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In a study published in the International Journal of Molecular Sciences, researchers Dr. Ádám Sturm and Dr. Tibor Vellai from Eötvös Loránd University have made a significant leap forward in our understanding of the aging process. Their latest research reveals a hidden epigenetic mechanism in mitochondrial DNA (mtDNA) that could revolutionize how we approach aging research and diagnostics.

This new discovery builds upon the researchers’ previous landmark studies. In 2015, they published “The mechanism of aging: primary role of transposable elements in genome disintegration,” which established the crucial role of transposable elements in the aging process. They followed this up in 2023 with “Downregulation of transposable elements extends lifespan in Caenorhabditis elegans,” further solidifying the connection between transposable elements and longevity.

The Mitochondrial Epigenetic Clock

The current study unveils a previously hidden DNA modification called N6-methyladenine (6mA) that accumulates progressively in mtDNA as organisms age. This phenomenon was observed across diverse species, including the nematode Caenorhabditis elegans, the fruit fly Drosophila melanogaster, and dogs. The consistency across these different species suggests an evolutionarily conserved mechanism in the aging process that could potentially apply to all animal species, including humans.

Dr. Sturm describes this discovery as a “mitochondrial epigenetic clock.” This clock ticks at different rates depending on the lifespan of the organism, providing a new perspective on how aging is regulated at the cellular level. The implications of this discovery are far-reaching, offering new insights into the molecular mechanisms of aging and potential avenues for intervention.

Overcoming Methodological Challenges

One of the key achievements of this study was the development of a novel, reliable PCR-based method for detecting 6mA modifications. This technique allows for accurate, sequence-specific measurement of 6mA levels in mtDNA, overcoming limitations of previous methods that had led to disputes about the existence of 6mA in animal genomes.

The researchers’ method relies on PCR-directed amplification of target sequences digested by a 6mA-dependent restriction enzyme and ligated to a linker stretch. This approach eliminates artifacts due to detection noise or bacterial contamination, providing unambiguous identification of 6mA sites in mitochondrial (and potentially nuclear) genomes.

Figure1. The new PCR based 6ma quantification method. Detailed description here. 

Linking 6mA Accumulation to Lifespan

A particularly intriguing finding of the study was that long-lived C. elegans mutants, which live twice as long as wild-type worms, accumulate 6mA at half the rate of their normal counterparts. This observation strongly links the rate of 6mA accumulation to the aging process and lifespan regulation. The researchers propose that N6-adenine methylation may serve as a mitochondrion-specific epigenetic clock. The gradual increase in mtDNA 6mA levels during aging is so robust and evolutionarily conserved that it can serve as a reliable epigenetic mark to determine age from a given tissue sample.

Single-Molecule Real-Time (SMRT) Sequencing Analysis

The researchers performed SMRT sequencing analysis on mtDNA samples isolated from C. elegans at different adult stages, specifically days 1 and 5. This technique allows for the direct detection of DNA modifications, including N6-methyladenine, during the sequencing process.
The results of the SMRT sequencing were striking. By identifying N6-methylated adenine sites and comparing the total 6mA levels of each chromosome with those of the mitochondrial genome within each age group, the team observed that N6-adenine methylation in the mtDNA relative to the nuclear genome was more robust in the aged sample.

Figure 2. SMRTseq analyses confirm that N6-methyadenine levels in the C. elegans mtDNA increase with age.

Implications for Aging Research and Diagnostics

The discovery of this mitochondrial epigenetic clock opens up new avenues for understanding and potentially intervening in the aging process. Dr. Vellai emphasizes that this epigenetic clock in mtDNA could serve as a more accessible and cost-effective way to measure biological age, compared to existing methods.

Current age-determining biomarkers and methods, such as Horvath’s clock (based on cytosine methylation), transcriptome-based aging clocks, inflammatory aging clocks, and metabolomic aging clocks, while informative, are often expensive and time-consuming. In contrast, the mtDNA 6mA-detecting PCR-based method presented in this study is relatively inexpensive and fast to perform, providing an opportunity for widespread use in scientific research, clinical diagnostics, and even forensics.

The Lifespan Limit Line

One of the most intriguing findings of the study is the discovery of what the researchers call a “Lifespan Limit Line.” This represents a point where the epigenetic marker (6mA) in mitochondrial DNA reaches a peak before declining, signaling the rapid onset of the aging process with severe health deteriorations.

This concept of a Lifespan Limit Line could have profound implications for our understanding of the maximum potential lifespan of different species and individuals. It may also provide insights into the biological processes that occur as organisms approach the end of their natural lifespan.

Figure 3. Discovery of a Lifespan Limit Line across species. The figure illustrates a surprising finding of a “Lifespan Limit Line”—a point where an epigenetic marker in mitochondrial DNA reaches a peak before declining, signaling the rapid onset of the aging process with severe health deteriorations.

Future Directions

The study paves the way for future research on how environmental factors, lifestyle choices, and potential interventions might influence the rate of 6mA accumulation in mtDNA. Understanding these epigenetic changes could lead to novel strategies for promoting healthier aging and potentially extending healthspan.

Moreover, the connection between this newly discovered epigenetic clock and the researchers’ previous work on transposable elements opens up intriguing questions about the interplay between different cellular and molecular mechanisms in the aging process.

After years of theoretical work, scientists from Eötvös Loránd University, Dr. Ádám Sturm and Dr. Tibor Vellai, prove their aging theory experimentally in a major breakthrough published in Nature Communications.

“Jumping genes”, also known as transposable elements (TE), account for more than half of our DNA. As we age, these jumping genes become more active because of an epigenetic DNA change called ‘adenine methylation’ or ‘6mA’. When these genes move and insert themselves into new locations within the DNA, they can interfere with the operation of other genes or important regions. This can change how those genes work or even stop them from working altogether, affecting the overall function and health of the cell. Credit: Sturm, Á., et al., 2023

Pioneering Hungarian aging researchers Dr. Ádám Sturm and Dr. Tibor Vellai have taken a monumental leap in the understanding of aging and, potentially, biological immortality. Their latest publication in Nature Communications comes as the culmination of years of theoretical and experimental works, suggesting the pivotal role of transposable elements (TEs) in aging and the Piwi-piRNA pathway inhibiting these elements in the non-aging biological systems.

The Piwi-piRNA pathway, has previously been proposed by the duo as the hidden mechanism behind the non-aging feature of the germline, cancer stem cells, and notably, the enigmatic Turritopsis dohrnii, commonly known as the “immortal jellyfish.”

In previous landmark articles entitled “The mechanism of aging: primary role of transposable elements in genome disintegration” (2015) and “The Piwi-piRNA pathway: road to immortality” (2017), Dr. Sturm and Dr. Vellai theorized the profound relationship between the Piwi-piRNA system and intriguing concept of biological immortality.

“Through years of dedicated research, we’ve long hypothesized the connection between the Piwi-piRNA pathway and non-aging biological systems. Today, we’ve experimentally proven its role in extending lifespan in a multicellular organism,” said Dr. Ádám Sturm commenting on the experiment in which an element of the Piwi-piRNA system from the non-aging germline was inserted and overexpressed in aging somatic cells. It was discovered that this intervention significantly increases the animal’s lifespan.

In C. elegans, somatic expression of prg-1 (Piwi-related gene)/Piwi, which is normally active in the non-aging germline only, significantly extends lifespan. Credit: Sturm, Á., et al., 2023

Dr. Tibor Vellai commented, “Our findings bridge the gap between theory and empirical evidence, ushering in a new era of genetic exploration. By harnessing the power of this pathway, the possibilities for extending life and enhancing healthspan are monumental.”

The new paper entitled “Downregulation of transposable elements extends lifespan in Caenorhabditis elegans” focuses on the mobility of TEs and their role in causing genomic instability. The research team used the nematode Caenorhabditis elegans as a model organism to explore this relationship.

“We’ve always been intrigued by the fundamental problem of whether the increasing mobilization of TEs is a cause or a consequence of aging,” said Dr. Sturm. “With this study, we’ve made a pivotal discovery: downregulating active TE families indeed extends lifespan. This indicates that TE mobilization drives the aging process.”

Dr. Vellai added, “It’s a significant step forward. The role of TEs in aging hasn’t been this comprehensively understood before. The results showcase that TEs are a crucial genetic determinant of lifespan.”

For those unfamiliar, TEs are segments of DNA that can change their positions within the genome, possibly leading to mutations that affect the host organism’s health and longevity. The Piwi-piRNA pathway effectively suppresses TEs in the non-aging germline (cells that are set aside during early development and later become eggs or sperm). However, as somatic cells age and lose the pathway’s effects, TEs become gradually mobile, causing genomic instability.

The team not only demonstrated that the expression of TEs is related to aging using a research-specific total RNA sequencing method, but they also extensively employed RNA interference-mediated downregulation to study specific active TE gene families, such as Tc1 and Tc3, in worms. They discovered that silencing TEs slows down the aging process. They also revealed that if several active TE families were silenced at the same time, the effect was additive, thus further proving that extreme longevity can be achieved if all active TEs are silenced simultaneously, as in the case of non-aging biological systems.

Downregulation of active transposable element (TE) families extends lifespan in C. elegans. Inhibition of Tc1 (a), Tc3 (b) and both TE families (c) promotes longevity. Simultaneous downregulation of Tc1 and Tc3 displays an additive effect. Credit: Sturm, Á., et al., 2023

“In our lifespan assays, by merely downregulating TEs or somatically overexpressing the Piwi-piRNA pathway elements, we observed a statistically significant lifespan advantage,” Dr. Vellai explained. “This opens the door to a myriad of potential applications in the world of medicine and biology.”

Furthermore, the research introduced the idea that DNA N6-adenine methylation, which is an epigenetic modification, rises with age at TE stretches specifically. This modification was observed using Single-molecule real-time (SMRT) sequencing and a newly developed PCR-based method for detecting N6-adenine methylation, which indicated an increase in their transcription as the animal aged.

Dr. Sturm emphasized the potential implications of this discovery. “This epigenetic modification may pave the way for a method to determine age from DNA, providing an accurate biological clock.”

The study provides not just a glimpse into the intricate mechanisms of aging but also offers avenues for further research into how controlling TE activity might affect overall health and longevity.

Both Dr. Sturm and Dr. Vellai urged researchers worldwide to delve deeper into this discovery, believing it to be a starting point for further revelations about aging. “This is just the beginning,” said Dr. Vellai, “the path to understanding and, perhaps, controlling aging and all diseases of old age is clearer than ever.”

 

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More information: Sturm, Á., et al. Downregulation of transposable elements extends lifespan in Caenorhabditis elegans. Nat Commun 14, 5278 (2023). https://doi.org/10.1038/s41467-023-40957-9

At Deep Biotech Solutions, we’re invested in advancing the field of genomic research. One such area of interest is the sequencing of the model organism Caenorhabditis elegans (C. elegans). In this post, we’ll be taking a deep dive into the potential and difficulties involved in Whole Genome Sequencing (WGS) and RNA-Seq analyses of C. elegans.

Genome and transcriptome sequencing of C. elegans offers a multitude of scientific opportunities:

1. Understanding Genetic Diversity: Using WGS, we can explore the genetic variation within C. elegans populations, which helps us understand the impacts on characteristics, evolutionary processes, and population dynamics.
2. Studying Gene Function: RNA-Seq analysis can reveal which genes are active under different conditions or developmental stages, providing a functional layer of information on top of the genetic blueprint.
3. Identifying Mutations: WGS and RNA-Seq are powerful tools for identifying mutations. By studying changes in DNA or gene expression patterns, we can pin down the genetic basis of various traits or conditions.
4. Modeling Diseases and Finding New Treatments: C. elegans is often used to model human diseases. Coupling this with WGS or RNA-Seq can lead to the discovery of new disease-related genes and potential drug targets.

 

However, like all scientific endeavors, C. elegans sequencing isn’t without its challenges:

1. Data Handling: The volume of data generated by WGS and RNA-Seq can be immense. Storing, processing, and analyzing these data require substantial computational resources and bioinformatics expertise.
2. Technical Noise: All sequencing methods can introduce some level of bias and noise. It’s essential to design experiments in ways that minimize these issues and ensure accurate, reproducible results.
3. Interpreting Results: While we can generate a lot of genomic and transcriptomic data, understanding what these data mean functionally can be complex and requires additional experimental investigation.
4. Genome Complexity: Despite being a simple organism, the C. elegans genome has its complexities. Repeat sequences and similar genes can make analyzing sequencing data tricky.

 

At Deep Biotech Solutions, we offer Whole RNA Sequencing services that can help researchers navigate the challenges and unlock the potential of C. elegans genomic research.

The promise and challenges of C. elegans sequencing reflect the broader landscape of genomics research – a field of vast potential punctuated by intricate challenges. As we continue to improve our technologies and understanding, we get ever closer to fully unlocking the secrets held within the genomes of organisms like C. elegans.

At Deep Biotech Solutions, we’re dedicated to exploring the boundless potential of genomics. One of the key services we provide is Whole RNA Sequencing, a revolutionary tool that allows us to look at the entire set of RNA molecules in a cell or tissue. One of the factors that can influence the success of this procedure is the number of reads. In this post, we’re diving deep into the role of read numbers and their importance in RNA Sequencing (RNA-Seq).

What are “reads”? Simply put, reads refer to the sequence of RNA building blocks (nucleotides) that we can decode in one run of our sequencing process. The total number of reads gives us a broad picture of what’s happening in the RNA world of a cell.

So why are these read numbers so important in RNA Sequencing?

1. Understanding Gene Activity: The number of reads we get for each gene gives us an idea of how active that gene is. The more reads, the higher the gene activity. This is crucial when we’re trying to find out which genes are working in specific tissues, during different stages of growth, or in response to certain treatments or conditions.
2. Discovering New RNA Molecules and Forms: When we have more reads, we increase our chances of discovering new RNA molecules and spotting alternative versions of RNA from the same gene. This is like reading a story from different perspectives, and it helps us understand the complex picture of how genes work.
3. Spotting Rare RNA Molecules: More reads also help us find rare RNA molecules that might otherwise be overlooked. In research and healthcare, these rare finds can be extremely important, like finding a rare clue in a mystery.

 

How many reads do we need? It depends on what we’re looking for:

• For a quick overview of the most active genes with RNA-Seq, we might need only 5-25 million reads per sample.
• For a wider look at gene activity and some new RNA versions, we typically aim for 30-60 million reads per sample.
• For a deep dive into the RNA world, which can uncover new RNA molecules, we may require as many as 100-200 million reads per sample.
• For targeted RNA expression, fewer reads are required. For example, Illumina suggests 3 million reads per sample for certain panels.
• For small RNA Analysis or miRNA-Seq, even fewer reads might be needed, often ranging from 1-5 million reads per sample.

 

While more RNA-Seq reads can provide a deeper look into the RNA world, it’s also a balance between cost and benefit. Generating more reads requires more resources, so it’s important to choose a number of reads that matches your goals and resources.

At Deep Biotech Solutions, we tailor our Whole RNA Sequencing service to meet your specific needs. We adjust the number of reads based on your project’s goals, ensuring the best possible outcome.

At Deep Biotech Solutions, we are committed to pushing the boundaries of biotechnological innovation. Partnering with Femtonics Kft., we have developed a new-generation COVID test that offers unprecedented speed and mutant coverage. Our state-of-the-art test redefines the benchmark for SARS-CoV-2 detection.

Harnessing the power of personalized bioinformatics analysis, we determined a consensus sequence that covers approximately 99% of the over 2.5 million SARS-CoV-2 mutants currently catalogued in the international database. This groundbreaking approach leverages a dedicated software tool, ensuring our test’s unmatched reach when it comes to identifying COVID-19 variants.

Our team of scientists designed a set of primers specifically for viral genome detection. These primers are predicted to detect additional mutants, further enhancing the test’s ability to stay one step ahead of the evolving COVID-19 threat.

In addition to its comprehensive mutant coverage, our test offers a significant speed advantage. With a reaction time approximately 20% faster than that of regular qPCR tests and other commercially available tests based on the same principle, results can be obtained in just 4-10 minutes. This level of speed and efficiency will provide critical advantages in managing the pandemic, enabling quicker isolation of infected individuals and faster contact tracing.

Our new test maintains stringent accuracy standards, demonstrating nearly 100% specificity and 98% sensitivity. These measurements mean that the test is highly reliable in correctly identifying positive cases (sensitivity) and correctly identifying negative cases (specificity).

We have developed this reagent kit to be performed at a constant temperature of 65 °C, which simplifies the testing process, making it more accessible. Furthermore, our reagent kit offers excellent value for money compared to other PCR reagents, demonstrating our commitment to providing accessible solutions in the battle against COVID-19.

We are proud to announce that this product is CE and ISO 13485 certified, affirming its compliance with health, safety, and environmental protection standards in the European Economic Area.

The journey to develop this advanced COVID test involved a comprehensive application of our biotechnology Contract Research Organization (CRO) services. From initial concept to obtaining the CE license, we incorporated every element of our portfolio. This included personalized bioinformatics analysis, molecular cloning, and our suite of custom CRO services, underscoring the power of a multi-disciplinary approach to developing critical healthcare solutions.

At Deep Biotech Solutions, we’re at the forefront of genomic technologies and research, including Whole Genome Sequencing (WGS). An essential component that influences the success of this intricate process is sequencing depth. Today, we delve into the role of sequencing depth, aiming to illuminate its significance and its application in various research and clinical scenarios.

Sequencing depth is a straightforward yet powerful concept. It refers to the average number of times a specific part of the DNA is read during the sequencing process. The depth of sequencing influences several key outcomes:

1. Detecting Genetic Variants: With a higher sequencing depth, we have a better chance of identifying differences in the DNA sequence, known as “variants”. This is particularly critical in healthcare, where spotting genetic differences, for instance, in cancer cells, can guide the most effective treatment approach.

2. Identifying Heterozygous Variants: A common scenario in genetics is having two different copies of a gene – one from each parent. These are known as heterozygous variants. A high sequencing depth allows for accurate identification of these variants, contributing to our understanding of the genomic landscape.

3. Discovering Rare Variants: Occasionally, there are DNA variants that are exceedingly rare or found in a small subset of cells. A higher sequencing depth allows us to detect these elusive variants, proving beneficial when studying heterogeneous samples, such as tumor cells, or identifying rare microorganisms.

The determination of the appropriate sequencing depth often depends on the goals of the study. For a preliminary overview of the genome, a lower sequencing depth might be adequate. However, for a thorough exploration of genetic variants and ensuring their authenticity, a higher sequencing depth is necessary.

Typically, for many applications, a medium sequencing depth, referred to as 30x coverage, is employed. This offers a balance between cost and the accuracy of the data obtained.

For more nuanced studies, like those involving cancer research where rare genetic changes need to be identified, even higher sequencing depths (like 100x or 1000x) might be required.

At Deep Biotech Solutions, our Whole Genome Sequencing service is tailored to meet varying requirements. We adjust the sequencing depth based on the specific needs of the project, ensuring optimal outcomes.

Understanding the significance of sequencing depth and its role in genomic studies enhances the effectiveness of genomics research and clinical applications. As we continue to explore the world of genomics, the balance between sequencing depth and the accuracy of results remains a key consideration.

 

Advancements in the field of genetic engineering have brought us to an exciting time where we can precisely edit the genetic code of nearly any organism. Among the various tools available, CRISPR/Cas9 stands out for its precision and versatility. However, like all scientific techniques, CRISPR/Cas9 presents certain challenges. At Deep Biotech Solutions, we have harnessed the power of deep learning to optimize CRISPR/Cas9 target efficiency, paving the way for more successful gene editing endeavors.

CRISPR/Cas9’s efficiency varies significantly based on the target site and cell type. Unintended mutations may also occur in locations other than the intended target site, known as off-target effects. Plus, the complexity of biological systems can affect the efficiency of CRISPR/Cas9, complicating predictions. There is no shortage of obstacles when it comes to perfecting this technology.

However, with problems come opportunities. Technological advancements have led to the development of predictive algorithms that assess potential target sites based on various factors, significantly enhancing the efficiency of CRISPR/Cas9 experiments. Among these advancements, the use of machine learning and deep learning tools represents a significant leap forward.

This is where Deep Biotech Solutions comes in. We have developed a state-of-the-art deep neural network-based prediction algorithm in-house. Our tool excels at predicting the efficiency of guide RNAs, the molecules that guide the Cas9 protein to the right spot in the genome. With this capability, our algorithm outshines other available options, offering an unprecedented level of precision in CRISPR/Cas9 target prediction.

But what does this mean for you? By making the most of our innovative tool, we can produce genetically modified cell lines more efficiently than ever before. These cell lines have numerous applications in research, biotechnology, and medicine, such as studying disease mechanisms, screening for potential drugs, and developing gene therapies.

Remember, science doesn’t stand still – and neither do we. As we continue to refine our predictive algorithm and broaden our understanding of biological systems, we are always striving to make the future of gene editing even brighter.