Muscle’s master regulator also moonlights as a gene silencer, study reveals

For more than 30 years, scientists have studied how the myogenic determination gene number 1 (MYOD) protein binds DNA to modify the gene expression of muscle stem cells. Similar to the instant kung fu education Keanu Reeves downloaded in “The Matrix,” MYOD plugs into muscle stem cell DNA and reprograms the cells to build muscle.

MYOD also comes to the rescue when muscle tissue needs to be repaired after injury or to restore minor damage that occurs with athletic training or other physical activity. The transcription factor rallies nearby muscle stem cells to expand in number and become muscle cells capable of regenerating harmed muscle fibers.

Like Spiderman hiding in plain sight as the unassuming photojournalist Peter Parker, this activator of genes specific to muscle has been harboring a secret identity. Scientists at Sanford Burnham Prebys and their international colleagues published findings in Genes and Development demonstrating that MYOD has its own Jekyll-and-Hyde twist, turning from a gene activator to a gene silencer.

“If you think of a cell like a house, then gene expression can be seen as the furniture that plays a major part in defining its unique identity,” said Pier Lorenzo Puri, MD, professor in the Center for Cardiovascular and Muscular Diseases at Sanford Burnham Prebys and the senior and co-corresponding author of the study.

“We focus a lot on MYOD’s traditional role of bringing in the new furniture appropriate for a muscle cell, but there is a critical first step of clearing out the old furniture to reset the cell’s identity.”

The research team examined MYOD binding events in human fibroblast cells during the process of MYOD reprogramming them into skeletal muscle cells. This experimental setting mimics the physiological process of muscle stem cells reprogramming into the myogenic lineage that occurs during muscle regeneration, and indeed the results were validated within the context of muscle regeneration following myotrauma in a mouse model.

One-third of the binding events were found at the conventional MYOD binding sites (the myogenic E-box motifs) at regulatory elements of the genome, consistent with MYOD’s traditional role as a gene activator. More than one-half of the binding events, however, occurred at the regulatory elements of downregulated genes, where DNA is packaged in such a way as to be less accessible to being transcribed into proteins, and coincided with the presence of DNA binding sites other than the E-box motifs.

This finding challenges the dogma that historically restricts MYOD DNA binding properties to the E-box motifs.

Furthermore, the scientists observed that the MYOD binding events associated with gene repression were found at genes involved in cell growth, cell proliferation, cell-of-origin as well as alternative cell lineages. This observation fits into the proposed new role of MYOD as a driver of cell reprogramming by removing the cell’s prior gene expression “furniture.”

“We discovered that MYOD has the ability to promiscuously bind the DNA at previously unexpected places,” said Puri.

“These locations were occupied by transcription factors that were promoting the expression of the cell’s origin lineage genes, so MYOD is binding there to erase the previous lineage prior to turning cells into the myogenic lineage.”

Puri and the team see their findings as an opportunity to expand current ideas about how transcription factors operate.

“We have provided seminal evidence that the same transcriptional activator can also play a repressor role at the very beginning of the process of cell transdifferentiation or reprogramming,” said Puri.

“Transcription factors are way more versatile than we thought, and this newfound versatility is dictated by where and how they bind to DNA.”

Puri says that the group’s findings regarding cellular reprogramming may help advance efforts to develop regenerative medicine therapies and to better understand the process of cellular reprogramming itself.

“In regenerative medicine, we hope to treat certain medical conditions by turning one cell type into another, one pathological cell into one physiologically normal or even therapeutic cell,” said Puri.

“And now we know that an important task is the repression of the previous lineage’s gene expression furniture.”

Puri also emphasized MYOD’s role in filtering out competing biochemical signals during cellular reprogramming.

“There are a variety of growth factors or regeneration cues that typically encounter these cells during the regeneration period,” said Puri.

“MYOD is able to be very selective in repressing most of the gene expression that would be activated by these cues in order to curate the proper program for building muscle.”

Next, the research team plans to explore what happens when MYOD’s repression of the cell’s prior identity is incomplete. This phenomenon may help explain why some athletes’ muscles recover better as they get older or why some people suffer from the age-related muscle mass deterioration and frailty known as sarcopenia at a younger age.

“It may be that small alterations in MYOD’s silencing role are tolerated by the body but progressively impair muscle function,” said Puri.

“Better understanding this concept may have an enormous impact in terms of biomedical applications for regenerative and sports medicine for athletes and sarcopenia patients.”

Puri shared that children suffering from muscular dystrophy experience a transition period called the honeymoon. For a length of time that varies with each child, their bodies can still deal with the disease by regenerating their muscle.

“If we can better understand this honeymoon period, then we may be able to use regenerative medicine approaches to extend it for as long as possible,” said Puri.

Chiara Nicoletti, Ph.D., a postdoctoral associate in the Puri lab at Sanford Burnham Prebys, shares first authorship of the study with Jimmy Massenet, Ph.D., also a postdoctoral associate in the Puri lab.

For more than 30 years, scientists have studied how the myogenic determination gene number 1 (MYOD) protein binds DNA to modify the gene expression of muscle stem cells. Similar to the instant kung fu education Keanu Reeves downloaded in “The Matrix,” MYOD plugs into muscle stem cell DNA and reprograms the cells to build muscle.

MYOD also comes to the rescue when muscle tissue needs to be repaired after injury or to restore minor damage that occurs with athletic training or other physical activity. The transcription factor rallies nearby muscle stem cells to expand in number and become muscle cells capable of regenerating harmed muscle fibers.

Like Spiderman hiding in plain sight as the unassuming photojournalist Peter Parker, this activator of genes specific to muscle has been harboring a secret identity. Scientists at Sanford Burnham Prebys and their international colleagues published findings in Genes and Development demonstrating that MYOD has its own Jekyll-and-Hyde twist, turning from a gene activator to a gene silencer.

“If you think of a cell like a house, then gene expression can be seen as the furniture that plays a major part in defining its unique identity,” said Pier Lorenzo Puri, MD, professor in the Center for Cardiovascular and Muscular Diseases at Sanford Burnham Prebys and the senior and co-corresponding author of the study.

“We focus a lot on MYOD’s traditional role of bringing in the new furniture appropriate for a muscle cell, but there is a critical first step of clearing out the old furniture to reset the cell’s identity.”

The research team examined MYOD binding events in human fibroblast cells during the process of MYOD reprogramming them into skeletal muscle cells. This experimental setting mimics the physiological process of muscle stem cells reprogramming into the myogenic lineage that occurs during muscle regeneration, and indeed the results were validated within the context of muscle regeneration following myotrauma in a mouse model.

One-third of the binding events were found at the conventional MYOD binding sites (the myogenic E-box motifs) at regulatory elements of the genome, consistent with MYOD’s traditional role as a gene activator. More than one-half of the binding events, however, occurred at the regulatory elements of downregulated genes, where DNA is packaged in such a way as to be less accessible to being transcribed into proteins, and coincided with the presence of DNA binding sites other than the E-box motifs.

This finding challenges the dogma that historically restricts MYOD DNA binding properties to the E-box motifs.

Furthermore, the scientists observed that the MYOD binding events associated with gene repression were found at genes involved in cell growth, cell proliferation, cell-of-origin as well as alternative cell lineages. This observation fits into the proposed new role of MYOD as a driver of cell reprogramming by removing the cell’s prior gene expression “furniture.”

“We discovered that MYOD has the ability to promiscuously bind the DNA at previously unexpected places,” said Puri.

“These locations were occupied by transcription factors that were promoting the expression of the cell’s origin lineage genes, so MYOD is binding there to erase the previous lineage prior to turning cells into the myogenic lineage.”

Puri and the team see their findings as an opportunity to expand current ideas about how transcription factors operate.

“We have provided seminal evidence that the same transcriptional activator can also play a repressor role at the very beginning of the process of cell transdifferentiation or reprogramming,” said Puri.

“Transcription factors are way more versatile than we thought, and this newfound versatility is dictated by where and how they bind to DNA.”

Puri says that the group’s findings regarding cellular reprogramming may help advance efforts to develop regenerative medicine therapies and to better understand the process of cellular reprogramming itself.

“In regenerative medicine, we hope to treat certain medical conditions by turning one cell type into another, one pathological cell into one physiologically normal or even therapeutic cell,” said Puri.

“And now we know that an important task is the repression of the previous lineage’s gene expression furniture.”

Puri also emphasized MYOD’s role in filtering out competing biochemical signals during cellular reprogramming.

“There are a variety of growth factors or regeneration cues that typically encounter these cells during the regeneration period,” said Puri.

“MYOD is able to be very selective in repressing most of the gene expression that would be activated by these cues in order to curate the proper program for building muscle.”

Next, the research team plans to explore what happens when MYOD’s repression of the cell’s prior identity is incomplete. This phenomenon may help explain why some athletes’ muscles recover better as they get older or why some people suffer from the age-related muscle mass deterioration and frailty known as sarcopenia at a younger age.

“It may be that small alterations in MYOD’s silencing role are tolerated by the body but progressively impair muscle function,” said Puri.

“Better understanding this concept may have an enormous impact in terms of biomedical applications for regenerative and sports medicine for athletes and sarcopenia patients.”

Puri shared that children suffering from muscular dystrophy experience a transition period called the honeymoon. For a length of time that varies with each child, their bodies can still deal with the disease by regenerating their muscle.

“If we can better understand this honeymoon period, then we may be able to use regenerative medicine approaches to extend it for as long as possible,” said Puri.

Chiara Nicoletti, Ph.D., a postdoctoral associate in the Puri lab at Sanford Burnham Prebys, shares first authorship of the study with Jimmy Massenet, Ph.D., also a postdoctoral associate in the Puri lab.