Work on Allotopic Expression of Mitochondrial Genes is Spreading

Gene expression is the process of generating proteins from the blueprints encoded in DNA. Most DNA is in the cell nucleus, but thirteen genes can be found in the mitochondria, the powerplants of the cell that were once, long ago, symbiotic bacteria. Alloptic expression is a form of genetic engineering wherein one or more of those mitochondrial genes is copied into nuclear DNA, and the resulting proteins transported back to the mitochondria where they are needed. It is that transportation that is the hard part, not yet accomplished for more than a couple of mitochondrial genes.

Why should we care about allotopic expression as anything more than a technical curiosity? Because mitochondrial DNA damage is one of the root causes of aging. Mutations that disable some mitochondrial genes, thus depriving mitochondria of necessary protein machinery, lead to a chain of unfortunate events that progressively produces ever more dysfunctional cells and damage to tissues and organs over the years. If researchers could create a backup source of the necessary proteins in the cell nucleus, then this contribution to aging could be completely removed - and even reversed in its later stages.

The SENS Research Foundation is more or less the only group coordinating work on allotopic expression for the treatment of aging, but a number of unaffiliated labs are using the approach in a more limited way in an attempt to address the genetic disease of Leber hereditary optic neuropathy (LHON). LHON is caused by a defective mitochondrial gene, so many of the efforts taken to cure it are also somewhat applicable to the issue of mitochondrial mutations in aging. Here researchers demonstrate effectiveness and safety of allotopic expression in this case:

We developed a novel strategy for treatment of Leber hereditary optic neuropathy (LHON) caused by a mutation in the nicotinamide adenine dinucleotide dehydrogenase subunit IV (ND4) mitochondrial gene. In a series of laboratory experiments, we modified the mitochondrial ND4 subunit of complex I in the nuclear genetic code for import into mitochondria. The protein was targeted into the organelle by agency of a targeting sequence (allotopic expression). The gene was packaged into adeno-associated viral vectors and then vitreally injected into rodent, nonhuman primate, and ex vivo human eyes that underwent testing for expression and integration.

We tested for rescue of visual loss in rodent eyes also injected with a mutant G11778A ND4 homologue responsible for most cases of LHON. We found human ND4 expressed in almost all mouse retinal ganglion cells by 1 week after injection and ND4 integrated into the mouse complex I. In rodent eyes also injected with a mutant allotopic ND4, wild-type allotopic ND4 prevented defective adenosine triphosphate synthesis, suppressed visual loss, reduced apoptosis of retinal ganglion cells, and prevented demise of axons in the optic nerve. Injection of ND4 in the ex vivo human eye resulted in expression in most retinal ganglion cells. Primates undergoing vitreal injection with the ND4 test article and followed up for 3 months had no serious adverse reactions.



That is very good news that this research is spreading.

Does anyone know if:

1 ~ The use of a targeting sequence to get the protein into the mitochondria is the same approach as that being studied by Matthew O'Conner and the SENS foundation?

2 ~ Looking at Matthew "Oki" O'Conner request for funding from I see that ND4 is part of Complex 1 which unfortunately has 6 other proteins from genes kept in the mitochodria rather than the nucleus. Will this approach work for the other 6 proteins in complex 1? What is to stop this working for all 13 proteins/genes?

Posted by: Jim at March 24th, 2014 6:40 AM

@Jim: My understanding is that each of the proteins is sufficiently different that the current more established methods of transport have to be essentially completely reworked for each. A long process of discovery and experimentation is needed to find the right approach within the method.

There have been early signs of more general methods that might work for all of the proteins with comparatively little rework, but nothing has robustly emerged there yet. e.g. see:

For background on the issue:


One of the challenges to executing this plan is that the 13 remaining proteins would actually be quite difficult for the mitochondria to import, because once they are formed in the main body of the cell, they quickly fold up on themselves, making it difficult or impossible for them to be threaded through the pores in the mitochondrial membrane. This is probably why evolutionary pressure hasn’t already forced the genes to relocate.

But there are several potential ways around this problem. One is to look to solutions discovered by evolution in other organisms. In several cases, evolution has favored the retention of modified versions of the genes for the very same proteins that our mitochondria still encode in their own DNA, because the proteins made by these modified genes have less tendency to snarl up in the cell body. This resistance to snarling means that when the genes for the modified version of the proteins are encoded in the nucleus, they can be produced in the main body of the cell, and still thread their way into the mitochondria. It’s possible that suitably-modified versions of these organisms’ nuclear-encoded mitochondrial genes could work for us, too.

Alternatively, we may be able to insert disposable molecular “braces” called inteins into the sequence of the proteins, that would temporarily hold them straight enough to let them pass through the membranes.

A third approach, which SENS Research Foundation is now most actively researching, was pioneered by Professor Marisol Corral-Debrinski at the Institut de la Vision at Pierre and Marie Curie University, Paris. She altered the genes for the proteins that need to be moved to the nucleus so that the protein would be “decoded” from its instructions very close to the mitochondrial surface, instead of far away in the cell body. This approach allows the allotopically-expressed proteins to be threaded directly into the mitochondria before they have the chance to twist up too much.


Posted by: Reason at March 24th, 2014 6:08 PM
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