👤 Sabrina Dumont

Mobile elements (ME) can transpose by copy-and-paste mechanisms. A heterozygous insertion in APOB exon 3 coding sequence was suspected in a patient with hypobetalipoproteinemia (HBL), by gel electrophoresis of the PCR products. An insertion of a 85 bp fragment flanked by a polyA stretch and a target replication site duplication was identified as a ME insertion (MEI) from the AluYa5 subfamily, NM₀₀₀₃₈₄.3(APOB):c.135₁₃₆ins(160). Then, the DNA was reanalyzed using our NGS custom panel. Routine analysis did not reveal any causative variant, but manual inspection of the alignments and MELT enabled us to detect this MEI from NGS data. A functional study revealed that this MEI introduces a stop codon p.(Phe46Alafs*2) and additionally leads to p.(Lys41Serfs*2) due to an exon skipping. This is the first report of a MEI into APOB, as a cause of HBL. Furthermore, our study highlights the value of including MEI-callers in routine pipelines to improve primary dyslipidemia diagnosis. Show less

Computer vision is increasingly used in farmers' fields and agricultural experiments to quantify important traits. Imaging setups with a sub-millimeter ground sampling distance enable the detection and tracking of plant features, including size, shape, and colour. Although today's AI-driven foundation models segment almost any object in an image, they still fail for complex plant canopies. To improve model performance, the global wheat dataset consortium assembled a diverse set of images from experiments around the globe. After the head detection dataset (GWHD), the new dataset targets a full semantic segmentation (GWFSS) of organs (leaves, stems and spikes) covering all developmental stages. Images were collected by 11 institutions using a wide range of imaging setups. Two datasets are provided: i) a set of 1096 diverse images in which all organs were labelled at the pixel level, and (ii) a dataset of 52,078 images without annotations available for additional training. The labelled set was used to train segmentation models based on DeepLabV3Plus and Segformer. Our Segformer model performed slightly better than DeepLabV3Plus with a mIOU for leaves and spikes of ca. 90 ​%. However, the precision for stems with 54 ​% was rather lower. The major advantages over published models are: i) the exclusion of weeds from the wheat canopy, ii) the detection of all wheat features including necrotic and senescent tissues and its separation from crop residues. This facilitates further development in classifying healthy vs. unhealthy tissue to address the increasing need for accurate quantification of senescence and diseases in wheat canopies. Show less

Optimal molecular diagnosis of primary dyslipidemia is challenging to confirm the diagnosis, test and identify at risk relatives. The aim of this study was to test the application of a single targeted next-generation sequencing (NGS) panel for hypercholesterolemia, hypocholesterolemia, and hypertriglyceridemia molecular diagnosis. NGS workflow based on a custom AmpliSeq panel was designed for sequencing the most prevalent dyslipidemia-causing genes (ANGPTL3, APOA5, APOC2, APOB, GPIHBP1, LDLR, LMF1, LPL, PCSK9) on the Ion PGM Sequencer. One hundred and forty patients without molecular diagnosis were studied. In silico analyses were performed using the NextGENe software and homemade tools for detection of copy number variations (CNV). All mutations were confirmed using appropriate tools. Eighty seven variations and 4 CNV were identified, allowing a molecular diagnosis for 40/116 hypercholesterolemic patients, 5/13 hypocholesterolemic patients, and 2/11, hypertriglyceridemic patients respectively. This workflow allowed the detection of CNV contrary to our previous strategy. Some variations were found in previously unexplored regions providing an added value for genotype-phenotype correlation and familial screening. In conclusion, this new NGS process is an effective mutation detection method and allows better understanding of phenotype. Consequently this assay meets the medical need for individualized diagnosis of dyslipidemia. Show less

The Wnt/β-catenin pathway is the most frequently deregulated pathway in hepatocellular carcinoma (HCC). Inactivating mutations of the gene encoding AXIN1, a known negative regulator of the Wnt/β-catenin signaling pathway, are observed in about 10% of HCCs. Whole-genome studies usually place HCC with AXIN1 mutations and CTNNB1 mutations in the group of tumors with Wnt/β-catenin activated program. However, it has been shown that HCCs with activating CTNNB1 mutations form a group of HCCs, with a different histology, prognosis and genomic signature to those with inactivating biallelic AXIN1 mutations. We aimed to elucidate the relationship between CTNNB1 mutations, AXIN1 mutations and the activation level of the Wnt/β-catenin program. We evaluated two independent human HCC datasets for the expression of a 23-β-catenin target genes program. We modeled Axin1 loss of function tumorigenesis in two engineered mouse models and performed gene expression profiling. Based on gene expression, we defined three levels of β-catenin program activation: strong, weak or no activation. While more than 80% CTNNB1-mutated tumors were found in the strong or in the weak activation program, most of the AXIN1-mutated tumors (>70%) were found in the subgroup with no activation. We validated this result by demonstrating that mice with a hepatocyte specific AXIN1 deletion developed HCC in the absence of β-catenin induction. We defined a 329-gene signature common in human and mouse AXIN1 mutated HCC that is highly enriched in Notch and YAP oncogenic signatures. AXIN1-mutated HCCs occur independently of the Wnt/β-catenin pathway and involve Notch and YAP pathways. These pathways constitute potentially interesting targets for the treatment of HCC caused by AXIN1 mutations. Liver cancer has a poor prognosis. Defining the molecular pathways involved is important for developing new therapeutic approaches. The Wnt/β-catenin pathway is the most frequently deregulated pathway in hepatocellular carcinoma (HCC). Mutations of AXIN1, a member of this pathway, represent about 10% of HCC mutations. Using both human HCC collections and engineered mouse models of liver cancers with AXIN1 mutation or deletion, we defined a common signature of liver tumors mutated for AXIN1 and demonstrate that these tumors occur independently of the activation of the Wnt/β-catenin pathway. Show less

Blood polyunsaturated fatty acid (PUFA) levels are determined by diet and by endogenous synthesis via Δ5- and Δ6-desaturases (encoded by the FADS1 and FADS2 genes, respectively). Genome-wide association studies have reported associations between FADS1-FADS2 polymorphisms and the plasma concentrations of PUFAs, HDL- and LDL-cholesterol, and triglycerides. However, much remains unknown regarding the molecular mechanisms explaining how variants affect the function of FADS1-FADS2 genes. Here, we sought to identify the functional variant(s) within the FADS gene cluster. To address this question, we (1) genotyped individuals (n = 540) for the rs174547 polymorphism to confirm associations with PUFA levels used as surrogate estimates of desaturase activities and (2) examined the functionality of variants in linkage disequilibrium with rs174547 using bioinformatics and luciferase reporter assays. The rs174547 minor allele was associated with higher erythrocyte levels of dihomo-γ-linolenic acid and lower levels of arachidonic acid, suggesting a lower Δ5-desaturase activity. In silico analyses suggested that rs174545 and rs174546, in perfect linkage disequilibrium with rs174547, might alter miRNA binding sites in the FADS1 3'UTR. In HuH7 and HepG2 cells transfected with FADS1 3'UTR luciferase vectors, the haplotype constructs bearing the rs174546T minor allele showed 30% less luciferase activity. This relative decrease reached 60% in the presence of miR-149-5p and was partly abolished by cotransfection with an miR-149-5p inhibitor. This study identifies FADS1 rs174546 as a functional variant that may explain the associations between FADS1-FADS2 polymorphisms and lipid-related phenotypes. Show less

Blood levels of polyunsaturated fatty acids (PUFAs) are under control of endogenous synthesis via Δ5- and Δ6-desaturases, encoded by the FADS1 and FADS2 genes, respectively and of diet. Genome-wide associations studies (GWAS) reported associations between polymorphisms in FADS1-FADS2 and variations in plasma concentrations of PUFAs, HDL- and LDL-cholesterol and triglycerides. However, it is not established whether dietary PUFAs intake modulates these associations. We assessed whether dietary linoleic acid (LA) or α-linolenic acid (ALA) modulate the association between the FADS1 rs174547 polymorphism (a GWAS hit) and lipid and anthropometric phenotypes. Dietary intakes of LA and ALA, FADS1 rs174547 genotypes, lipid and anthropometric variables were determined in three French population-based samples (n = 3069). These samples were stratified according to the median dietary LA (<9.5 and ≥9.5 g/d) and ALA (<0.80 and ≥0.80 g/d) intakes. The meta-analysis was performed using a random-effect. Our meta-analysis confirmed the association between rs174547 and plasma lipid levels and revealed an association with waist circumference and body mass index. These associations were not modified by dietary ALA intake (all p-interaction > 0.05). In contrast, the associations with HDL-cholesterol levels, waist circumference and BMI were modulated by the dietary intake of LA (p interaction < 0.05). In high LA consumers only, the rs174547 minor allele was significantly associated with lower HDL-cholesterol levels (β = -0.05 mmol/L, p = 0.0002). Furthermore, each copy of the rs174547 minor allele was associated with a 1.58 cm lower waist circumference (p = 0.0005) and a 0.46 kg m The present study suggests that dietary LA intake may modulate the association between the FADS gene variants and HDL-cholesterol concentration, waist circumference and BMI. These gene-nutrient interactions, if confirmed, suggest that subjects carrying the rs174547 minor allele might benefit from low dietary LA intakes. Show less

Molecular alterations of the MAPK pathway are frequently observed in papillary thyroid carcinomas (PTCs). It leads to a constitutive activation of the signalling pathway through an increase in MEK and ERK phosphorylation. ERK is negatively feedback-regulated by Dual Specificity Phosphatases (DUSPs), especially two ERK-specific DUSPs, DUSP5 (nuclear) and DUSP6 (cytosolic). These negative MAPK regulators may play a role in thyroid carcinogenesis. MAPK pathway activation was analyzed in 11 human thyroid cancer cell lines. Both phosphatases were studied in three PCCL3 rat thyroid cell lines that express doxycycline inducible PTC oncogenes (RET/PTC3, H-RASV12 or BRAFV600E). Expression levels of DUSP5 and DUSP6 were quantified in 39 human PTCs. The functional role of DUSP5 and DUSP6 was investigated through their silencing in two human BRAFV600E carcinoma cell lines. BRAFV600E human thyroid cancer cell lines expressed higher phospho-MEK levels but not higher phospho-ERK levels. DUSP5 and DUSP6 are specifically induced by the MEK-ERK pathway in the three PTC oncogenes inducible thyroid cell lines. This negative feedback loop explains the tight regulation of p-ERK levels. DUSP5 and DUSP6 mRNA are overexpressed in human PTCs, especially in BRAFV600E mutated PTCs. DUSP5 and/or DUSP6 siRNA inactivation did not affect proliferation in two BRAFV600E mutated cell lines, which may be explained by a compensatory increase in other phosphatases. In the light of this, we observed a marked DUSP6 upregulation upon DUSP5 inactivation. Despite this, DUSP5 and DUSP6 positively control cell migration and invasion. Our results are in favor of a stronger activation of the MAPK pathway in BRAFV600E PTCs. DUSP5 and DUSP6 have pro-tumorigenic properties in two BRAFV600E PTC cell line models. Show less

To build the spindle at mitosis, motors exert spatially regulated forces on microtubules. We know that dynein pulls on mammalian spindle microtubule minus-ends, and this localized activity at ends is predicted to allow dynein to cluster microtubules into poles. How dynein becomes enriched at minus-ends is not known. Here, we use quantitative imaging and laser ablation to show that NuMA targets dynactin to minus-ends, localizing dynein activity there. NuMA is recruited to new minus-ends independently of dynein and more quickly than dynactin; both NuMA and dynactin display specific, steady-state binding at minus-ends. NuMA localization to minus-ends involves a C-terminal region outside NuMA's canonical microtubule-binding domain and is independent of minus-end binders γ-TuRC, CAMSAP1, and KANSL1/3. Both NuMA's minus-end-binding and dynein-dynactin-binding modules are required to rescue focused, bipolar spindle organization. Thus, NuMA may serve as a mitosis-specific minus-end cargo adaptor, targeting dynein activity to minus-ends to cluster spindle microtubules into poles. Show less

A growing body of evidence suggests that microRNAs (miRNAs) are involved in Alzheimer's disease (AD) and that some disease-associated genetic variants are located within miRNA binding sites. In the present study, we sought to characterize functional polymorphisms in miRNA target sites within the loci defined in earlier genome-wide association studies (GWAS). The main objectives of this study were to (1) facilitate the identification of the gene or genes responsible for the GWAS signal within a locus of interest and (2) determine how functional polymorphisms might be involved in the AD process (e.g., by affecting miRNA-mediated variations in gene expression). Stringent in silico analyses were developed to select potential polymorphisms susceptible to impairment of miRNA-mediated repression, and subsequent functional assays were performed in HeLa and HEK293 cells. Two polymorphisms were identified and further analyzed in vitro. The AD-associated rs7143400-T allele (located in 3' untranslated region [3'-UTR] of FERMT2) cotransfected with miR-4504 resulted in lower protein levels relative to the rs7143400-G allele cotransfected with the same miRNA. The AD-associated rs9909-C allele in the 3'-UTR of NUP160 abolished the miR-1185-1-3p-regulated expression observed for the rs9909-G allele. When considered in conjunction with the findings of previous association studies, our results suggest that decreased expression of FERMT2 might be a risk factor in the etiopathology of AD, whereas increased expression of NUP160 might protect against the disease. Our data therefore provide new insights into AD by highlighting two new proteins putatively involved in the disease process. Show less

The glucose-activated transcription factor carbohydrate response element binding protein (ChREBP) induces the expression of hepatic glycolytic and lipogenic genes. The farnesoid X receptor (FXR) is a nuclear bile acid receptor controlling bile acid, lipid, and glucose homeostasis. FXR negatively regulates hepatic glycolysis and lipogenesis in mouse liver. The aim of this study was to determine whether FXR regulates the transcriptional activity of ChREBP in human hepatocytes and to unravel the underlying molecular mechanisms. Agonist-activated FXR inhibits glucose-induced transcription of several glycolytic genes, including the liver-type pyruvate kinase gene (L-PK), in the immortalized human hepatocyte (IHH) and HepaRG cell lines. This inhibition requires the L4L3 region of the L-PK promoter, known to bind the transcription factors ChREBP and hepatocyte nuclear factor 4α (HNF4α). FXR interacts directly with ChREBP and HNF4α proteins. Analysis of the protein complex bound to the L4L3 region reveals the presence of ChREBP, HNF4α, FXR, and the transcriptional coactivators p300 and CBP at high glucose concentrations. FXR activation does not affect either FXR or HNF4α binding to the L4L3 region but does result in the concomitant release of ChREBP, p300, and CBP and in the recruitment of the transcriptional corepressor SMRT. Thus, FXR transrepresses the expression of genes involved in glycolysis in human hepatocytes. Show less

Two rate-limiting enzymes in PUFA biosynthesis, Δ5- and Δ6-desaturases, are encoded by the FADS1 and FADS2 genes, respectively. Genetic variants in the FADS1-FADS2 gene cluster are associated with changes in plasma concentrations of PUFA, HDL- and LDL-cholesterol, and TG. However, little is known about whether dietary PUFA intake modulates these associations, especially in adolescents. We assessed whether dietary linoleic acid (LA) or α-linolenic acid (ALA) modulate the association between the FADS1 rs174546 polymorphism and concentrations of PUFA, other lipids, and lipoproteins in adolescents. Dietary intakes of LA and ALA, FADS1 rs174546 genotypes, PUFA levels in serum phospholipids, and serum concentrations of TG, cholesterol, and lipoproteins were determined in 573 European adolescents from the HELENA study. The sample was stratified according to the median dietary LA (≤9.4 and >9.4 g/d) and ALA (≤1.4 and >1.4 g/d) intakes. The associations between FADS1 rs174546 and concentrations of PUFA, TG, cholesterol, and lipoproteins were not affected by dietary LA intake (all P-interaction > 0.05). Similarly, the association between the FADS1 rs174546 polymorphism and serum phospholipid concentrations of ALA or EPA was not modified by dietary ALA intake (all P-interaction > 0.05). In contrast, the rs174546 minor allele was associated with lower total cholesterol concentrations (P = 0.01 under the dominant model) and non-HDL-cholesterol concentrations (P = 0.02 under the dominant model) in the high-ALA-intake group but not in the low-ALA-intake group (P-interaction = 0.01). These results suggest that dietary ALA intake modulates the association between FADS1 rs174546 and serum total and non-HDL-cholesterol concentrations at a young age. Show less

Genetic variability in the FADS1-FADS2 gene cluster [encoding delta-5 (D5D) and delta-6 (D6D) desaturases] has been associated with plasma long-chain PUFA (LCPUFA) and lipid levels in adults. To better understand these relationships, we further characterized the association between FADS1-FADS2 genetic variability and D5D and D6D activities in adolescents. Thirteen single nucleotide polymorphisms (SNPs) were genotyped in 1,144 European adolescents (mean +/- SD age: 14.7 +/- 1.4 y). Serum phospholipid fatty acid levels were analyzed using gas chromatography. D5D and D6D activities were estimated from the C20:4n-6/C20:3n-6 and C20:3n-6/C18:2n-6 ratios, respectively. Minor alleles of nine SNPs were associated with higher 18:2n-6 levels (1.9E-18 Show less