1. Introduction

The laws of genetics were outlined by the monk Gregor Mendel as early as 1856. In the garden of the Abbey of St Thomas in Brno, in the south-east of what is now the Czech Republic, he turned to the study of the hybridisation of peas, because the Bishop, his hierarchical superior, asked him to stop being interested in the hybridisation of mice, the breeding he carried out in his cell being a nuisance to his fellow monks. In 1866, after 10 years of meticulous work, Mendel set out, in a very interesting article entitled “ Experiments on plant hybrids ”, the theoretical foundations of the laws of genetics and modern heredity. Often presented as early as the 19th century, Mendel’s experiments have frequently been poorly taken up and misinterpreted. Mendel’s fundamental contribution, far more than the discovery of these laws, lies in the assertion that it is not the traits themselves that are transmitted, but something else that Mendel designates by the term “ Faktoren ” (factors, which the Danish geneticist Wilhelm Johannsen would call genes, in 1909).

For a long time, scientists believed that an organism’s genetic code alone determined its biological characteristics, its development and its adaptation to its environment.

The discovery of DNA in 1953 by the Briton Francis Crick and the American James Watson was a tremendous advance in the understanding of genetics. Thanks to technological advances in biology and with the help of highly powerful computers, scientists managed to map the genomes of many organisms, including the bee. They then realised that there are significant differences between what an organism’s genetic heritage dictates and the appearance or functioning of that same organism.

Since ~2010, epigenetics has been investigating the mechanisms regulating genome expression without modification/alteration of the genetic code itself.

This branch of biology studies, in a sense, the role of the conductor who interprets the score of a concertante symphony by directing the musicians of his orchestra with a baton…

Under the influence of various chemical or “external” factors, epigenetics can not only modify an organism’s appearance or behaviour, but can also transmit some of these modifications to subsequent generations…

Beekeepers are well aware of the mechanism underlying the differentiation of worker/queen castes. A fertilised egg laid by a queen will give rise either to a worker or to a queen, depending on the available diet (pure royal jelly or not). It is also known that a worker begins her life as a nurse and ends it as a forager. These two activities are completely different and involve clearly distinct skills. Beekeepers are nonetheless struck by the epigenetic modifications brought about by the resumption of the queen’s egg-laying in spring or by queenlessness. The reversibility of these epigenetic modulations fascinates us: a forager can become a nurse again if needed; a worker can even start laying eggs!

 

1.1 A few definitions to get started

Heritage: property that one holds by inheritance from one’s ancestors.

Genetics: the science that studies individuals’ hereditary traits, their transmission across generations and their variations (mutations).

Epigenetics: a branch of biology that studies the nature of mechanisms that modify gene expression in a reversible, transmissible and adaptive way without changing the nucleotide sequence.

Chromosome: a complex molecular structure composed of DNA and proteins, whose configuration evokes the letter “ X – or Y ”.

DNA: a helical molecule (twisted double helix) carrying the entirety of the genetic information (genome) contained in the nucleus of a cell of a living organism (animal or plant).

RNA: a molecule consisting of a chain of ribonucleotides (twisted single helix) whose order is dictated by DNA sequences, which it copies as a template.

Enzyme: a protein capable of catalysing chemical reactions in cells

Nucleic acid: an assembly of macromolecules or a polymer whose basic unit, or monomer, is a nucleotide and whose nucleotides are linked to one another by phosphodiester bonds.

Nucleoside: a molecule resulting from the linkage of a nucleic base to a sugar.

Nucleotide: an organic molecule composed of a nucleic base, a five-carbon sugar and finally one to three phosphate groups.

 

A cell consists of a plasma membrane or envelope containing cytoplasm, which is formed by an aqueous solution (cytosol) in which there are many biomolecules such as proteins and nucleic acids, organised or not within organelles. Many living beings consist of only a single cell: these are unicellular organisms such as bacteria, archaea and most protists.

Others consist of several cells: these are multicellular organisms such as plants and animals. The latter contain a highly variable number of cells from one species to another; the human body contains about one hundred trillion (10¹⁴) cells, but is colonised by a two- to tenfold greater number of bacteria, which are part of its microbiota and are much smaller than human cells. Most animal cells are visible only under a microscope, with a diameter between 10 and 100 µm.

The nucleus is a cellular structure present in the majority of eukaryotic cells and in all eukaryotic organisms, containing most of the cell’s genetic material (DNA). Its main function is to store and protect the nuclear genome as well as the machinery necessary for chromosome replication and for the expression of the information contained in genes. It disappears temporarily during the process of cell division and is reconstituted in daughter cells. Its diameter varies from 5 to 7 micrometres.

Ribosomes are ribonucleoprotein complexes (i.e. composed of proteins and RNA) present in eukaryotic and prokaryotic cells. Extremely well conserved over the course of evolution, their function is to synthesise proteins, like true production plants, by decoding the information contained in messenger RNA. Ribosomes are made up of ribosomal RNA, which carries the catalytic activity, and ribosomal proteins. Ribosomes consist of two subunits: a smaller one that “reads” the messenger RNA and a larger one that carries out the polymerisation of amino acids to form the corresponding protein.

Mitochondria are often described as the “power stations” of cells, insofar as they contribute to most cellular ATP production via β-oxidation, the Krebs cycle and the respiratory chain as part of oxidative phosphorylation, ATP being the ubiquitous energy molecule used in a very large number of chemical reactions of metabolism, notably anabolism (biosyntheses).

The Golgi apparatus is a cellular organelle located near the endoplasmic reticulum and the nucleus. It stores the proteins and lipids produced by the reticulum, modifies them through the action of enzymes, sorts them and transports them within the intra- or extracellular environment, depending on their final destination.

The smooth endoplasmic reticulum, devoid of ribosomes, is the site of lipid synthesis and sugar metabolism. Containing specialised enzymes, the reticulum also participates in cell detoxification.

The rough endoplasmic reticulum assembles and transports proteins destined for membranes and for cellular secretion.

 

1.2 Some astonishing figures

 

The length of the genome (fully extended) of a human cell (Ø ~5–50 µm) reaches 2 metres for a diameter of 2 µm (µ =10-6) and comprises more than 3 billion bases coding for more than 23’000 different proteins.

Human chromosome no. 1, the largest of the human chromosomes, contains about 220 million base pairs for a linear length of more than 7 cm.

Neither the number of chromosomes, nor genome size (the amount of DNA contained in a copy of a genome), nor the number of genes is correlated with the complexity of the organism: this is the C-value paradox. The C-value represents genome size, expressed as the number of base pairs, or in picograms.

 

2. Each human cell contains 46 chromosomes

Each human cell contains 46 chromosomes (like the olive tree, the gecko, the sea sponge, the bat) versus 32 in the bee’s cell and 16 in the drone, which is haploid. The Australian ant has 2, the mosquito has 6, the elephant 56, the goldfish 100, the Azuré de l’Atlas butterfly 440, the proteus amoeba around 1000!!!

Chromosome telomeres are protected by “caps” to slow their ageing and prevent fraying that could lead to fusions between chromosomes and thus mutations.

In humans, telomeres shorten with age, inflammation and stress. Studies have shown that short telomeres are associated with a higher risk of age-related diseases (cancerous tumours).

One study1 is said to have shown that beekeepers have a longer lifespan than non-beekeepers. It turns out that the telomere length of the tested beekeepers is significantly longer than that of non-beekeepers.

Cellular oxidation phenomena and excessive production of free radicals contribute to premature telomere shortening. This oxidation is amplified by certain factors such as tobacco, alcohol, regular medication intake, pollution, UV, electromagnetic radiation, physical and intellectual overwork, stress, malnutrition, etc. The consumption of antioxidants would slow this early wear. As part of a health-oriented diet, consumption of hive products is an asset. The antioxidant properties of honey, pollen and royal jelly have been confirmed by several recent studies highlighting the virtues of polyphenols in a preventive context.

In the bee, telomerase, the enzyme that maintains telomere length, could be modulated by epigenetics, which would explain the difference in lifespan between summer bees and winter bees, and in queens.

1 ) Nasir, N. F. M., Kannan, T. P., Sulaiman, S. A., Shamsuddin, S., Azlina, A., & Stangaciu, S. (2015). The relationship between telomere length and beekeeping among Malaysians. AGE, 37(3), 1-6

 

One may try to imagine DNA as a ladder. The uprights of the “ladder” correspond to sugars and the rungs to nitrogenous bases A-T-C-G. One more small visualisation effort is then needed to twist this ladder around itself and make it into a twisted double helix.

		Each human cell contains 46 chromosomes (like the olive tree, the gecko, the sea sponge, the bat) versus 32 in the bee’s cell and 16 in the drone, which is haploid. The Australian ant has 2, the mosquito has 6, the elephant 56, the goldfish 100, the Azuré de l’Atlas butterfly 440, the proteus amoeba around 1000!!!
		The rungs of the ladder can dissociate (open like a zip/zip fastener), then be read, copied and exported to the production plants (ribosomes) to synthesise all the proteins necessary for the life of the cell. It is the sequences of these rungs, taken three by three (triplets or codons), that encode the manufacture of proteins.

The structure of the horizontal rungs of the ladder is formed by four distinct and paired nitrogenous bases (rather simple molecules, carbon- and nitrogen-based, very common and involved in many biochemical processes):

A = Adenine with T = Thymine ; C = Cytosine with G = Guanine

The sequence of these base pairs (genetic code) corresponds to the manufacturing plan for each of the proteins our cells may need in order to exist, function... and keep us alive!

These plans—our genes—are therefore kept safely inside the cell nucleus in the form of DNA. The factories that synthesise proteins—ribosomes—are located outside this nucleus.

 

Thus, protein manufacture is not carried out from the original plans, but by relying on their “duplicate”: messenger RNA (for messenger ribonucleic acid).

Thus, when a cell needs a protein, the manufacturing plan for that protein is “photocopied” or “transcribed”. The copy thus generated—a messenger RNA—is then exported out of the nucleus and reaches the ribosomes where it enables synthesis of the required protein. Very unstable and fragile, this copy is then rapidly destroyed.

If DNA is the medium of heredity in living organisms, the gene is a segment of DNA that contains the information necessary for the production of a functional unit (for example a protein). It is the functional unit of heredity.

3. Transcription and translation

DNA stores genetic information in the long term.

By replication, during cell division (mitosis), it transmits all the genetic information to new somatic cells (and only half the genome, by meiosis, to reproductive cells or gametes, also called egg or sperm cells).

By transcription, DNA is “ photocopied ” and “ printed ” into a single-stranded RNA chain (one upright of the ladder as opposed to the two uprights of DNA). This “ photocopy ”, called messenger RNA (everyone has heard of it in connection with mRNA vaccines against COVID-19), is exported out of the nucleus and directed towards adjacent ribosomes for protein assembly (translation).

 

3.1 Figuratively, to see more clearly the relationship between genetics and epigenetics

Let us use as an analogy a user manual describing how to assemble a car on an assembly line.

If your responsibility is to fit the wheels, you do not need information about installing the windscreen, assembling the seats, or assembling the engine. To make your task easier, the manual is divided into parts, chapters, sections and paragraphs so that you can focus only on the information required to fit the wheels. The user manual is analogous to your genome—it is composed of all the information you need to form a human. The layout and organisation of the manual reflect the function of the epigenome, that is, the chemical biology that indicates to different cell types which portions of the genome are to be read. In this way, by working on the same information (your genome), your cells can organise this information (using epigenetics) and work in concert, each with its own role to form and maintain all the tissues and organs of your body.

In its most scientifically rigorous definition, a chromosome corresponds to a fully condensed chromatin structure (DNA thread), with a fibrous appearance like a tightly wound ball of wool.

In this definition, the chromosome is present only during mitosis (cell division), during which it reaches its maximum degree of condensation.

The rest of the time (outside mitosis), chromatin is more or less condensed in the nucleus and does not form a chromosome.

 

This DNA thread is twisted and wound around itself and around “ pulleys ”, or histones.

If the “pulleys” are pressed tightly against one another by methylation, the DNA strand is not accessible for reading by RNA to produce proteins. Conversely, if the DNA strand is largely unwound by acetylation or epigenetic factors, the reading of the different genes becomes possible and protein production starts.

4. Epigenetic mechanisms

Top: the addition of methyl groups to DNA or histones compacts DNA winding. Transcription factors cannot bind to DNA and genes are not expressed. Bottom: histone acetylation allows DNA to unwind. Transcription factors can bind to DNA and genes are expressed.

To simplify:

When DNA genes coding, for example, for pollen collection cannot be read, the proteins/enzymes are not produced and the worker carries out her tasks inside the hive.

Conversely, when the segment of DNA that codes to boost a forager is well exposed and readable, the machinery kicks in, changes the behaviour of the nurse, who becomes a forager, and pollen collection is in full swing…     Figuratively, one can represent the genome (the DNA thread) as a magnetic tape on which the entire “Beethoven’s Symphony No. 9” or the entire genetic code of the cell has been recorded.

Epigenetics makes it possible to mask or unmask certain bars of the score and thus to modulate the interpretation of the piece of music.

 

5. Genetics vs epigenetics

From the point of view of genetics, a gene can be modified by an alteration of the genetic code, for example after a genetic mutation due to a mutagen (UV, X-rays…) or spontaneously, following an error in DNA replication during cell division. This modification of the genetic heritage is definitive and irreversible. It is transmitted to descendants.

From the point of view of epigenetics, there is no modification of the genetic code ; a gene can be masked or temporarily overexpressed by the induction of external factors (environment, others…). This modification is totally reversible and can be transmitted to descendants.

The plasticity of the worker’s epigenome is markedly greater than that of queens: workers alternate several roles/trades over the course of their short life: cleaners, nurses, ladies-in-waiting, wax producers, ventilators, storekeepers, guards, scouts, foragers;  for its part, the queen does nothing but eat and lay eggs... J !

The sequence of these tasks is easily modulated, so that a forager can become a nurse again if needed (!), for example when the queen resumes egg-laying after the winter months. Likewise, a nurse can quickly become a forager (!) after the division of a colony (the foragers from both hives returning to the parent hive…). The astonishing complexity of epigenetics arises when swarming fever ignites and several factors coexist (lack of space, area of open/capped brood, weather, season, age of the queen, deficit of marking pheromones, etc…)

► see the article: Reversible switching between epigenetic states in the behavioural suborganisms of bees

 

 

Examples of external or epigenetic factors :

• Royal jelly
• Pheromones
• Juvenile hormone
• Vitellogenin
• Ethyl oleate
• Food abundance
• Temperature
• Stress
• Pesticides
• Toxins, infections
• Parasites
• Age polyethism
• Demography of sub-castes…

 

Royal jelly is a secretion product of the cephalic glandular system (hypopharyngeal glands and mandibular glands) of worker bees, between the fifth and fourteenth day of life. This whitish, gelatinous substance is rich in fatty acid (10HDA) stimulating the development of the reproductive system, in several vitamins and trace elements, and in royalactin (57-kDa protein) which determines worker/queen caste differentiation.

Juvenile hormone is a hormone that controls post-embryonic development in insects. It owes its name to the fact that it maintains juvenile characteristics, by promoting larval moults and delaying metamorphosis. However, it does not act only in larvae, since it also plays a very important role in the adult insect, where it regulates reproduction, in particular vitellogenesis and oogenesis.

Vitellogenin is a protein highly present in winter bees; it is stored in their overdeveloped fat body; this molecule stimulates their immune system. Moreover, there is a correlation between vitellogenin level and the chances of winter survival of bee colonies. The vitellogenin level also determines the type of foraging. Thus, high vitellogenin levels in young bees will favour late foraging oriented towards pollen collection, and lower levels will allow earlier foraging oriented towards nectar collection. Moreover, vitellogenin reduces oxidative stress in the bee by trapping free radicals, thereby extending the lifespan of workers and the queen.

Ethyl oleate (EO), a true pheromone produced by foragers, is a molecule that plays an essential role in the maturation and transformation of the youngest bees: it acts as a chemical inhibitor that delays the age of foraging. This pheromone thus inhibits the transformation of young bees into foragers as follows:

• In the case of a strong nectar flow and fine weather, foragers are outside “at work” and the young bees remaining in the hive are therefore not exposed to ethyl oleate. They then transform more quickly into foragers, because a colony knows how to mobilise its forces to take advantage of a strong nectar flow. This results in an understaffing of nurses in the hive and encourages the queen to increase egg-laying.
• In the case of bad weather, by contrast, foragers are confined in the hive and diffuse ethyl oleate to young bees, which then remain longer at the nurse stage. There is then a large population confined in the hive, with a very high number of young nurses. This imbalance in bee castes very often triggers swarming fever and, very frequently, as soon as fine weather returns, swarming occurs.

Interfering RNA is a ribonucleic acid whose interference with a specific messenger RNA leads to its degradation and a decrease in its translation into protein. Insofar as RNA plays a crucial role in gene expression, interfering RNA makes it possible to block it by rendering a given gene “silent”. It would presumably be a product of evolution enabling organisms to defend themselves against the introduction of foreign genomes, notably viral, or to modulate gene expression.

 

Although not experimentally verified, it can be observed that when larvae are taken for grafting from a colony acclimatised at altitude (around 1’500 m), queens moved to the lowlands (at 500 m) will show a delay in their egg-laying process the following spring (up to 2 brood frames difference on the same date compared with other queens from the lowlands).

One may hypothesise that information in the form of an epigenetic mark present in the larva could condition its future as a queen (in this case, delayed colony development); this mark being potentially labile under the effect of the new environment, the trait is no longer observed in the second year. The effect of the geographical environment could have consequences for the performance of a queen purchased from a breeder located in a different environment.

Consequently, it could be risky to judge a queen’s performance without taking into account the geographical origin of her mother. Following this line of reasoning, one might think that ecotypes adapted to different regions could correspond to colonies that are genetically very close but differentially marked by their natural habitat. These forms of adaptation, drawing on epigenetic phenomena, would therefore be potentially reversible.

6. In conclusion

Epigenetic mechanisms confer plasticity on living beings with respect to environmental changes and have certainly contributed to the evolution of species over time. In the bee, these phenomena also exist and play a major role in behavioural changes. It is therefore also possible that the traits observed in bee colonies are not solely linked to the presence of certain alleles but also to epigenetic imprints positioned on the parents’ genome; in this case, the traits observed by the beekeeper at colony level would be potentially reversible.

7. Annexes

 

		Universal genetic code : by reading the letters from the centre to the periphery of the disc, one obtains a triplet that provides ribosomes with the information to synthesise a very specific amino acid. For example, “ AGG ” yields arginine. The apposition of several amino acids will form a protein. Reading a DNA gene always begins with a start code (AUG) and always ends with one of the three stops (UAA, UAG, UGA), a bit like in Morse code writing (… 3 dots between letters ; ……. 7 dots between words).
“ True ” twins: identical in terms of their genome (genetic heritage) but different in terms of their phenotype (appearance).		Epigenetic marks vary from one individual to another. Even monozygotic twin sisters (from the same egg) are not identical from this point of view. Monozygotic twins are often very physically similar. As they age, monozygotic twins differentiate following personal choices such as diet, physical and intellectual activities... as well as life experiences. Schooling also constitutes an important step in this differentiation process.

In scientific research, the comparative study of two monozygotic twin individuals makes it possible to highlight epigenetic mechanisms that play an important role in the regulation of gene expression during ontogenesis. Thus, although they are clones in the biological sense of the term, they have different fingerprints and biological constants from the first weeks of gestation. Over the course of their lives, modulation of the expression of their genes is influenced by the environment in the broad sense (lifestyle hygiene, diet, profession, geographical location, etc.), and their phenotype makes it easier to differentiate them (cf. the Bogdanoff brothers).

 

 

 

Sources:

 https://www.docteur-eric-sebban.fr/cancer-du-sein/diagnostic-cancer-sein/implication-des-modifications-epigenetiques-sur-le-cancer-du-sein/

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 https://lejournal.cnrs.fr/articles/lepigenetique-mene-le-genome-a-la-baguette

 https://thisisepigenetics.ca/fr/blogs/quest-ce-que-lepigenetique

 https://lejournal.cnrs.fr/articles/edith-heard-ou-la-revolution-epigenetique

 https://aide-a-la-procreation.fr/au-sujet-de-la-fertilite/epigenetique/

 https://fr.wikipedia.org/wiki/Cellule_(biologie)

 https://thisisepigenetics.ca/fr/blogs/quest-ce-que-lepigenetique

 http://www.astrosurf.com/luxorion/Bio/cellule-3d-dwg-raven.jpg

 https://www.dropbox.com/s/vbvr8fxkg6cofxs/1461823611-Epigenetique.pdf?dl=0