Erasing Mitochondrial Mutations

Researchers develop a method to selectively remove mutated mitochondrial DNA from the murine germline and single-celled mouse embryos.

By | April 23, 2015


Mutations in mitochondrial DNA (mtDNA) can be specifically targeted and removed by transcription activator-like effector nucleases (TALENs) in murine oocytes, single-celled mouse embryos, and fused human-mouse hybrid cells, providing proof of principle for a method that could one day be used to treat certain hereditary mitochondrial disorders in people, according to a study published today (April 23) in Cell.

“It’s an extremely important step,” said Valerio Carelli of the University of Bologna, Italy. “The results are very relevant and very convincing.”

Between 1,000 and 100,000 mitochondria power each human cell. Often, mitochondria in the same cell have different genomes, or haplotypes, a condition known as heteroplasmy. Certain haplotypes include mutations that impact mitochondrial function and cause disease, particularly in energy-hungry organs such as the brain and heart. Because mitochondria segregate randomly as cells divide, it is impossible to determine early in embryonic development how a mix of wild-type and mutated mitochondria inherited from the mother will affect an organism.

To rid mitochondria of these harmful mutations, researchers have used restriction enzymes as well as zinc-finger nucleases (ZFNs) and TALENs, which can be designed to recognize any DNA sequence, to cut and eliminate mutated mitochondrial genomes from heteroplasmic cells.

“Because the cell likes keeping the number of mtDNA molecules constant, after elimination of the faulty ones, the wild-type copy will repopulate the cell,” explained Michal Minczuk of the Medical Research Council (MRC) Mitochondrial Biology Unit in Cambridge, U.K., who helped develop ZFN mtDNA-editing.

Now, an international team led by researchers at the Salk Institute for Biological Studies in La Jolla, California, has used mitochondria-targeting restriction enzymes and TALENs in the mammalian germline and early-stage mouse embryos for the first time. Injecting mRNAs encoding each enzyme into mouse cells with two different wild-type mtDNA haplotypes selectively removed the targeted genome variant, and the edited embryos grew into normal mice. The team did not observe any off-target effects.

To determine whether the enzymes could be used to edit human mtDNA, the researchers fused mouse oocytes with fibroblast cells from patients with one of two mitochondrial disorders—Leber’s hereditary optic neuropathy or neurogenic muscle weakness, ataxia, and retinitis pigmentosa. Unlike in the mouse experiment, mutant mtDNA was still detectable, albeit at lower levels, after TALEN mRNA injection. Mutated mtDNA usually only causes disease if more than 60 percent to 75 percent of a cell’s mitochondria harbor the error, so “the reduction that we observed was more than enough for the phenotype to disappear,” study coauthor Juan Carlos Izpisua Belmonte of the Salk Institute said.

The scientists hope to offer the technique as an alternative to mitochondrial replacement therapy—a procedure also known as “three-parent IVF” that was recently green-lighted by the U.K. parliament. Injecting an mRNA is similar to routine injections of sperm in fertility clinics and therefore more practical than the complicated process of adding the nucleus from the egg of a mother with mitochondrial disease to an enucleated egg from a woman with normal mitochondria, Belmonte said.

While mitochondrial replacement advocate Doug Turnbull of Newcastle University, U.K., praised this latest advance in an e-mail as “elegant and exciting,” he cautioned that the technique “may be of limited value for those women whose oocytes have either large amounts of mutated mitochondrial DNA or all mutated mitochondrial DNA.” Carelli agreed, but said he believes enzyme treatment would be much better for heteroplasmic diseases because it does not require a third donor and therefore, unlike mitochondrial donation, does not combine different sources of mitochondria.

Belmonte’s group is already testing its editing technique on discarded embryos of patients with mutated mtDNA. It must be shown that mitochondria are not severely depleted during the procedure, Carelli said. Minczuk wondered whether the method would work as well on diseased haplotypes as it did on the two healthy mouse variants. There are ethical questions, too, as many scientists have called for a moratorium on genome editing in the human germline.

Belmonte envisions that the technique could also play a role in gene therapy for other disorders. “If we can edit the mitochondrial DNA to make a better mitochondrion, I am convinced this will have an impact on so many other processes,” he said.

P. Reddy et al., “Selective elimination of mitochondrial mutations in the germline by genome editing,” Cell, doi:10.1016/j.cell.2015.03.051, 2015.

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Avatar of: Keith Loritz

Keith Loritz

Posts: 27

April 24, 2015

Cool stuff is happening! s.l.o.w.l.y.

"scientists have called for a moratorium on genome editing in the human germline". Don't tell mother nature about this one. I think we call it evolution (the hard way). Nature is too slow, too dirty and too inefficient for mans' needs. This fact will become all too apparent to the general public as we keep running into walls that restrict what we MUST do. GMOs. . . nature does it by accident, we must do it with understanding, it is the only way forward.

Avatar of: JimmyJ


Posts: 3

April 24, 2015

I wonder if this can be used in vivo for somatic gene therapy? It could be used to establish whether or not aging is in part caused by point mutations (deletions) in mitochondira which then cause the the diseased mitochondria to take over the cell, eventually exporting ROS.

Avatar of: Juntaceo


Posts: 2

Replied to a comment from Keith Loritz made on April 24, 2015

April 24, 2015

Most Bioethicists would argue that the potential for great harm if these experiments go awry , outweighs any laudable but unproven potential benefits.

Evolution is random, and currently Modern Human evolution is poorly understood, and no general concesus exists.

Evolution is not in any comparable to the kiinds of rewriting of the genome you are suggesting.

I don't think you get to speak for "Man's needs"

Avatar of: JonRichfield


Posts: 134

April 28, 2015

<siiiiigh....!!!> "many scientists have called for a moratorium on genome editing in the human germline", have they?

Pardon me while I get the heaves.

If you try to please everybody with something, someone won't like your trying to please everybody with anything...

I don't know what these so-called scientists think a moratorium would solve; it won't get us any nearer to either a solution or an understanding; it won't make anything worse than things are already, and it also forbids people suffering in an unacceptable status quo from risking an unacceptable consequence -- or a termination -- or a cure.

And it is for this that our species developed logic, science and ethics??? Not to mention technology, courage and hope?

A great darkness of the spirit descends upon my eyes as my gorge rises.

But I'll abstain from trying to do anything about it, for dread that someone might not like it and call for a moratorium on something... or anything...

Avatar of: Charles Barnard

Charles Barnard

Posts: 1

August 3, 2017

As I write this, the first human embryonic editing for disease has been announced.

Of course, this has been possible for some time, so the milestone lies primarily in the feeling of security of the researchers to announce their work, rather than the work itself.

I would like to see more work concentrating upon error correction routines for DNA, since such a correction could easily extend useful life of replicated DNA manyfold, without a CRISPR module having to scan the entire strand to locate any possible defects.

Proper error correction could provide a CRISPR based tool with the exact location, type and correction for each error.

For those who feel this has come slowly, rememer that once this genie is out of the bottle, I can carry materials to make literally millions of DNA changes in the space of a flashdrive. Properly encoded, I can fit it all onto a few strands of artificial DNA, or into a bacterium.

It is only a matter of weeks from the first simple cosmetic changer (eye-color, hair restorer, whatever,) to it becoming available around the world.

How do we even tell who is who when you can change your appearance easily in a matter of days or weeks?

How do you stop terrorists who bomb you with DNA mutating diseases?

Someone can now design a delivery system and tool which can selectively eliminate all people of a particular branch of humanity...or any of a million things.

You think computer malware is a problem....?

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