User:Kinkreet/Role of microRNA processing in adipose tissue in stress defense and longevity

Background
Aging is a process which can be defined as an inability to maintain homeostasis, respond to stress, as well as a general metabolic deterioration.

Calorie restriction (CR) is a dietary regimen that restricts calorie intake (but not malnutrition), and has been widely-accepted to be a method of increasing lifespan in mice, where mice which undergo CR live 60% longer than ad libitum-fed mice, as well as across different species. The effects of calorie restriction is vast, and includes decreased production of reactive oxygen species, reduced levels of circulating proinflammatory cytokines, increased expression of protein chaperones, increased detoxification pathways, enhanced DNA repair processes, decreased apoptosis, and reduced cellular senescence. The insulin/IGF-1 signaling and the heat shock pathways have been attributed as the effectors of CR; in this paper, the author attempts to attribute microRNAs (miRNAs) also as partly responsible for the effects of CR.

To study the role of miRNA in CR, Kahn chose to used adipose tissues as the model tissue used in this study. This is because it has previously been shown that the removal of adipose tissue mimicked the effect of CR of increasing longevity. And indeed obesity is associated with accelerated onset of age-related diseases . In particular, visceral adipose tissue accretion is a hallmark of aging. Additionally, a high proportion of adipocytes in the bone marrow reduces the success rate of haematopoietic stem cell transplantation and prevents stem cell self-renewal - signs of senescence. And so adipose tissue, Kahn argues, is the most prominent tissue affected by aging. Furthermore, it has been shown that adipocytes secrete vesicles containing small RNAs. If miRNAs do in fact confer effects of CR, it'd explain how aging at one part of the body can potentially spread to all cells of the organism through RNA trafficking. This spread of RNA from one cell to the next has been observed in plants  and now also in rodents. This might also be an alternative explanation of how the serum of young mice is able to rejuvenate old mice in parabiosis experiments.

microRNAs
Over the past decade, small non-coding RNAs (sncRNAs) have emerged to have functions in regulating gene expression. The most prominent families of these sncRNA have been the small interfering RNAs (siRNAs) and microRNAs (miRNAs). miRNA begins life as a transcribed (from RNAPII ) primary miRNA molecule (pri-miRNA), which is then processed by Drosha in the nucleus to generate precursor miRNAs (pre-miRNAs); the pre-miRNAs are then exported from the nucleus into the cytoplasm using exportin-5, to be finally cleaved by the Dicer complex to give mature miRNAs. These miRNAs then binds to their perspective target mRNA and inhibit their translation and/or lead to their degradation.

miRNAs and aging
There are many correlations between miRNA and aging - as mice ages, or when aging is induced by stressors such as H2O2, glucose oxidase, and paraquat, the global level of mature miRNAs, as well as the mRNA levels of the microRNA processing enzymes (Dicer and exportin-5) in adipose tissues are decreased. This effect has also been observed in human preadipocytes and nematodes intestine (a close analog of fat in mammals). What is more striking, perhaps, is that CR was able to reverse this decline.

Having observed the effect of aging on the miRNA system, Kahn and colleagues tested the reciprocal. Loss-of-function mutation in, or the KO of Dicer, the a central component in miRNA generation, leads to cells exhibiting signs of senescence and sensitivity to oxidative stress. The opposite is true when Dicer were overexpressed. This has been affirmed in C. elegans.

Possible mechanism
The mechanism behind this relationship is unclear, however there maybe some clues. The lin-4/miR-125 family has shown in C. elegans and mammalian cells to increase lifespan if overexpressed, and decrease lifespan if knocked down. It is proposed that lin-4/miR-125 function by negatively regulating p53.

p53 is a tumor suppressor protein which acts as a transcriptional activator for p21, which in turn interacts with cell division-stimulating protein (cdk2). Cdk2 can be bound by cyclins E and A, to permit G1/S transition and S phase progression; however, the presence of p21 means cdk2 binds to p21 instead of the cyclins, preventing cell cycle progression and cell proliferation. Tumor cells require p21, and thus p53 to be inactive, in order to proliferate and build up a large enough mass for metastasis.

When a cell is unstressed, cell cycle progression is favored, and p53 is in the inactive, phosphorylated state ; p53 is constantly being monoubiquitinated by MDM2, subsequently leading to its proteasome-dependent degradation.

miR-125b binds to the 3' UTC of the p53 mRNA and inhibits its function. Because the level of miRNA processing enzymes, and consequently of miR-125b, decreases with age, inhibition of p53 also decreases with age, and so cells are no longer prevented from entering the cell cycle, indicating that the cell is more susceptible to entering damaged cells into the cell cycle, leading to proliferation of these damaged cells. Because these aged, Dicer-deficient cells have a higher proliferative potential that Dicer-sufficient cells, if these cells have also acquired insulin resistance, then it will lead to systemic insulin resistance and cause diabetes. Even if the cells do not have insulin resistance, its high proliferative property gives a higher risks of cancers. Thus a mechanism dependent on miRNAs might be able to explain the correlation between senescence and diseases which have a higher incidence at higher ages, such as cancers, inflammatory diseases, and diabetes.

Clinical Relevance
Overexpression of Dicer in patients with cancers or diabetes might be possible way to prevent damaged tissues from spreading and slow down aging without actual change in diet through calorie restriction. Indeed, overexpression of Dicer in the intestine of C. elegans do prevent lin-4/miR-125-dependent aging and increase stress resistance.

If miRNAs are in fact suppressors of senescence and inducer of stress resistance, and if miRNAs do in fact circulate to other parts of the body via small vesicles, then it is possible that circulating miRNAs can act as a biomarker for age-related diseases and cancers.

Non-specificity of Dicer
Dicer is a RNase III endoribonuclease which is not only involved in miRNA processing, but also in RNA interference where it cleaves dsRNAs into 20-25 double-stranded nucleotide fragments; the sense strand is degraded leaving a single-stranded small non-coding RNA termed small interfering RNAs (siRNAs). Both siRNAs and miRNAs act to silence genes, but perform this function using different mechanisms. The anti-sense siRNAs hybridize with its complementary sequences sense mRNA, which causes the mRNA to be cleaved by Argonaute. The mRNA is fragmented and can no longer be translated into the full protein. miRNA binds to complementary sequences at the 3' untranslated region (3' UTR) of mRNA and promote its degradation and/or physically prevent the mRNA from being translated by the ribosome.

Dicer and Argonaute, and exportin-5 are involved in the siRNA and miRNA processing and exportation from the non-nucleolar regions of the nucleus, respectively. The level of exportin-5 is highly correlated with the rate of silencing and is thus likely to be the rate-limiting step in vivo. Furthermore, exogenous introduction of shRNA and/or siRNA inhibit the function of miRNA, probably by competing with miRNA to bind to Exp5, preventing miRNA's translocation into the cytoplasm. Indeed, when Exp5 is overexpressed, this inhibition is lifted.

Therefore, Dicer, Argonaute and Exp5 are common components in the processing of many sncRNA, not restricted to only miRNAs. And because the processing of these sncRNAs are so intimately linked, one cannot attribute the effect of calorie restriction at only the class of miRNA without also studying the effect on the levels of other sncRNAs.

The relationship between senescence and miRNAs is not stringent
In the liver, kidney, and skeletal muscle, there appear to be no significant correlation between the level of Dicer and age.

Although CR does increase the overall level of miRNA, this relationship is not stringent.

Out of the 265 miRNAs that were detectable from the subcutaneous adipose tissue of C57BL/6J mice, only 51% decreased with age while 10% actually increased with age. Furthermore, the majority of these decreases occurs during the first 6 months of the mice's life, and considering the average lifespan of C57BL/6J to be ~800 days, the decrease in miRNAs might be more appropriately correlated to development rather than senescence.

Of the 136 miRNA that was seen to decrease with age, a significant 37% showed no increase at all after CR; while other miRNA increased to a level higher than observed in young tissues.