Archives des Bases moléculaires et structurales du vivant

Vulnerability of the placenta to air pollution: what effects on the unborn child’s development?

Posted on May 7, 2024May 13, 2024 by Graziella Cara

How does exposure to air pollution affect the proper course of pregnancy and the development of the unborn child? A research team from Inserm and Université Grenoble Alpes investigated the potential effects on placental DNA of exposure to three major airborne pollutants. When comparing the data obtained from around 1 500 pregnant women, it observed that exposure to these pollutants during pregnancy was associated with epigenetic changes[1] liable to alter the development of the foetus, particularly at the metabolic, immune and neurological levels. Its findings, to be published in The Lancet Planetary Health, also show that the periods of susceptibility to air pollutants differ according to the sex of the foetus, thereby impacting the development of girls and boys in different ways.

Exposure to outdoor air pollution presents a major risk for the proper course of pregnancy. It is particularly suspected of causing cardiometabolic, respiratory and neuropsychological pathologies in unborn children. However, while its physiological effects have been studied, the molecular mechanisms involved remain poorly understood.

The placenta is an organ that plays a key role in foetal development. Particularly vulnerable to many chemical compounds, it can be likened to an ‘archive’ that reflects the child’s prenatal environment: the epigenetic changes occurring in its cells partially reflect the mother’s environmental exposures during pregnancy. To study these changes, we usually measure the level of DNA methylation, one of the best known epigenetic mechanisms involved in gene control and expression.

A team led by Inserm researcher Johanna Lepeule at the Institute for Advanced Biosciences (Inserm/CNRS/Université Grenoble Alpes) investigated the impact of three air pollutants – nitrogen dioxide (NO₂) and particulate matter (PM_2.5and PM₁₀) – on placental DNA methylation. Thanks to data from three French mother-child cohorts[2], it was able to compare exposure to these pollutants and methylation levels in over 1 500 participants during their pregnancy.

Its findings show a significant impact of exposure to the three airborne pollutants on placental DNA methylation levels concerning genes involved in foetal development. One third of these changes were directly associated with indicators of child development (birthweight, birth length, head circumference, duration of pregnancy, etc.).

Other placental changes concerned genes involved in the development of the nervous system, immune system and the metabolism – including genes involved in the onset of neonatal diabetes or obesity.

While these alterations in methylation are present in both sexes, the scientists also identified changes that have an additional impact and affect different genes depending on the sex of the unborn child. Two different gestation periods that are particularly vulnerable to epigenetic changes under the effect of the pollutants have emerged from this research: the start of pregnancy (1st trimester) in boys and the end of pregnancy (3rd trimester) in girls.

‘Our findings show that exposure to air pollution during pregnancy induces changes in placental DNA methylation specific to each of the two sexes, says Lepeule. This differentiated impact could contribute to alterations in the development and course of pregnancy that differ according to the sex of the unborn child.’

Thus in boys, significant alterations in methylation have been detected in genes that are critically involved in the development of the nervous system and intellect.

‘These observations support the growing number of studies linking exposure to air pollution during pregnancy with impaired neurodevelopment and/or reduced cognitive capacities, with greater vulnerability among male children,’ explains Lucile Broséus, Inserm researcher and first author of the publication.

In girls, the methylation affected genes involved in foetal development and the regulation of oxidative stress. It could therefore be associated with developmental defects liable to increase the risk of developing chronic metabolic diseases (hypertension, diabetes, obesity, etc.) later in life, but also with the occurrence of miscarriage or pre-eclampsia in the mother[3].

This research therefore provides new data on the epigenetic mechanisms involved in the deregulation of foetal growth under the effect of air pollution and which could cause long-term changes in the metabolism.

‘Future studies may investigate whether placental epigenetic changes caused by exposure to air pollution during pregnancy persist after childbirth and how they could influence development during childhood, adds Lepeule. In addition, as this research has been carried out on French cohorts, its findings will have to be verified in populations from other geographical regions and with different genetic profiles,’ concludes the researcher.

[1] Epigenetic changes are materialised by biochemical marks present on DNA. Reversible, they do not lead to changes in DNA sequence but do induce changes in gene expression. They are induced by the wider environment: the cell receives signals informing it about its environment, and specialises itself accordingly, or adjusts its activity.

[2]EDEN, led by Inserm, Poitiers University Hospital and Nancy University Hospital; PELAGIE, led by Inserm; and SEPAGES, led by Inserm and Grenoble Alpes University Hospital.

[3]Pre-eclampsia is a complication of pregnancy characterised by elevated blood pressure and increased protein in the urine. It can occur in the middle of the second trimester or later, shortly before delivery or sometimes even after. Responsible for one third of the births of very preterm infants in France, this syndrome is a major cause of intrauterine growth retardation. If left untreated, it can lead to the death of the mother, child, or both.

Cell contraction drive the initial shaping of human embryos

Posted on May 1, 2024May 2, 2024 by Graziella Cara

Human embryo at the blastocyst stage ready to implant. The nuclear envelope of the cells appears in blue and the actin cytoskeleton in orange. © Julie Firmin et Jean-Léon Maître (Institut Curie, Université PSL, CNRS UMR3215, INSERM U934)

Human embryo compaction, an essential step in the first days of an embryo’s development, is driven by the contractility of its cells. This is the finding of a team of scientists from CNRS, Institut Curie, Inserm, AP-HP and the Collège de France. Published in the 1^st May edition of Nature, these results contradict the presupposed driving role of cell adhesion in this phenomenon and pave the way for improved assisted reproductive technology (ART).

In humans, embryonic cell compaction is a crucial step in the normal development of an embryo. Four days after fertilisation, cells move closer together to give the embryo its initial shape. Defective compaction prevents the formation of the structure that ensures the embryo can implant in the uterus. In assisted reproductive technology (ART), this stage is carefully monitored before an embryo is implanted.

An interdisciplinary research team¹ led by scientists at the Genetics and Developmental Biology Unit at the Institut Curie (CNRS/Inserm/Institut Curie) studying the mechanisms at play in this still little-known phenomenon has made a surprising discovery: human embryo compaction is driven by the contraction of embryonic cells. Compaction problems are therefore due to faulty contractility in these cells, and not a lack of adhesion between them, as was previously assumed. This mechanism had already been identified in flies, zebrafish and mice, but is a first in humans.

By improving our understanding of the early stages of human embryonic development, the research team hopes to contribute to the refinement of ART as nearly one third of inseminations are unsuccessful today.²

The results were obtained by mapping cell surface tensions in human embryonic cells. The scientists also tested the effects of inhibiting contractility and cell adhesion and analysed the mechanical signature of embryonic cells with defective contractility.

Human embryo at the 4-cell stage. Cell DNA appears in red and their actin cytoskeleton in blue. The cell on the right has just split its genome into two and is about to divide. © Julie Firmin et Jean-Léon Maître (Institut Curie, Université PSL, CNRS UMR3215, INSERM U934)

1 – Scientists from the following entities also took part in the study: the Centre interdisciplinaire de recherche en biologie (CNRS/Collège de France/Inserm), the Reproductive Biology Department – CECOS (AP-HP) and the Institut Cochin (CNRS/Inserm/Université Paris Cité).

2 – Source: Agence de la biomédecine

Obesity: opt for omega 3 fatty acids to prevent the associated risks

Posted on February 20, 2024March 8, 2024 by Graziella Cara

In yellow, microglia (immune cells of the brain) activated by the pro-inflammatory nature of a sunflower oil-enriched diet (fluorescence microscopy). © Clara Sanchez/Inserm

Obesity is a major public health problem, affecting around 650 million adults worldwide[1], and is often associated with systemic and cerebral inflammation as well as anxiety and cognitive disorders, such as memory deficits. In a new study, researchers from Inserm, CNRS and Université Côte d’Azur at the Institute of Molecular and Cellular Pharmacology tried to understand more precisely how diet can cause obesity, and its associated comorbidities. They focused more specifically on omega 6 (ω6) and omega 3 (ω3) fatty acids, exploring the health effects of various diets with different ratios of fatty acids (see box below). Their findings indicate that a diet enriched with ω6 (in this case, sunflower oil) is strongly associated with changes in metabolism, inflammation and cognitive functions, whereas one enriched with ω3 (in this case, rapeseed oil) has certain preventive effects. This research makes it possible to envisage dietary interventions based on a low ω6/ω3 ratio (thus with a preference for rapeseed oil over sunflower oil) to combat obesity and its associated neurological disorders. These findings have been published in Brain Behavior and Immunity.

According to the WHO, cases of obesity have almost tripled in number worldwide since 1975. Obesity is associated with numerous comorbidities (type 2 diabetes, cardiovascular diseases, osteoarthritis, cancer and cognitive disorders) and high mortality. While its causes are complex and involve the interaction of several factors, dietary imbalance is recognised as being the major contributing factor.

What is more, previous studies[1] have shown that obesity is associated not only with metabolic dysfunction, but also chronic inflammation in the peripheral organs (adipose tissues, liver, skeletal muscles and pancreas), as well as in the central nervous system (neuroinflammation). This neuroinflammation in obesity is characterised by an increase in pro-inflammatory markers in the region of the hypothalamus, an area of the brain known to control dietary behaviour[2]. However, the nature of the nutritional lipids that could be responsible for this neuroinflammation has not yet been elucidated.

In a new study, researchers from Inserm, CNRS and Université Côte d’Azur specifically focused on certain fatty acids that are essential for our bodies to function properly, and are known for having anti- and pro-inflammatory properties: omega 3 and 6 (see box below). Their objective was to better understand whether omega 3 and 6 are involved in the phenomenon of neuroinflammation in the context of a high-fat diet (so-called ‘obesogenic diet’), and whether they can be associated with the development of obesity.

Their research is also based on the observation of an increasingly strong trend in developed countries towards an excessive consumption of omega 6, whose inflammatory properties are abundantly documented in the scientific literature[4].

Omega 3 and omega 6: the importance of getting the right balance

Omega 3 and omega 6 fatty acids are essential for the correct functioning of the body, which is unable to produce or synthesise them on its own. They therefore need to come from the diet and respect a certain balance (referred to as the omega 6/omega 3 ratio), in order to combine the pro-inflammatory properties of omega 6 with the anti-inflammatory properties of omega 3.

Omega 6 fatty acids: e.g. linoleic and gamma-linolenic acids, are found in many oils, such as sunflower and corn oils
Omega 3 fatty acids: e.g. eicosapentaenoic and docosahexaenoic acids, are found in oily fish, and alpha-linolenic acid in oils such as flax, hemp, rapeseed, walnut or soya

In animal models, the scientists evaluated the health effects of three obesogenic diets – high in lipids – each with a different fatty acids ratio.

In these diets, the researchers used vegetable oils that can be found in the shops, namely rapeseed (rich in omega 3) and sunflower (rich in omega 6). The first diet contained a high omega 6/omega 3 ratio, meaning that it was highly enriched in omega 6 and therefore in sunflower oil. The second had an intermediate ratio, with balanced levels of omega 3 and omega 6. And the third was highly enriched in omega 3 and therefore in rapeseed oil.

Through different examinations, the scientists measured the various effects of these diets on weight gain and fat storage, glucose homeostasis[5] response, development of anxiety and cognitive disorders, and brain inflammation.

At the end of the experiment, which lasted up to 5 months, the scientists observed (results summarised in the diagram below):

altered metabolism, neuroinflammation and cognitive functions, including increased anxiety and spatial memory disorders in the obese mice fed a diet enriched with omega 6, and therefore sunflower oil,
a protective effect of the omega 3-enriched, high-rapeseed oil diet, on weight gain, regulation of glucose homeostasis and the development of cognitive disorders.

‘While obesity was previously attributed to an increase in the inflammatory state, our study shows that such a state depends on the type of diet to which the animal is exposed. In other words, it is a diet high in omega 6 that is responsible for the inflammatory phenomena observed and not the obesity itself,’ explains Clara Sanchez, Inserm post-doctoral researcher and first author of the article.
‘This study also shows, for the first time, the protective effect against obesity and the associated inflammatory phenomena that a lipid-enriched diet can present, provided it promotes the consumption of omega 3. This research makes it possible to envisage dietary interventions based on a low ω6/ω3 ratio to combat obesity and its associated neurological disorders,’ explains Carole Rovère, Inserm researcher and last author of the article.

In their discovery, the scientists also observed in these mice a change in the shape of certain brain cells located in the hypothalamus – known as microglia – which appear to activate in response to a high-omega 6 diet. Their research will now focus on better understanding the specific role of these cells in obesity.

[1]WHO, 2016

[2]Gregor and Hotamisligil, 2011; Thaler et al., 2012

[3] Baufeld et al., 2016; Cansell et al., 2021; De Souza et al., 2005; Le Thuc et Rovère, 2016; Salvi et al., 2022

[4] The WHO recommends consuming a ratio of omega 6 to omega 3 of 5:1. However, in Western societies our actual consumption is more like 15:1!

[5]Glucose homeostasis is a state of balance between the intake of glucose (intestinal absorption following a meal or production of glucose by the liver) and its use (glucose entry and use in the organs).

Inflammation and cancer: identifying the role of copper paves the way for new therapeutic applications

Posted on April 26, 2023April 28, 2023 by Graziella Cara

The research team developed a “drug prototype” capable of mitigating both the mechanisms of inflammation and the processes potentially involved in metastatic spread. © Institut Curie / BELONCLE Frank

For the first time, researchers from Institut Curie, the CNRS and Inserm have uncovered a previously unknown chain of biochemical reactions. This chain involves copper and leads to metabolic and epigenetic alterations[1] that activate inflammation and tumorigenesis. But there is more; the research team developed a “drug prototype” capable of mitigating both the mechanisms of inflammation and the processes potentially involved in metastatic spread. Published in the journal Nature on April 26, 2023, these results provide hope for new therapeutic opportunities to control inflammation and cancer.

Inflammation is a complex biological process that can eradicate pathogens and promotes repair of damaged tissues. However, deregulation of the immune system can lead to uncontrolled inflammation and produce lesions instead. Inflammation is also involved in cancer. The molecular mechanisms underlying inflammation are not fully understood, and so developing new drugs represents a significant challenge.

As far back as 2020, Dr. Raphaël Rodriguez, CNRS research director and head of the Chemical Biology team at Institut Curie (Equipe Labellisé Ligue Contre le Cancer) at the Cellular and Chemical Biology laboratory (Institut Curie/CNRS/Inserm), had shed new light on a membrane receptor called CD44, which marks immune responses, inflammation and cancer progression. Dr. Rodriquez and his team showed that CD44 helped import iron into cell[2], triggering a series of reactions leading to activation of genes involved in the metastatic process.

“This is a cell plasticity phenomenon we continued to study, investigating other metals potentially internalized by CD44, notably copper,” he explains.

Copper causing epigenetic alterations

Along with his colleagues[3], Dr. Rodriguez has now reached a new milestone.

The research team managed to identify a signaling pathway involving copper and leading to the expression of pro-inflammatory genes in macrophages, the cells present in all tissues and playing an important role in innate immunity.

Once internalized in macrophages, copper enters into the mitochondria (the organelle responsible for cell respiration and energy production), where it catalyzes the oxidation of NADH into NAD⁺ (nicotinamide adenine dinucleotide, a molecule needed for the activity of certain enzymes). The increase of NAD⁺ in cells enables the activity of certain enzymes involved in the production of metabolites essential for epigenetic regulation. These metabolites thus, contribute to the activation of genes involved in inflammation.

Inflammation and cancer: shared molecular mechanisms

The scientists did not stop there, they also designed molecules able to bind to copper, inspired from the structure of metformin.[4] By testing these new molecules on models of acute inflammation, they found that a synthetic dimer of metformin, LCC-12 (also termed Supformin), reduced activation of macrophages and attenuated inflammation.

“Our work has enabled us to develop a drug prototype that inactivates copper chemistry in the cell’s metabolic machinery, thus blocking expression of the genes involved in inflammation”, explains Dr. Rodriguez.

To finish, they applied this therapeutic strategy to cancer cell models engaged in an epithelial-mesenchymal transition[5]. Here again, Supformin blocked the cellular mechanism and thus the cell transformation.

“The genes activated in cancer cells are not the same as those expressed in immune cells, but the chain reaction leading to epigenetic alterations is identical”, explains Dr. Rodriguez.

These results thus reveal the role of copper in cancer cells and their ability to adopt a metastatic nature.

Dr. Raphaël Rodriguez concludes: “Our study reveals that the inflammatory and cancer processes depend on similar molecular mechanisms and could therefore in the future benefit from similar innovative therapies, such as those tested with Supformin.”

The explanations of Dr. Raphaël Rodriguez in video :

[1]Epigenetics is the study of the mechanisms at play in gene regulation, which is essential to the action of cells and to maintaining their identity. Unlike genetic mutations, which are permanent, epigenetic modifications on DNA or histones are reversible.

[2] Read the press release “Cancer: a new mechanism that regulates cell activity involving iron”: https://curie.fr/sites/default/files/medias/documents/2020-08/CPCNRS-CD44ferCancer-FR-emb.pdf

[3] This study was conducted at Institut Curie, in the Cellular and Chemical Biology unit (Institut Curie, CNRS, Inserm), in collaboration with UVSQ, Raymond Poincaré hospital (AP-HP), Gustave Roussy hospital, the Institut de chimie moléculaire et des matériaux d’Orsay (CNRS/University Paris-Saclay), the Multimodal Imaging Center (CNRS/Institut Curie/Inserm/University Paris-Saclay), the Center for Infection and Immunity of Lille (CNRS/Inserm/Institut Pasteur de Lille/CHU of Lille/University of Lille), Institute of Pharmacology and Structural Biology (CNRS/University of Toulouse III) along with British and Australian researchers.

[4]Metformin is a treatment used for Type-2 diabetes, and is able to form a bimolecular complex with copper.

[5] Epithelial-mesenchymal transition is the first step in enabling cancer cells to metastasize.

Extreme Temperatures During Pregnancy: A Possible Impact on the Lung Development of Newborn Girls

Posted on March 17, 2023April 3, 2023 by Graziella Cara

Exposure to extreme temperatures from the fetal stage could impact health. © Fotalia

Exposure to extreme temperatures from the fetal stage could impact health. This is what suggests a study by researchers from Inserm, Université Grenoble Alpes and CNRS, based on the SEPAGES cohort[1], intended to study the impact of various environmental factors on the health of pregnant women and their children. In this research, to be published in JAMA Network open, associations were found in newborn girls, between in utero exposure to very high or very low ambient temperatures from the second trimester of pregnancy and the alteration of several respiratory parameters.

The thermoregulation implemented by the body in response to variations of temperature requires the adaptation of maternal blood flow and cardiac function which, when this occurs during pregnancy, can be to the detriment of the fetus. Physiological alterations have also been observed in animals in response to heat stress exposures, such as impaired placental development with reduced blood flow, or oxidative stress which, outside of normal conditions, may affect the health of mother and child. External temperature could therefore have an impact on embryo-fetal development.

A team led by Inserm researchers Johanna Lepeule and Ariane Guilbert at the Institute for Advanced Biosciences (Inserm/Université Grenoble Alpes/CNRS), wished to verify this hypothesis using data from the SEPAGES cohort (Assessment of Air Pollution Exposure During Pregnancy and Effect on Health). Made up of pregnant women and children from their pregnancies, this cohort makes it possible to study the effect of various environmental factors on health.

Exposure Modeled Throughout Pregnancy

The researchers modeled the exposure to ambient temperatures of 343 women and their children, from conception to their first weeks of life. At the same time, they evaluated the respiratory function of the newborns at around 6 to 7 weeks after birth. Various measurements were used to calculate the tidal volume (volume of air that enters and leaves with each breath), respiratory rate (number of breaths per minute), and functional residual capacity (FRC) (volume of air remaining in the lungs after an expiration)[2].

Since fetal development and respiratory function differ slightly according to sex, the research team also compared outcomes between girls and boys.

Associations That Vary According to Sex

In boys, the scientists did not observe any significant alterations in lung function associated with external temperature during pregnancy. However, they found that girls exposed in utero from the second trimester of pregnancy to the highest or lowest temperatures had a lower FRC and a higher respiratory rate than those exposed to temperatures closer to the average.

In addition, girls exposed to very low temperatures in utero had decreased tidal volume.

“Although the observed variations are not pathological in nature and do not make it possible to predict a future respiratory disorder, explains Lepeule, the various lung function measurements all converge towards an association between in utero exposure to high or low temperatures and poorer lung performance in newborn girls. “

New analyses of the respiratory data collected in children at 3 and 8 years of age will be needed in order to determine whether these associations persist over the long term or whether they are reversible over time.

In the meantime, “these findings underpin the importance of developing public policies to protect pregnant women and their children from extreme temperatures, particularly in the current context of climate change,” concludes Lepeule.

[1] The SEPAGES couple-child cohort (Assessment of Air Pollution Exposure During Pregnancy and Effect on Health), coordinated by Inserm and Université Grenoble Alpes, aims to characterize the exposure of pregnant women and their children to environmental contaminants and study their effect on the health of pregnant women, fetuses, and children.

[2] This residual volume plays an essential role in the maintenance of lung function: as the lungs are elastic, they retract during the muscle relaxation that enables expiration. At the end of expiration, the residual volume makes it possible to limit the retraction forces placed on the lungs so that the pulmonary territories remain open to gas exchange (O₂ and CO₂ essentially). Otherwise, the lungs would close on themselves, and the alveoli would collapse, meaning that gas exchange could no longer take place.

A “Nano-Robot” Built Entirely from DNA to Explore Cell Processes

Posted on July 28, 2022December 2, 2022 by Léa Surugue

Scientists have designed a “nano-robot” made up of three DNA origami structures. © Gaëtan Bellot/Inserm

Constructing a tiny robot from DNA and using it to study cell processes invisible to the naked eye… You would be forgiven for thinking it is science fiction, but it is in fact the subject of serious research by scientists from Inserm, CNRS and Université de Montpellier at the Structural Biology Center in Montpellier[1]. This highly innovative “nano-robot” should enable closer study of the mechanical forces applied at microscopic levels, which are crucial for many biological and pathological processes. It is described in a new study published in Nature Communications.

Our cells are subject to mechanical forces exerted on a microscopic scale, triggering biological signals essential to many cell processes involved in the normal functioning of our body or in the development of diseases.

For example, the feeling of touch is partly conditional on the application of mechanical forces on specific cell receptors (the discovery of which was this year rewarded by the Nobel Prize in Physiology or Medicine).

In addition to touch, these receptors that are sensitive to mechanical forces (known as mechanoreceptors) enable the regulation of other key biological processes such as blood vessel constriction, pain perception, breathing or even the detection of sound waves in the ear, etc.

The dysfunction of this cellular mechanosensitivity is involved in many diseases – for example, cancer: cancer cells migrate within the body by sounding and constantly adapting to the mechanical properties of their microenvironment. Such adaptation is only possible because specific forces are detected by mechanoreceptors that transmit the information to the cell cytoskeleton.

At present, our knowledge of these molecular mechanisms involved in cell mechanosensitivity is still very limited. Several technologies are already available to apply controlled forces and study these mechanisms, but they have a number of limitations. In particular, they are very costly and do not allow us to study several cell receptors at a time, which makes their use very time-consuming if we want to collect a lot of data.

DNA origami structures

In order to propose an alternative, the research team led by Inserm researcher Gaëtan Bellot at the Structural Biology Center (Inserm/CNRS/Université de Montpellier) decided to use the DNA origami method. This enables the self-assembly of 3D nanostructures in a pre-defined form using the DNA molecule as construction material. Over the last ten years, the technique has allowed major advances in the field of nanotechnology.

This enabled the researchers to design a “nano-robot” composed of three DNA origami structures. Of nanometric size, it is therefore compatible with the size of a human cell. It makes it possible for the first time to apply and control a force with a resolution of 1 piconewton, namely one trillionth of a Newton – with 1 Newton corresponding to the force of a finger clicking on a pen. This is the first time that a human-made, self-assembled DNA-based object can apply force with this accuracy.

The team began by coupling the robot with a molecule that recognizes a mechanoreceptor. This made it possible to direct the robot to some of our cells and specifically apply forces to targeted mechanoreceptors localized on the surface of the cells in order to activate them.

Such a tool is very valuable for basic research, as it could be used to better understand the molecular mechanisms involved in cell mechanosensitivity and discover new cell receptors sensitive to mechanical forces. Thanks to the robot, the scientists will also be able to study more precisely at what moment, when applying force, key signaling pathways for many biological and pathological processes are activated at cell level.

“The design of a robot enabling the in vitro and in vivo application of piconewton forces meets a growing demand in the scientific community and represents a major technological advance. However, the biocompatibility of the robot can be considered both an advantage for in vivo applications but may also represent a weakness with sensitivity to enzymes that can degrade DNA. So our next step will be to study how we can modify the surface of the robot so that it is less sensitive to the action of enzymes. We will also try to find other modes of activation of our robot using, for example, a magnetic field,” emphasizes Bellot.

[1] Also contributed to this research: the Institute of Functional Genomics (CNRS/Inserm/Université de Montpellier), the Max Mousseron Biomolecules Institute (CNRS/Université de Montpellier/ENSCM), the Paul Pascal Research Center (CNRS/Université de Bordeaux) and the Physiology and Experimental Medicine: Heart-Muscles laboratory (CNRS/Inserm/Université de Montpellier).

A Gene Therapy Studied in Steinert’s Disease

Posted on February 10, 2022November 23, 2022 by Léa Surugue

Myotonic dystrophy type 1 (DM1) or Steinert’s disease is a rare and debilitating genetic neuromuscular disease affecting multiple organs and with a fatal outcome. No treatment is available at present. Encouraged by previous research into its molecular causes, researchers from Inserm, CNRS, Sorbonne Université, Lille University Hospital and Université de Lille, in partnership with the Institute of Myology, at the Center for Research in Myology and the Lille Neuroscience & Cognition center, have developed and tested a promising gene therapy that acts directly at the origin of the disease. Initial findings published in Nature Biomedical Engineering show correction of molecular and physiological alterations in mouse skeletal muscle1.

Myotonic dystrophy type 1 (DM1), otherwise known as Steinert’s disease, is a rare, hereditary and genetic neuromuscular condition affecting around 1 in 8,000 people. Debilitating and fatal, it is referred to as a “multisystem” condition because it simultaneously affects the muscles (muscle weakening and atrophy called “dystrophy”; muscle relaxation impairment called “myotonia”) and other organs (cardiorespiratory, digestive and nervous systems, etc.). The expression and course of the disease vary from one patient to the next and no treatment exists as yet.

It is caused by the abnormal repetition of a small DNA sequence (triplet CTG2) in the DMPK (DM1 Protein Kinase) gene located on chromosome 19. In healthy individuals, this sequence is present but repeated between 5 and 37 times. However, in patients with DM1, a mutation occurs whereby the number of triplets increases, producing up to several thousand repetitions.

About the mechanisms enabling gene expression

To obtain the production of a protein, a gene (located in the cell nucleus) is first transcribed into a molecule of RNA. To become a messenger RNA (mRNA), it undergoes maturation, particularly involving splicing. This basically means that the molecule is cut into pieces, some of which are eliminated and others attached. Thanks to this finely regulated process, one gene can lead to the synthesis of different mRNA and therefore of different proteins. After splicing, the mature mRNA will eventually be translated into protein, outside of the cell nucleus.

In Steinert’s disease, the mutated gene is transcribed but the mutant mRNA are retained in the cell nuclei as characteristic aggregates. In the cells of people with DM1, the MBNL1 proteins that normally bind to certain RNA in order to regulate their splicing and maturation are “captured” by the RNA that carry the mutation.

Thus sequestered in the aggregates, it is impossible for them to perform their functions, resulting in the production of proteins that function less well or not at all, some of which have been linked to clinical symptoms.

The team led by CNRS Research Director Denis Furling at the Myology Research Center (Inserm/Sorbonne Université/Institute of Myology) in association with that of Nicolas Sergeant, Inserm Research Director at the Lille Neuroscience & Cognition Centre (Inserm/Université de Lille/Lille University Hospital), focused on a therapeutic strategy to restore the initial activity of MBNL1 in skeletal muscle cells expressing the mutation responsible for Steinert’s disease.

To do this, the scientists engineered modified proteins that present, like the protein MBNL1, binding affinities for the RNA carrying the mutation and as a consequence act as a decoy for these RNA.

When expressing these decoy proteins in vitro in muscle cells from patients with DM1, they observed that they were captured by the mutated RNA instead of the MBNL1 proteins. The latter were then released from the aggregates of mutated RNA and regained their normal function. As a result, the splicing errors initially present in these cells disappeared. Finally, the mutated RNA bound to the decoy proteins proved to be less stable and could be more easily and effectively eliminated by the cell.

Aggregates of mutant DMPK-RNA containing pathological triplet (red) repetitions visualized by FISH-immunofluorescence in the nuclei (blue) of muscle cells (green) isolated from patients with Myotonic dystrophy type1 © Denis Furling and Nicolas Sergeant

The research team then transposed this technique into an animal model in order to verify the validity of this approach in vivo. With the help of viral vectors used in gene therapy, the decoy proteins were expressed in the skeletal muscle of mouse models of Steinert’s disease. In these mice, a single injection was effective, over a long period of time and with few side effects, in correcting the muscle damage associated with the disease, particularly the splicing errors, myopathy and myotonia.

“Our findings highlight the efficacy on Steinert’s disease symptoms of a gene therapy based on the bioengineering of RNA-binding decoy proteins with strong affinity for the pathological repetitions present in the mutated RNA, in order to release the MBNL1 proteins and restore their regulatory functions,” declares Furling. However, the authors point out that additional studies are needed before this therapy can be transposed into a clinical study. “This research paves the way for the development of therapeutic solutions in the context of other diseases in which pathological RNA repetitions cause splicing regulation dysfunction,” concludes Sergeant.

1 The striated skeletal muscle is the muscle that is attached to the skeleton by tendons and which, due to its ability to contract, enables the performance of precise movements in a well-defined direction.

2 The coding sequence of a gene consists of the chaining of different combinations of four nucleic acids: adenine, guanine, cytosine, and thymine (replaced by uracil in RNA). These are organized in triplets (or codons) whose correct “reading” by the cell machinery enables the expression of a protein.

Human “Jumping Genes” Caught in the Act!

Posted on May 2, 2019November 23, 2022 by Graziella Cara

Over the course of evolution, the genomes of most living organisms have grown more complex thanks to transposable elements, a.k.a. “jumping genes,” or DNA fragments that can move and copy themselves from one chromosome location to another. Researchers from Inserm, the CNRS, Université Côte d’Azur, and Université de Montpellier were able to capture these “jumping genes” just after they moved. The researchers compared their observations with existing databases. Their work, to be published in Molecular Cell, shows that the integration of “jumping genes” in humans is not random. Instead, it is thought to be influenced by specific genome properties. These results open up new perspectives for interpreting whole genome sequencing data.

Transposable elements, also known as “jumping genes,” are small DNA fragments that can multiply and move in the chromosomes of most living organisms. They have proliferated so intensely in mammals and primates that they make up more than half of our chromosomes! Of course, they don’t jump all at once in all of our cells. Of all the copies present in our DNA, only a small fraction remain active. All the rest are molecular remnants reflecting millions of years of evolution, during which harmful insertions were eliminated and beneficial ones retained.

In humans, the most active jumping genes are L1 retrotransposons. They can alter or destroy genes when they jump, triggering the manifestation of genetic diseases like hemophilia and muscular dystrophy. L1 retrotransposons are also particularly active in some forms of cancer, and could be involved in cellular aging or in some mental illnesses.

Do L1 retrotransposons target specific chromosome regions, or do they choose their positions at random? Teams led by Inserm head researchers Gaël Cristofari and Simona Saccani working at the Nice Institute for Research on Cancer and Aging (IRCAN, Inserm, CNRS, Université Côte d’Azur), along with their colleagues at Université de Montpellier, were able to use a “high-speed” genome sequencing technique to catch actively jumping genes right after they jumped to a new position.

After comparing their observations with genomic and epigenomic databanks, the researchers were able to identify which genome characteristics influenced the integration of the L1 retrotransposons. The most notable characteristic was DNA replication, and natural selection phenomena after integration played a preponderant role.

“We already knew that L1 retrotransposons tend to accumulate in specific areas of our chromosomes, especially heterochromatin. But we didn’t know whether that reflected a particular attraction to those regions, or if they are simply tolerated in those regions and eliminated elsewhere through natural selection. When we know where they jump to and which copies are retained over the course of evolution, we can discover – by deduction – the regions where they can do damage,” explains Cristofari.

Their results make it easier to understand how jumping genes can trigger mutations in humans, and how they contribute to the evolution of our genetic heritage. In the future, this research could be used to interpret whole genome sequencing data, particularly in personalized medicine and vast sequencing programs.

The research was made possible with financial support from the Fondation pour la Recherche Médicale, Cancéropôle PACA, the European Research Council, the French National Research Agency, the Labex Signalife, the Groupement de Recherche sur les Eléments Transposables (CNRS, GDR 3546), the FHU OncoAge, and the European Erasmus Mundus Mobility with Asia program.

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Category: Bases moléculaires et structurales du vivant