In order to survive, flourish and successfully reproduce, organisms rely on a high degree of genetic stability. Mutagenic agents, which can threaten the integrity of the genetic code by causing mutations in DNA, pose a serious risk to human health. They have long been implicated in a range of genetically inherited afflictions, as well as cancer, aging and neurodegenerative diseases like Alzheimer’s.

It now appears that mutagenic threats to a cell’s subtle machinery may be far more widespread than previously appreciated. In a new study, Arizona State University's Michael Lynch and his colleagues demonstrate that DNA mutation itself may represent only a fraction of the health-related havoc caused by mutagens. Michael Lynch is the director of the Biodesign Center for Mechanisms of Evolution and professor at ASU's School of Life Sciences. Download Full Image

The study highlights the ability of mutagenic compounds to also affect the process of transcription, during which a DNA sequence is converted (or transcribed) to mRNA, an intermediary stage preceding translation into protein.

The research findings, which highlight mutagenic transcription errors in yeast, worms, flies and mice, suggest that the harmful effects of mutagens on transcription are likely much more pervasive than previously appreciated — a fact that may have momentous implications for human health.

“Our results have the potential to completely transform the way we think about the consequences of environmental mutagens,” Lynch said.

Professor Lynch is the director of the Biodesign Center for Mechanisms in Evolution and a researcher in ASU’s School of Life Sciences.

The research results appear in the current issue of the journal PNAS.

Cells under threat

Due to their important role in disease processes, mutagenic compounds have long been a topic of intensive scientific study. Such agents include sunlight and other sources of radiation, chemotherapeutics, toxic byproducts of cellular metabolism or chemicals present in food and water.

Mutagens can inflict damage to the DNA, which can later snowball when cells divide, and DNA replication multiplies these errors. Such mutations, if not corrected through DNA proofreading mechanisms, can be passed to subsequent generations and depending on the location at which they appear along the human DNA strand’s 3-billion-letter code may seriously impact health, in some cases, with lethal results.

But even if repaired prior to replication, transiently damaged DNA can also interfere with transcription — the process of producing RNA from a DNA sequence. This can happen when RNA polymerase, an enzyme that moves along a single strand of DNA, producing a complementary RNA strand, reads a mutated sequence of DNA, causing an error in the resulting RNA transcript.

Because RNA transcripts are the templates for producing proteins, transcription errors can produce aberrant proteins harmful to health or terminate protein synthesis altogether. It is already known that even under the best of conditions, transcript error rates are orders of magnitude higher than those at the DNA level.

RNA: A string of errors?

While the existence of transcription errors has long been recognized, their quantification has been challenging. The new study describes a clever technique for ferreting out transcription errors caused by mutagens and separating these from experimental artifacts — mutations caused during library preparation of RNA transcripts through processes of reverse-transcription and sequencing. 

The method described involves the use of massively parallel sequencing technology to identify only those errors in RNA sequence directly caused by the activity of a mutagen. The results demonstrate that at least some mutagenic compounds are potent sources of both genomic mutations and abundant transcription errors.

The circular sequencing assay outlined in the study creates redundancies in the reverse-transcribed message, providing a means of proofreading the resultant linear DNA. In this way, researchers can confirm that the transcription errors observed are a result of the mutagen’s effects on transcription and not an artifact of sample preparation.

The DNA molecule has been shown to be particularly vulnerable to a class of mutagens known as alkylating agents. One of these, known as MNNG, was used to inflict transcriptional errors on the four study organisms. The effects observed were dose-dependent, with higher levels of mutagen causing a corresponding increase in transcriptional errors.

Hidden mistakes may be costly to health

Transcription errors differ from mutations in the genome in at least one vital respect. While DNA replication during cell division acts to amplify mutations to the genome, transcription errors can accumulate in nondividing cells, with a single mutated DNA template giving rise to multiple abnormal RNA transcripts. 

The full effects of these transcription errors on human health remain largely speculative because they have not been amenable to study until now. Using the new technique, researchers can mine the transcriptome — the full library of a living cell’s RNA transcripts, searching for errors caused by mutagens.

While the new research offers hope for a more thorough understanding of the relationship between various mutagens and human health, it is also a cautionary tale.  A preoccupation with mutational defects in DNA sequence may have blinded science to the potential effects of agents that result in transcription errors without leaving permanent traces in the genome.

This fact raises the possibility that a broad range of environmental factors as well as chemicals and foods deemed safe for human consumption are in need of careful reevaluation based on their potential for producing transcriptional mutagenesis. Further, transcriptional errors in both dividing and nondividing cell types are likely key players in the complex processes of physical aging and mental decline.