Mitochondria


Nature’s most recent “Insight” supplement is devoted to a topic near and dear to our hearts, even when spelled with that superfluous UK “e”: Ageing. From the introductory editorial:

Ageing, the accumulation of damage to molecules, cells and tissues over a lifetime, often leads to frailty and malfunction. Old age is the biggest risk factor for many diseases, including cancer and cardiovascular and neurodegenerative diseases. … Ageing research is clearly gaining momentum, as the reviews in this Insight testify, bringing hope that at some time in the future we will be able to keep age-related diseases at bay by suppressing ageing itself.

The five reviews are all by prominent scholars — many of whose work we’ve discussed here — and cover a wide range of subjects within gerontology and biogerontology:

As always, Nature Insight supplements are free-access, so even if you don’t have access to a university subscription, you can still read these articles.

(For a previous aging-related Nature Insight on DNA repair, see here.)

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Here are the biogerontological reviews from the last month or so that I’ve found interesting and noteworthy. The field as a whole continues to massively overproduce review papers; by my totally unscientific estimate, these represent less than ten percent of the review abstracts that crossed my desk since Thanksgiving.

The last installment of review roundup can be found here. As always, each Review Roundup is guaranteed to contain at least one link to a review you will find highly educational, or your money back.

Comparative biogerontology:

A while back I attended a NAKFI meeting about aging. Along with a few others, I applied for (and got) a seed grant to use comparative zoology to study aging — in a nutshell, to study the various ways that nature has solved various problems that arise during aging, and see whether we might learn something that could be applied to enhancing human healthspan or lifespan.

The initial small grant funded a series of meetings, culminating in a large-scale gathering of scientist with wide expertise not only in biogerontology but also zoology, evolutionary biology, metabolomics, and other disparate fields. While this conference didn’t end up leading to the creation a single comprehensive Comparative Biogerontology Initiative, as some of my fellow applicants had hoped, it did provoke a great deal of excellent discussion. There are a few smaller-scale efforts currently underway, initiated by people who came together to talk about the original idea.

Two of the attendees of the big meeting have published reviews recently. I haven’t asked them personally but I am assuming that they’re discussing ideas that germinated at the CBI conferences.

Gene regulation:

Inflammation:

Mitochondria:

One of the authors of the first paper is Thomas Nyström, whose lab recently described the role of cell polarity in sorting protein aggregates preferentially into the mother cell during cell division. That story lacked a significant mitochondrial component, so this review is a nice complement to the primary study published earlier this year.

Nuclear organization:

Stem cells:

Leanne Jones, the senior author on this review, is one of the folks writing the proverbial book on the critical interactions between stem cells and the tissue microenvironment. Her lab uses the Drosophila gonad as a model system.

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Yesterday we learned that the most well-characterized mammalian sirtuin, SIRT1, is involved in the control of behavior in response to food availability. SIRT1 is just one of seven sirtuins in mammalian genomes, each of which has a characteristic expression pattern, subcellular localization, and physiological importance.

Today we’re going to talk about another member of the sirtuin family, SIRT3, which has been known for a while now to localize to the mitochondria. Now, Kim et al. have shown that the SIRT3 protein acts as a tumor suppressor:

SIRT3 Is a Mitochondria-Localized Tumor Suppressor Required for Maintenance of Mitochondrial Integrity and Metabolism during Stress

The sirtuin gene family (SIRT) is hypothesized to regulate the aging process and play a role in cellular repair. This work demonstrates that SIRT3−/− mouse embryonic fibroblasts (MEFs) exhibit abnormal mitochondrial physiology as well as increases in stress-induced superoxide levels and genomic instability. Expression of a single oncogene (Myc or Ras) in SIRT3−/− MEFs results in in vitro transformation and altered intracellular metabolism. Superoxide dismutase prevents transformation by a single oncogene in SIRT3−/− MEFs and reverses the tumor-permissive phenotype as well as stress-induced genomic instability. In addition, SIRT3−/− mice develop ER/PR-positive mammary tumors. Finally, human breast and other human cancer specimens exhibit reduced SIRT3 levels. These results identify SIRT3 as a genomically expressed, mitochondria-localized tumor suppressor.

So, the absence of the SIRT3 gene disrupts mitochondrial function and destabilizes the mitochondrial genome, but the causal relationship is unclear: one can imagine a derangement of mitochondrial morphology or physiology causing the genomic instability, but one can also imagine a primary defect in DNA metabolism causing the physiological defects — and one could also imagine both, operating in a vicious cycle.

The significance of the “single oncogene” observation requires a bit of explanation: In a normal cell, simply turning on a single tumor-promoting gene isn’t sufficient to transform the cell, i.e., create a tumor — if a cell detects a hyperphysiological level of a growth factor or internally generated mitogenic signal, it will undergo senescence, a permanent cell cycle arrest that (among other things) prevents mutated cells from turning into cancers.

In the case of the SIRT3 knockout, however, a single oncogene is enough to transform an otherwise normal cell, i.e., the loss of SIRT3 function appears to potentiate the transformation process. Furthermore, this implies that the mitochondria are involved in the decision to undergo senescence. This is not to say that SIRT3 is directly involved in the senescence fate decision: the SIRT3 knockout has broadly deranged mitochondria, and it’s not clear which mitochondrial function is involved in senescence.

While this report does not fully elucidate the mechanism of SIRT3 action, it is clear that oxidation must be central to the issue: reactive oxygen species (ROS) levels are high in the SIRT3 knockout, but increasing the dose of an antioxidant enzyme prevents the single-oncogene transformation. We don’t yet know whether the ROS themselves are causing oncogenic mutations, or the loss of the mitochondrial function in senescence is allowing cells that would have arrested to progress further down the path to cancer. (Could be both, obviously, but it’s likely that one of the two is more important.)

These findings are consistent with what we already know about the connection between mitochondrial ROS, cellular damage and lifespan (ideas that are central to one strategy for developing longevity-enhancing drug compounds by targeting antioxidants to mitochondria), but they raise a question: how much of the “longevity regulation” we owe to antioxidant genes can be attributed to tumor suppression?

ResearchBlogging.orgKim, H., Patel, K., Muldoon-Jacobs, K., Bisht, K., Aykin-Burns, N., Pennington, J., van der Meer, R., Nguyen, P., Savage, J., & Owens, K. (2010). SIRT3 Is a Mitochondria-Localized Tumor Suppressor Required for Maintenance of Mitochondrial Integrity and Metabolism during Stress Cancer Cell, 17 (1), 41-52 DOI: 10.1016/j.ccr.2009.11.023

There’s lots more about SIRT3 in a recent post at @ging.

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A prominent scholar of the CLK-1 story has called the coroner on the mitochondrial free radical theory of aging (MFRTA). From Lapointe & Hekimi:

When a theory of aging ages badly

According to the widely acknowledged mitochondrial free radical theory of aging (MFRTA), the macromolecular damage that results from the production of toxic reactive oxygen species (ROS) during cellular respiration is the cause of aging. However, although it is clear that oxidative damage increases during aging, the fundamental question regarding whether mitochondrial oxidative stress is in any way causal to the aging process remains unresolved. An increasing number of studies on long-lived vertebrate species, mutants and transgenic animals have seriously challenged the pervasive MFRTA. Here, we describe some of these new results, including those pertaining to the phenotype of the long-lived Mclk1 +/− mice, which appear irreconcilable with the MFRTA. Thus, we believe that it is reasonable to now consider the MFRTA as refuted and that it is time to use the insight gained by many years of testing this theory to develop new views as to the physiological causes of aging.

MFRTA recently turned 50, and consequently has received a lot of attention lately; q.v. this review and this retrospective by Denham Harman, the originator of the theory. The thesis of most pieces seems to be that the theory hasn’t been demonstrated to explain the bulk of age-related decline, but that there’s still life in the idea. In contrast, the authors of this review argue that the relevant experiments have been performed and that the theory has been falsified — in other words, we’ve done our scientific duty and it’s now time to move on.

I doubt very much that this article will put a permanent end to the controversy. Data reported fairly recently have breathed new life into oxidative theories in general and the MFRTA in particular. While these authors contend that the CLK-1 mouse mutant contradicts the underlying mechanisms of the MFRTA, other recently reported work on this pathway supports the claim that inhibiting mitochondrial respiration delays aging, a key prediction of MFRTA.

Furthermore, if mitochondrially generated oxidative radicals are truly not playing a causative role in aging, it becomes much harder to explain how mitochondrially targeted antioxidants can extend lifespan in mammals.

ResearchBlogging.orgLapointe, J., & Hekimi, S. (2009). When a theory of aging ages badly Cellular and Molecular Life Sciences DOI: 10.1007/s00018-009-0138-8

I’m sitting in the Drexler Auditorium of the Buck Institute, where I’ve been working over the last six months. Today we’re being treated to an off-schedule “Special Institute Lecture” by Harvard’s Gregory Verdine. These are my notes about the talk; below, I’m paraphrasing Verdine’s words, not writing my own.

To fix it you have to find it: Repairing oxidative damage to DNA.

Mitochondrial metabolism generates oxygen radicals, which damage DNA and increase the risk of mutation. The primary oxidative adduct of guanine, 8-oxoG, differs by only two atoms from the original guanine, but this small difference is still enough to change the residue’s base-pairing characteristics (i.e., from G=C to 8-oxoG=A). 8-oxoG is repaired by the enzyme Ogg1, which displaces the damaged residue by covalently bonding to the DNA backbone.

Ogg1 has a tough job: oxoG-C base pairs are perfectly stable, and from the outside they look just like G-C pairs, so the detection of rare lesions within the genome poses a tremendous challenge. In crystallographic studies that exploited a catalytically dead Ogg1 enzyme (which can recognize but not cleave DNA), Verdine’s lab has shown that the binding of Ogg1 at a G-C base pair results in the extrusion of G or oxoG to the exterior of the double helix, setting the stage for repair — but how does Ogg1 find the lesion site in the first place? To answer this question, Verdine’s group visualized single molecules of Ogg1 diffusing in one dimension along a double helix. They observed that Ogg1 moves so quickly that it can’t be checking every G-C base pair along its path. Instead, Ogg1 (and other lesion-repair proteins) may exploit subtle rearrangements in the DNA backbones near lesion sites. The enzyme amplifies these local structural changes into substantial conformational changes, leading to base extrusion and starting the process of repair.

We want a new drug: Synthetic biologicals as novel pharmaceuticals

Small-molecule drugs have “good geography,” in the sense that they can cross cell membranes. However, they’re limited, in that they can only target proteins that engulf them (e.g., in hydrophobic pockets or active sites). Proteins, on the other hand, have much more diverse function, but terrible geography — they simply can’t get into cells, and they’re therefore useless for intracellular targets. Both classes of drug can attack (generously) only ~10% of prospective targets. Therefore, we need an entirely new class of drug: synthetic biologics like stapled peptides and RIPtides, which combine the bioavailability of small molecules with the functional diversity of proteins.

Stapled peptides are essentially the interaction domains of proteins, conformationally restrained in such a way that they still retain the active structure. (Think of a protein as a delivery system for an interaction domain, in which the non-interacting portions serve primarily to hold the ID in place.) An interaction domain alone would be too floppy to have a biological effect; conversely, the intact protein has the desired function but can’t cross the membrane. Solution: replace the main body of the protein with a hydrocarbon “staple” that keeps the interactive domain in the active conformation, without substantially increasing its size. Surprisingly, stapled peptides are taken up by cells via an energy-dependent active transport process, one upshot of which is that they don’t need to be uncharged and hydrophobic in order to cross the membrane. Drugs of this kind have already been used in animal studies to suppress leukemia by activating apoptotic factors in tumor cells.

Declaring open season on transcription factors

A brief concluding note: Transcription factors are among the most well-validated prospective targets, but they have historically been outside the scope of drug developers. TFs function primarily by protein-protein interactions that aren’t amenable to interference by small-molecule drugs. Recently, however, Verdine and others have been able to use synthetic biologicals to interfere with a specific oncogenic transcription factors.

My own comments

OK, so, not a lot of specific about either aging or cancer, but the idea of a novel class of pharmaceuticals that could be used to attack the “missing 80%” of validated prospective drug targets is still very exciting.

The free radical theory of aging (FRTA) was first advanced by Denham Harman more than 50 years ago. The theory proceeds logically from a small number of straightforward assumptions, based on observations from radiation biology. From the Science of Aging Timeline:

Harman’s logic proceeds from three observations: (1) irradiation causes premature aging; (2) irradiation creates oxygen radicals, which may mediate its effects; and (3) cells produce oxygen radicals under normal conditions. From these premises, he theorized that aging could be caused by endogenously generated oxygen radicals.

Over a half-century, the FRTA has evolved substantially (eventually focusing on the mitochondria as a major source of the initially postulated endogenous radicals), and has lately been the subject of several reviews evaluating its explanatory power and extent of current acceptance.

A unique perspective on the FRTA’s history has recently been provided by none other than its initiator, Denham Harman, who is retired but still intellectually active. From his review:

Origin and evolution of the free radical theory of aging: a brief personal history, 1954–2009

Aging is the progressive accumulation in an organism of diverse, deleterious changes with time that increase the chance of disease and death. The basic chemical process underlying aging was first advanced by the free radical theory of aging (FRTA) in 1954: the reaction of active free radicals, normally produced in the organisms, with cellular constituents initiates the changes associated with aging. The involvement of free radicals in aging is related to their key role in the origin and evolution of life. The initial low acceptance of the FRTA by the scientific community, its slow growth, manifested by meetings and occasional papers based on the theory, prompted this account of the intermittent growth of acceptance of the theory over the past nearly 55 years.

It’s a very personal account, starting with the educational experiences that Harman credits with putting him in the right place at the right time, continuing with a description of the origins of the theory, and paying a great deal of attention to the “fits-and-starts” advancement of the theory toward broad acceptance (though not without effort and extensive modification). Pieces like these, in which the originator of a hugely influential theory provides their individual perspective on the consequences of their work, are rare indeed — hence this is a must-read for students and practitioners of biogerontology.

ResearchBlogging.orgHarman, D. (2009). Origin and evolution of the free radical theory of aging: a brief personal history, 1954–2009 Biogerontology DOI: 10.1007/s10522-009-9234-2

Here’s the latest in our (infrequent and irregular) series of “review roundups” — links, without extensive further comment, to the reviews I found most intriguing over the past few weeks. For the previous foray into the secondary literature, see here.

Remember, each Review Roundup is guaranteed to contain at least one link to a review you will find highly educational, or your money back.

Autophagy:

Chaperones:

Evolution:

Glycation:

Immunology:

Mitochondria:

Neurodegeneration:

Resveratrol:

Senescence:

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