Metformin and exercise shown to promote telomere transcription and integrity via AMPK activation: Connection between Progeria, aging, & HIV-1 latency

"Hutchinson-Gilford Progeria Syndrome" by The Cell Nucleus and Aging: Tantalizing Clues and Hopeful Promises. Scaffidi P, Gordon L, Misteli T; https://commons.wikimedia.org/wiki/File:HIV-budding-Color.jpg#/media/File:HIV-budding-Color.jpg. "HIV-budding-Color" by Photo Credit: C. Goldsmith. Content Providers: CDC/ C. Goldsmith, P. Feorino, E. L. Palmer, W. R. McManus.

Metformin and exercise shown to promote telomere transcription and integrity via AMPK activation: Connection between Progeria, aging, & HIV-1 latency

A recently published study in the journal Science Advances in 2016 strikingly demonstrated that activation of AMPK by the anti-diabetic drug metformin and other pharmacological AMPK activators promoted telomere transcription in vitro, leading to a significant upregulation of telomeric repeat–containing RNA (TERRA), molecules that play a critical role in telomere protection [1]. Cycling endurance exercise, which is associated with AMPK activation, also led to increased TERRA levels in vivo in healthy human subjects. Interestingly, telomere length has been shown to be significantly reduced in cells derived from patients with the accelerated aging disorder Hutchinson-Gilford progeria syndrome (HGPS) and telomere shortening in normal cells that occurs during cellular senescence activates progerin production, a toxic protein that leads to an accelerating aging phenotype in children with HGPS. Metformin has also recently been found to alleviate pathological aging in HGPS cells by reducing the levels of progerin and SRSF1, a gene splicing factor that has been shown to increase progerin production. As increased SRSF1 splicing activity also inhibits reactivation of latent HIV-1 (preventing immune system detection and destruction of the virus), this study provides further substantiation that AMPK activation beneficially links normal aging, accelerated aging, and HIV-1 latency, a hypothesis I first proposed and published in 2015 and 2016 [25,26].

Telomeres are specialized regions of repetitive nucleotide sequences located at the ends of eukaryotic chromosomes that protect chromosomal ends from deterioration [2]. However, continuous cell division leads to telomere shortening, impeding the replenishment of tissues and triggering cellular senescence (i.e. cells cease to divide). Although human telomeres shorten with age, telomeres may be lengthened by the enzyme telomerase [2]. Human telomeres are also protected by a protein complex known as shelterin and by telomeric repeat–containing RNA (TERRA), molecules that that partly associate with telomeres and are transcribed from subtelomeric promoters [1].

In the Science Advances study, Diman et al. initially demonstrated binding of nuclear respiratory factor 1 (NRF1) to all predicted NRF1 binding sites in subtelomeric sequences using a LB37 non–small cell lung carcinoma cell line. NRF1 is a transcription factor that plays a critical role in modulating the expression of nuclear-encoded genes necessary for mitochondrial respiration as well as mitochondrial DNA transcription and replication [3]. AMPK activation has also been shown to increase NRF1 expression [4]. The authors also subjected 10 healthy volunteers to a cycling endurance exercise for 45 min to determine if AMPK activation is associated with upregulation of TERRA and PGC-1α (peroxisome proliferator–activated receptor γ coactivator 1α). PGC-1a is transcription factor that is activated during exercise or caloric restriction, acts as a transcriptional coactivator for NRF1, and is also upregulated via AMPK activation [1,5]. Indeed, low (50% VO2 peak) and high (75% VO2 peak) intensity exercise led to a significant increase in phosphorylated acetyl–coenzyme A carboxylase (ACC, a marker of AMPK activation), PGC-1α nuclear translocation (indicating its activation), and TERRA induction (186% for high intensity and 131% for low intensity exercise). Post-exercise (2.5 hours) blood lactate levels, which correlated with AMPK activity, were also significantly correlated with TERRA induction, indicating that AMPK activation promotes telomere transcription [1].          

Additionally, knockdown of NRF1 in Huh-7 cells reduced endogenous TERRA levels by 25 to 45 % whereas NRF1 overexpression stimulated luciferase activity in subtelomeric sequences, indicating that NRF1 plays a role in promoting basal telomere transcription. Furthermore, treatment of Huh-7 cells with phenformin, a biguanide similar to metformin that also activates AMPK, increased ACC phosphorylation, induced nuclear accumulation of PGC-1α , increased TERRA levels (185 to 400% of expression in untreated cells), and also increased subtelomeric luciferase activity, indicating that AMPK activation increases TERRA levels by promoting NRF1 and PGC-1α -mediated telomere transcription [1].  

Most importantly, the authors demonstrated that AMPK activation also induces telomere transcription and TERRA upregulation in healthy non-dividing muscle cells. Fluorescence in situ hybridization (FISH) experiments revealed that myotubes expressed an average of 36 telomeric signals and 38 TERRA foci per nucleus, with an average of 35 TERRA foci colocalizing with telomeres, suggesting that a majority of telomeres may be covered by TERRA [1]. Strikingly, myotube treatment with the AMPK activators metformin, phenformin, or AICAR increased ACC phosphorylation and significantly upregulated TERRA levels by 1.6 to 2.2, providing compelling evidence that pharmacological- or exercise-induced AMPK activation promotes telomere integrity by increasing TERRA levels through induction of telomere transcription [1]. 

As noted above, telomere maintenance has also been shown to play a significant role in the production of progerin, a toxic protein produced in both normal humans and patients diagnosed with HGPS via aberrant alternative splicing of the LMNA gene [6]. Normal lamin A plays a critical role in supporting nuclear architecture and morphology. Lamin A binding to subtelomeric repeats also localizes telomeres to the nuclear periphery and loss of lamin A leads to defects in telomeric heterochromatin, altered nuclear distribution and shortening of telomeres, inefficient processing of dysfunctional telomeres by non-homologus end joining, and increased genomic instability [7]. Indeed, telomere length has been found to be significantly reduced in fibroblasts derived from HGPS patients and a recent study also confirmed that in normal human fibroblasts, progressive telomere damage that occurs during cellular senescence activates progerin production and also leads to extensive changes in alternative splicing of many other genes, highlighting a striking similarity between normal aging and accelerated aging in HGPS patients [6,8].

As the splicing factor SRSF1 has been shown to increase progerin production by promoting the use of a cryptic splice site located in the LMNA gene, metformin was recently shown to significantly reduce the expression of SRSF1 and progerin and also improve the nuclear architecture of HGPS cells, indicating that AMPK activation by metformin beneficially alters gene splicing in HGPS cells by modulating SRSF1, a hypothesis that I first proposed and published in 2014 [9,26]. Also, p32, a splicing-associated protein that is an endogenous inhibitor of SRSF1 and is critical for the maintenance of mitochondrial functionality and oxidative phosphorylation, has also been shown to be essential for rapamycin- or starvation-induced autophagy mediated by ULK1 [10,11]. Because rapamycin, also an AMPK activator in vivo, improves accelerated aging defects in HGPS cells by reducing progerin levels via autophagy induction, AMPK activation likely also beneficially modulates the activity of p32, leading to inhibition of SRSF1 splicing activity and enhancement of mitochondrial functionality [12,13]. Moreover, PGC-1α, which is activated by AMPK and promotes telomere transcription and increased TERRA levels, is downregulated in HGPS cells, leading to significant mitochondrial dysfunction [14]. Methylene blue, which activates AMPK in vivo, was shown to increase PGC-1α levels, induce progerin solubility, and alleviate accelerated aging defects in HGPS cells [14,15].

The inhibition of SRSF1 and the promotion of telomere transcription by metformin via AMPK activation also connects HGPS and telomere integrity with HIV-1 latency. Increased splicing activity of SRSF1 inhibits reactivation of latent HIV-1 residing in infected immune cells, preventing immune system detection and destruction of the virus [16]. Reactivation of latent HIV-1 leads to a reduction in SRSF1 but an increase in p32 activity and bryostatin-1 (a PKC modulator) has been shown to reactivate latent HIV-1 via AMPK activation [16,17]. Interestingly, p32 modulation via AMPK activation may also enhance and stabilize the splicing activities of hnRNPA1, a heteroribonuclear protein that associates with p32, antagonizes the splicing function of SRSF1, prevents splicing of the HIV-1 genome (promoting viral reactivation), and participates in the maintenance and preservation of telomeres [18-21]. hnRNPA1 is also decreased in senescent human fibroblasts and antagonizes cellular senescence and the senescence-associated secretory phenotype (SASP) via increasing SIRT1 expression [22,23]. SIRT1, a histone deacetylase that plays a role in a number of age related diseases and in the extension of lifespan, is also activated by AMPK [5].

The evidence presented in the Science Advances publication further substantiates that AMPK activation represents a central node that connects the therapeutic benefits of chemically distinct compounds in diseases as seemingly dissimilar as HGPS and HIV-1 latency. Indeed, AMPK activators including metformin promote telomere transcription and integrity, decrease the splicing activity of SRSF1 that increases progerin production and prevents latent HIV-1 reactivation, and potentially beneficially modulates the activity of the SRSF1 inhibitor p32 and the ribonucleoprotein hnRNPA1. As AMPK activators (e.g. ionomycin) induce human oocyte activation (giving rise to normal healthy children) and AMPK is localized throughout the entire acrosome in human sperm (likely promoting the acrosome reaction), AMPK activation links normal human aging, Progeria, and HIV-1 latency with the creation of all human life [24-27].

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