recapitulated the differences between AD patients and HC. Functional analysis of the genes linked to these communities revealed their association with key cytokine signaling pathways, MAP kinase signaling and protein ubiquitination. Our findings support that epigenetic mechanisms play a role in the pathogenesis of AD by affecting inflammatory signaling molecules in skin homing -CD4(+)CLA(+) T cells and uncover putative molecules participating in AD pathways.
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| 4. |
- Acevedo, Nathalie
(författare)
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Epigenetic mechanisms of asthma and allergy
- 2015
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Allergic diseases such as atopic eczema (AE) and asthma are chronic inflammatory disorders in which the regulation of the immune system is altered, in concert with disease-specific defects in target organs. Their inception requires gene-environment interactions but the molecular mechanisms contributing to the pathogenesis remain unclear. During the last decades there was a dramatic increase in the prevalence of allergic diseases worldwide and several environmental conditions have been implicated (e.g. pollution, changes in lifestyle, reduced biodiversity). These exposures are supposed to affect leukocyte function, possibly through epigenetic mechanisms, which are the ultimate connection between the cell and the environment. The overall aim of this thesis was to elucidate the DNA methylation signatures as an epigenetic marker associated to cell lineage (I) and increasing age (IV) in primary blood leukocytes, and to compare DNA methylation (II, III) and miRNA expression (II) between healthy controls (HC) and patients with AE and asthma in connection with changes in mRNA expression. In study I, we found a particular DNA methylation landscape for each population of blood leukocytes that discriminates their lineage and that lymphoid cells are more methylated than myeloid cells. At the DNA methylation level, CD19+ B cells are the most different among all leukocytes. This study revealed that differences in cell composition may bias DNA methylation analyses conducted in DNA extracted from whole blood and provided a catalog of lineage-specific CpG sites. In study II, we compared DNA methylation and miRNA levels between AE patients and HC in four T cell populations isolated from peripheral blood. We discovered that circulating skin-homing memory CD4+CLA+ T cells isolated from AE patients have significant DNA methylation differences in 49 CpG sites and differential expression of 16 miRNAs compared to HC. Integrative data analyses with the differentially expressed mRNAs revealed that epigenetic signatures in AE patients are connected to the IL-10/IL-13 pathway. In study III, we analyzed the relation between genotype and epigenotype in the GSDMB/ORMDL3 asthma-susceptibility locus. We found significant differences in DNA methylation between children with controlled persistent asthma and HC. Methylation levels in three of these differentially methylated CpG sites correlate with the mRNA levels of ORMDL3. Causal inference tests revealed that both, single nucleotide polymorphisms and non-polymorphic CpG sites have independent effects on ORMDL3 mRNA expression. We also discovered that CD8+ T cells contain a differentially methylated region in the 5’UTR of ORMDL3 that is less methylated compared to other leukocyte subtypes, suggesting that methylation differences in ORMDL3 might promote lymphocyte driven inflammation. In study IV, we performed a pilot genome-wide DNA methylation analysis in whole blood, aiming to elucidate which genes had longitudinal changes in DNA methylation between 3 and 60 months after birth in healthy children. After filtering by lineage-CpG sites, we found 330 age-methylated CpG sites and 464 age-demethylated CpG sites that reflect the immunological window and are located on genes encoding for cytokines and chromatin remodelers. In summary, we discovered DNA methylation signatures in blood leukocytes associated to terminal differentiation and development, and demonstrated that compared to HC, patients suffering of AE and asthma have differences in DNA methylation and miRNA levels that might have functional impact on mRNA expression. The findings of this thesis provide new insights on genomic loci containing epigenetic differences between allergic patients and HC, that may be studied in the future as candidates for the development of preventive strategies, disease biomarkers or novel pharmacological therapies.
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| 5. |
- Acevedo, Nathalie, et al.
(författare)
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Genetic Variants in CHIA and CHI3L1 Are Associated with the IgE Response to the Ascaris Resistance Marker ABA-1 and the Birch Pollen Allergen Bet v 1
- 2016
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Ingår i: plos one. - : Public Library of Science (PLoS). - 1932-6203. ; 11:12
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Tidskriftsartikel (refereegranskat)abstract
- Helminth infections and allergic diseases are associated with IgE hyperresponsiveness but the genetics of this phenotype remain to be defined. Susceptibility to Ascaris lumbricoides infection and antibody levels to this helminth are associated with polymorphisms in locus 13q33-34. We aimed to explore this and other genomic regions to identify genetic variants associated with the IgE responsiveness in humans. Forty-eight subjects from Cartagena, Colombia, with extreme values of specific IgE to Ascaris and ABA-1, a resistance marker of this nematode, were selected for targeted resequencing. Burden analyses were done comparing extreme groups for IgE values. One-hundred one SNPs were genotyped in 1258 individuals of two well-characterized populations from Colombia and Sweden. Two low-frequency coding variants in the gene encoding the Acidic Mammalian Chitinase (CHIA rs79500525, rs139812869, tagged by rs10494133) were found enriched in high IgE responders to ABA-1 and confirmed by genetic association analyses. The SNP rs4950928 in the Chitinase 3 Like 1 gene (CHI3L1) was associated with high IgE to ABA-1 in Colombians and with high IgE to Bet v 1 in the Swedish population. CHIA rs10494133 and ABDH13 rs3783118 were associated with IgE responses to Ascaris. SNPs in the Tumor Necrosis Factor Superfamily Member 13b gene (TNFSF13B) encoding the cytokine B cell activating Factor were associated with high levels of total IgE in both populations. This is the first report on the association between low-frequency and common variants in the chitinases- related genes CHIA and CHI3L1 with the intensity of specific IgE to ABA-1 in a population naturally exposed to Ascaris and with Bet v 1 in a Swedish population. Our results add new information about the genetic influences of human IgE responsiveness; since the genes encode for enzymes involved in the immune response to parasitic infections, they could be helpful for understanding helminth immunity and allergic responses. We also confirmed that TNFSF13B has an important and conserved role in the regulation of total IgE levels, which supports potential evolutionary links between helminth immunity and allergic response.
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| 6. |
- Bernal, Ximena E., et al.
(författare)
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Empowering Latina scientists
- 2019
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Ingår i: Science. - : American Association for the Advancement of Science (AAAS). - 0036-8075 .- 1095-9203. ; 363:6429, s. 825-826
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)
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| 7. |
- Bornelöv, Susanne, et al.
(författare)
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Rule-Based Models of the Interplay between Genetic and Environmental Factors in Childhood Allergy
- 2013
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 8:11, s. e80080-
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Tidskriftsartikel (refereegranskat)abstract
- Both genetic and environmental factors are important for the development of allergic diseases. However, a detailed understanding of how such factors act together is lacking. To elucidate the interplay between genetic and environmental factors in allergic diseases, we used a novel bioinformatics approach that combines feature selection and machine learning. In two materials, PARSIFAL (a European cross-sectional study of 3113 children) and BAMSE (a Swedish birth-cohort including 2033 children), genetic variants as well as environmental and lifestyle factors were evaluated for their contribution to allergic phenotypes. Monte Carlo feature selection and rule based models were used to identify and rank rules describing how combinations of genetic and environmental factors affect the risk of allergic diseases. Novel interactions between genes were suggested and replicated, such as between ORMDL3 and RORA, where certain genotype combinations gave odds ratios for current asthma of 2.1 (95% CI 1.2-3.6) and 3.2 (95% CI 2.0-5.0) in the BAMSE and PARSIFAL children, respectively. Several combinations of environmental factors appeared to be important for the development of allergic disease in children. For example, use of baby formula and antibiotics early in life was associated with an odds ratio of 7.4 (95% CI 4.5-12.0) of developing asthma. Furthermore, genetic variants together with environmental factors seemed to play a role for allergic diseases, such as the use of antibiotics early in life and COL29A1 variants for asthma, and farm living and NPSR1 variants for allergic eczema. Overall, combinations of environmental and life style factors appeared more frequently in the models than combinations solely involving genes. In conclusion, a new bioinformatics approach is described for analyzing complex data, including extensive genetic and environmental information. Interactions identified with this approach could provide useful hints for further in-depth studies of etiological mechanisms and may also strengthen the basis for risk assessment and prevention.
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| 8. |
- Cantù, Claudio, et al.
(författare)
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Mutations in Bcl9 and Pygo genes cause congenital heart defects by tissue-specific perturbation of Wnt/beta-catenin signaling
- 2018
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Ingår i: Genes & Development. - : COLD SPRING HARBOR LAB PRESS, PUBLICATIONS DEPT. - 0890-9369 .- 1549-5477. ; 32:21-22, s. 1443-1458
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Tidskriftsartikel (refereegranskat)abstract
- Bcl9 and Pygopus (Pygo) are obligate Wnt/beta-catenin cofactors in Drosophila, yet their contribution to Wnt signaling during vertebrate development remains unresolved. Combining zebrafish and mouse genetics, we document a conserved, beta-catenin-associated function for BCL9 and Pygo proteins during vertebrate heart development. Disrupting the beta-catenin-BCL9-Pygo complex results in a broadly maintained canonical Wnt response yet perturbs heart development and proper expression of key cardiac regulators. Our work highlights BCL9 and Pygo as selective beta-catenin cofactors in a subset of canonical Wnt responses during vertebrate development. Moreover, our results implicate alterations in BCL9 and BCL9L in human congenital heart defects.
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| 9. |
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| 10. |
- Gomez-Garrido, Jessica, et al.
(författare)
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Chromosome-level genome assembly of Lilford's wall lizard, Podarcis lilfordi (Günther, 1874) from the Balearic Islands (Spain)
- 2023
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Ingår i: DNA Research. - 1340-2838. ; 30:3
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Tidskriftsartikel (refereegranskat)abstract
- The Mediterranean lizard Podarcis lilfordi is an emblematic species of the Balearic Islands. The extensive phenotypic diversity among extant isolated populations makes the species a great insular model system for eco-evolutionary studies, as well as a challenging target for conservation management plans. Here we report the first high-quality chromosome-level assembly and annotation of the P. lilfordi genome, along with its mitogenome, based on a mixed sequencing strategy (10X Genomics linked reads, Oxford Nanopore Technologies long reads and Hi-C scaffolding) coupled with extensive transcriptomic data (Illumina and PacBio). The genome assembly (1.5 Gb) is highly contiguous (N50 = 90 Mb) and complete, with 99% of the sequence assigned to candidate chromosomal sequences and >97% gene completeness. We annotated a total of 25,663 protein-coding genes translating into 38,615 proteins. Comparison to the genome of the related species Podarcis muralis revealed substantial similarity in genome size, annotation metrics, repeat content, and a strong collinearity, despite their evolutionary distance (∼18-20 MYA). This genome expands the repertoire of available reptilian genomes and will facilitate the exploration of the molecular and evolutionary processes underlying the extraordinary phenotypic diversity of this insular species, while providing a critical resource for conservation genomics.
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