Wow that is a very impressive list!
I always thought Telomere length was part of a feedback loop for aging that includes DNA methylation/Endocrine Secretion, Telomere Length/TPE and circulating blood factors and that it might be possible through combination therapy to “reboot” the system for want of better words.
I do believe restoration of Telomere length is important (in particular due to gene expression via TPE) but without addressing the other two things it isn’t going to do that much bar rejuvenate some tissue and encourage some more favorable gene expression.
Parabiosis studies hint that youthful blood factors may have potential to reprogram the body into a younger age state, perhaps exposure to youthful factors eg, via plasma or small molecules for long enough could encourage a return to youthful gene expression? I am hoping Harold might be able to answer that once he begins his HPE study.
No doubt an over simplification of the situation of course.
From: James Watson To: Gerontology Research Group Sent: Monday, 23 March 2015, 7:52Subject: Re: [GRG] are short telomeres a cause or a result of aging?
Dear Walt, Tom, Josh, David, Harold, and others participating in the telomere discussion.
When I read through your comments on telomeres, I did not see much mentioned about the molecular biology
of telomeres that has been discovered in the past 5-10 years.
Here is a short summary of the 15 major molecular mechanisms that affect/regulate telomere length
which have not been discussed much on LA-GRG. (I purposely left off telomerase and hTERC,
which has been discussed at length).
I did not order them in order of importance (all of these discoveries are important).
Instead, I hope each of these major molecular mechanisms controlling telomere length will inspire each
of you to learn more and share with us what you learned. (please share what you learned with me).
I think we must “move on” from a simple discussion of “telomerase activation” to that of a much larger view
of what controls telomeres, which includes long non-coding RNA (TERRA), histone trimethylation (H3K9 and H4K20),
COMPASS (H3K4me3), histone H3K79 trimethylation, histone hyperacetylation, Rb proteins, subtelomeric DNA
methylation (DNMTs), the miR-290 family, Shelterin proteins, Rap-1, SIRT1, Tankyrases, SIRT6, and the ALT
mechanism are all very important aspects of telomere length control.
It is NOT all due to the enzyme telomerease or the p53-mediated DNA-damage response (DDR).
Here are the “top 15” things that I have not seen LA-GRG members discussing much about:
1. Both ends of the chromosome arm are important for lifespan!
Each chromosome arm as two ends – the pericentromeric area and the telomeric area. Both must be silenced
by the formation of heterochromatin. Pericentromeric heterochromatin (pericentromere DNA) and the telomeric
heterochromatin are both equally important in lifespan! Disruptions in histone protein modifications and with DNA
methylation in the pericentromeric region result in chromosomal instability, recombination, and aneuploidy. In old
age, as many as 80% of cells display features of aneuploidy in organs like the liver. Disruptions in histone protein
modifications and with DNA methylation of the subtelomeric regions likewise regulated telomere length and the ALT
pathway. I noted that LA-GRG discussions on telomere length rarely mentions pericentromeric DNA stability. Instability
in this area is a major factor in the development of aneuploidy, cancer, and aging. Let’s include in the discussion of the
“end of the chromosome arm” more information about the “beginning of the chromosome arm”.
Here is an illustration that compares the silencing mechanism of the “end of the chromosome arm” and the “pericentromeric DNA”
Ref for illustration: http://ift.tt/1EO9diB
2. TERRA is important for lifespan – TERRA is a long non coding RNA (lncRNA) that regulates telomerase activity.
I have seen very few mentioning TERRA in the LA-GRG discussions on telomeres and aging.
I hope all of you are reading a lot about TERRA because it is very important.
As of 2012, a total of 14,880 transcripts originating from 9,277 non-coding loci have been discovered in the
human genome for lncRNA. Of these, approximately 5,000 of the human genome lncRNA are NOT
evolutionarily conserved. (This means that these lncRNAs are very recent additions to the molecular
mechanisms by which genes are regulated….i.e. not something found in yeast). Whereas approximately
half of the lncRNAs are evolutionarily conserved (i.e. are ancient), the other half may be the clue to why
we are “human” and why we age at the rate we do and why we have heritable age-related phenotypes.
lncRNAs are transcribed from intergenic and gene-overlapping areas of the human genome. They also
have eons and introns, just like protein-coding genes. They are also regulated by chromatin modifications,
histone tail modifications, DNA methylation, and miRNA. They also undergo post-transcriptional changes
such as splicing out introns and alternative splicing. Unlike protein-coding genes, however, they are not
translated, they are not polyadenylated, and they have no open reading frame. Also, lncRNA “genes” tend
to have a “bias” towards two-exon transcripts. Even more interestingly, lncRNA can regulate protein-coding
genes nearby (called cis-regulatory mechanisms), genes that are a long ways away on other chromosomes
(called trans-regulatory mechanisms), and also by forming circular transcriptions (which regulate protein
coding genes by acting as a “microRNA sponge”). A good example of two lncRNAs that regulates over 900
genes by all three mechanisms (cis-regulatory, trans-regulatory, and microRNA sponge methods) are the
very important lncRNAs, ANRIL and HOTAIR.
TERRA is an acronym, and stands for TElomeric Repeat containing RNA, or TERRA for short.
TERRA functions as a telomerase inhibitor, “jamming” the site where the hTERC template fits into the telomerase enzyme.
If hTERC is considered to be a lncRNA (it is a little short for that),
The production of TERRA is due to the transcription of “unsilenced” telomeres and subtelomeric DNA.
This “unsilencing” occurs with telomeric/subtelomeric histones are not trimethylated and subtelomeric
DNA is not hypermethylated at CpG residues (remember that the telomeric repeats do not have cytosine in them).
Only subtelomeric DNA can be methylated, but both telomeric and subtelomeric histones can be trimethylated.
If this does not occur, then TERRA transcripts are produced from the transcription of telomeric DNA.
This is NOT good. This is why methylation of subtelomeric DNA is so important! This is also why
taking a supplement like TA-65 does not make a “dent” in lifespan or even lengthen telomeres in the
Patten protocol that used TA-65. A very effective way to do something about TERRA is to do what Dean
Ornish did for his low grade prostate cancer group, who participated in the 5-year study at the Preventative
Medicine Institute in Salsalito (see pdf copy of study below). In this study, the control group underwent a 5%
shortening of telomere length, whereas the experimental group lengthened their telomeres by 10%. (That is
a 15% difference between the two groups……without TA-65 or Product B!). Again, please read the references
below on TERRA.
Here is an illustration of the role of TERRA in shortened telomeres:
Illustration reference: http://ift.tt/1bqCZQN
For more information on this, go to:
3. Telomeric Histone H3K9 and H4K20 trimethylation are important for Lifespan
– Two important histone methyltransferases also regulate telomere length.
Histone tail modifications are very important in silencing telomeric DNA, since telomeric DNA is not methylated (there
are no cytosine nucleotides in mammalian telomeric repeats, whereas their are cytosines in subtelomeric DNA).
Both the telomere and the subtelomeric region are epigenetically regulated by histone protein tail modifications.
The lysine side chains on H3 and H4 subunits of the nucleosomes are modified by histone
methyltransferases of the “suppressor of variegation” family, aka SUV. (I like to call them “SUVs” for short).
SUVs maintain H3K9 trimethylation (by SUV39H) and H4K20 trimethylation (by SUV4-20H).
Unless both telomeric and subtelomeric histones are trimethylated by “the SUVs”, the telomeric
DNA is not silenced, thereby allowing for telomere and subtelomeric DNA to be transcribed.
The transcription of telomeric DNA leads to too much TERRA, which inhibits telomerase. This is why
keeping your telomeric DNA silenced is so important! Please folks, it is not up to small exogenous molecules
like astralagous or fish oil! It is up to your “histones” to silence your telomeric DNA! Please wake up!
Transcription of these parts of our DNA is NOT a good thing, except with cell division (mitosis and meiosis).
During mitosis and meiosis, telomeric DNA must be copied, of course.
Here are some articles on how histones silence telomeric DNA.
Here is an illustration that includes H3K9 and H4K20 trimethylation:
4. Mammalian COMPASS-like complex and H3K4 trimethylation are important for Lifespan
– This is a large macromolecular complex that regulates lysine 4 methylation on histone 3 (H3K4)
This particular form of histone trimethylation is associated with the silencing of genes that are near the base of the telomere (i.e. the TPE).
Unlike the H3K9 trimethylation and H4K20 trimethylation described above, there is another very old method by which telomeres
can silence nearby genes, such as the “telomere position effect” or TPE. This involves a macromolecular complex first described
in yeast as COMPASS, which is an acronym for “COMplex of Proteins ASsociated with Set1”. Set1 is a protein that has a “SET”
domain, which is a 130-140 amino acid sequence (i.e motif) in the protein that confers a specific function. In this case, Set1
appears to be critical to the methylation of lysine 4 on histone 3. H3K4 methylation is another method by which telomere-associated
gene silencing can occur. It occurs in both yeast and human telomeres, so this is an evolutionarily conserved method of gene
regulation that is very old. There are many other proteins found in the COMPASS-like complex, including Cps40, Cps60, Rad6,
Dot1, and Bre1. However, not all of these are important for H3K4 trimethylation. It appears that Cps50 and Cps30 are the two
most important proteins in the COMPASS-like complex that regulates H3K4 trimethylation. Here are some articles on the human
COMPASS-like complex. If you really want to know how to do something about aging, please read the articles below on COMPASS.
Here is an illustration of COMPASS
Illustration reference: http://ift.tt/1EO9dPA
5. Telomeric Histone H3K79 trimethylation is important for Lifespan
– this is another site on histone subunit 3 that regulates telomeric silencing
This histone modification “signature” is not associated with the TPE, but with the silencing of the telomeric DNA itself.
By now, I hope that all of you will see how important it is to “silence your telomeric DNA”, even if you don’t like the subject!
Telomeric DNA silencing also requires the trimethylation of a distant site on the histone tail of H3. This site is regulated
by the protein “Dot1” in yeast. There is a Dot1 homolog in human epigenetic silencing of telomeric DNA as well.
Here are some references on this. We must get out of the 1990s and move on to 2015. The scientists have “leap frogged
us” in terms of understanding telomeres!
Here is an illustration of how H3K79 works, via Rad6-Bre1
Illustration reference: http://ift.tt/1bqD2MD
6. Telomeric and subtelomeric histone hyperacetylation in shortening Lifespan
– Histone acetyltransferases (HATs) like p300 can hyperacetylate both telomeric and subtelomeric DNA
H3 and H4 lysine side chains if they loose their trimethylation. Acetylated lysine side chains of histone subgroups H3 and H4
allows for transcription to occur of these areas. Transcription of telomeric DNA means more TERRA. Transcription of
pericentrometic DNA means lots of micro satellite repeats are created, making cancer. This is why keeping heterochromatin/DNA
silent is so important. If you don’t the telomeric and subtelomeric DNA is transcribed, making more TERRA. If you don’t keep the
pericentromeric heterochromatin/DNA silent, you get aneuploidy! This is key. There are many very practical ways to reduce
hyperacetylation of telomeric, subtelomeric, and pericentromeric DNA. One is to activate SIRT6 (see #12 beow). SIRT6 is a
deacetylator of H3K9. Once H3K9 is deacetylated, then it can be trimethylated by SUV39H (see above). A 2nd way to reduce
histone hyperacetylation is to keep your EtOH intake to less than 2 glasses per day. EtOH is metabolized by a two-step process
into acetate, which is used by acetyl-CoA to “turn on” genes by histone acetyltransferases (However, 1 glass per day is good,
due to its hermetic effects). Please, LA-GRG members, lets talk about histone acetylation vs histone deacetylation in regulating
This is a very practical longevity issue that everyone can do!
Here is an illustration of telomeric histone hyperacetylation
Illustration reference: http://ift.tt/1bqD2MF
For more information on this, go to:
7. Rb protein family is important for Lifespan – Rb proteins regulate histone trimethylation.
As you may recall, cellular senescence is the end game for telomere shortening. In order for the “cellular senescence program” to
be triggered, both Rb and P53 pathways must be activated. This must occur before the cyclin-dependent kinase inhibitors
like p16INK4a can trigger cellular senescence. There is a very practical way to deal with this problem – eliminate senescent
cells! This will immediately make your “leukocyte telomere length test” show a “younger age”! Dr. Kirkland and colleagues
just showed us all a “blueprint” for how to eliminate senescent cells with a very practical method involving one drug and one
Here is an illustration of the Rb proteins and their role in telomeric biology
8. Subtelomeric DNA hypermethylation by DNMTs is important for Lifespan
– DNA methyltransferases regulate telomere length via methylation of the subtelomeric region
(telomeres do not have cytosine bases, but the subtelomeric region must be hypermethylated to avoid telomeric recombination,
accelerated telomeric shortening, and the telomere position effect (TPE).
Here is a nice illustration of subtelomeric DNA methylation:
Illustration reference: http://ift.tt/1bqD2MF
For more information on this, go to:
. miR-290 Cluster – microRNAs control Rbl2, which in turn regulates DMNT1, DNMT3a, and DNMT3b expression and resultant telomere recombination.
The miR-290 cluster controls DNA methylation of subtelomeric DNA. The miR-290 cluster includes miR-290,
miR-291-3p, miR-291-5p, miR-292-3p, miR-292-5p, miR-293, miR-294, and miR-295. When cells are transfected with miR-291-3p,
miR-292–3p, miR-294, or miR-295, there is a down regulation in Rbl2 and an up regulation in DNMT3a and DNMT3b expression.
(but not DNMT1 expression). Thus miRNA therapy would be a “viable strategy” for lengthening telomeres, not some ridiculous
supplement like TA-65! The LA-GRG “chat room” never mentions the microRNA regulation of telomere length, however, or how
microRNA could be used to lengthen lifepsan. It would be well for LA-GRG to discuss this miR-290 family, which clearly plays
a role in telomere length regulation. Please bring these microRNAs up in the LA-GRG discussion on telomeres.
Here is an illustration of miR-290 function:
Illustration reference: http://ift.tt/1EO9fXP
10. Shelterin proteins control Lifespan: TRF1, TRF2, RAP1, TIN1, POT1, and others
As all of you know, telomeric DNA forms a loop at its end. The 3D structure of the loop is largely formed by “cap proteins” called Shelterin proteins.
Several of the Shelterin proteins of the Shelterin complex that regulate telomere length by a cis-acting mechanism.
Specifically, TRF1 and POT1 are part of the Shelterin cap that protects the end of the telomere. TRF1
and POT1 are probably regulated epigenetically by histone tail modifications described above (H3K9 and H4K20 trimethylation).
Someone must educate the LA-GRG members of the role of Shelterin proteins in protecting telomeres and preventing telomeric
shortening. This can be done with meditation! Meditation has been shown to increase telomere length dramatically! Most
experts believe this can be done by reducing oxidative stress. Meditation is what the Nobel Laureate, Elizabeth Blackburn,
is advocating to lengthen telomeres. This appears to be the molecular mechanism of how meditation works to lengthen telomeres.
Please, LA-GRG chat room members, please listen to the Nobel Laureate who discovered telomerase!
This is a very practical way that has been scientifically proven to work!
Here is an illustration of the Shelterin protein complex and its role in the telomeric loop:
Here is a nice illustration that shows why the “loop” is so important and why Shelterin proteins are so important:
11. Rap-1 controls Lifespan – The human repressor activator protein (Rap-1) is a critical regulator of telomere length. Rap-1 was first thought to
only be associated with the Shelterin protein, TRF2, but it has been known now for over 10 years that the human Rap-1
complex actually includes TRF2, and a bunch of DNA repair proteins (Rad50, Mre11, PARP1, and Ku86/Ku70). When
Rap-1 mRNA is “knocked down” with siRNA, telomere length increases. The above molecular mechanism is called a
“cis-acting” mechanism of Rap1. However, more recently, a trans-acting molecular mechanism has been discovered
whereas Rap1 can both control telomere length and affect distant gene expression by binding to extratelomeric sites
throughout the genome! These extratelomeric sites where Rap-1 binds includes subtelomeric DNA, but also distant
genes and “non-gene” sites. 70% of the trans-acting binding sites for Rap1 were at genes and 30% were not located
at genes. These “non-gene” binding sites for Rap1 (that are outside of the telomere or subtelomeric region) are likely
to be long non coding RNA “gene” sites. Inhibiting Rap-1 also inhibits these lncRNAs. Thus Rap1 inhibitors is a viable
strategy to lengthen telomeres. Unfortunately, no one on the LA-GRG chat room has been talking about Rap-1 inhibitors.
Why? Just read the articles below to learn more about Rap-1.
Here is an illustration of how Rap-1 regulates telomeres:
Illustration reference: http://ift.tt/1EO9geh
For more information on this, go to:
12. SIRT1 controls telomeres – The NAD-dependet histone deacetylase, SIRT1, has many other roles besides deacetylating lysine side chains
of histone proteins. One of the roles of SIRT1 is its critical role in double-stranded DNA repair by the “high fidelity method”
called “homologous recombination” (HR). If SIRT1 is not adequately expressed (or NAD levels within the nucleus are low),
then the cell has to use the “low fidelity method” of DNA repair called non-homologous end joining, or NHEJ. When SIRT1
is over-expressed, more HR occurs at the centromere, the telomere, and in the chromosome arms between the centromere
and the telomere. This is why Sirtuins are part of the picture. In addition, SIRT1 is a H3K56 deacetylator, but it doesn’t do
this as well as SIRT6 (see section on SIRT6 below – #12). In addition, SIRT1 deacetylates FoxO3, there by preventing
cellular senescence in epithelial cells like the lung alveolar cells. Summary: SIRT1 helps maintain telomere length. Thus,
SIRT1 activators such as supplements, drugs, CR, fasting, exercise, sleep, NAD, NMN, NR, and many other things will help
maintain telomere length. I can supply you with a list of over 30 different SIRT1 activators, but none of them work as well
as sleep, exercise, fasting, CR, etc. Also reducing dietary nicotinamide, activating NAMPT to convert nicotinamide into NMN
with fasting, exercise, sleep, and reducing body fat will also help increase SIRT1 activity, since nicotinamide is a SIRT1-7
inhibitor. These arevery practical solutions to the telomere shortening problem that no one in LA-GRG is talking about!
Please! Start including SIRT1 and SIRT6 in your discussion (BTW, SIRT1, SIRT6, TIN1 and TIN2 all consume NAD).
Here is an illustration of how SIRT1 controls telomeres:
Here is how you can do something about SIRT1
13. Tankyrases control telomeres – Tankyrases are “telomere-specific” forms of Poly-ADP-ribose Polymerases. There are two isoforms of
this “telomere PARP”, tankyrase 1 (TIN1) and tankyrase 2 (TIN2). Tankyrases promote telomere elongation. Tankyrases
are NAD-dependent enzymes. If NAD levels are low in the cell nucleus, then the Tankyrases cannot keep the telomeres
long. You can take IV NAD or you can take oral NR or NMN. This is a very practical way to maintain your telomere length.
If you are poor and cannot afford NAD, NR, or NMN, you can double the level of NAD in your cells with a 2-day fast! You
can also increase your NAD levels by turning out the lights and going to bed at night. You can also increase NAD levels
in your cells by not eating 3 hours before you go to bed. You can also increase NAD levels with exercise. Unfortunately,
Steve Coles did not do any of the above……now he is dead……30-40 years too early! Why don’t all of you discuss more
about TIN1 and TIN2, LA-GRG? Maybe we can find some small molecules that specifically inhibit TIN1 or TIN2. PARP
inhibitors have already been discovered and synthesized. I don’t know if any Tankyrase inhibitors have been discovered.
These tankyrases are voracious NAD hogs! (BTW, tankyrases colocalize to the pericentriolar region during mitosis! Thus
TIN1 and TIN2 are important for both ends of the chromosome arm).
Here is a nice little diagram of Tankyrase-1 and how it affects telomere elongation:
14. SIRT6 regulates telomeres and the TPE – SIRT6 is very important for the Telomere Position Effect (TPE)
Most of the attention to the Sirtuin family has been on SIRT1, which is better understood than any of the other 6
members of the human SIRT family. When it comes to telomeres, however, SIRT6 may be much more important than
SIRT1 (SIRT1 is important for telomeres, but main via its role in double-stranded DNA break repair via the HR method).
SIRT6 knockdown results in premature cellular senescence and also leads to telomere dysfunction. Here is how that works.
Unlike SIRT1, SIRT6 is a histone H3 lysine 9 de-acetylator, whereas SIRT1 and SIRT6 are H3 lysine 56 de-acetylators.
Except during S-phase of the cell cycle, both of these H3 sites must be maintained in a deacetylated state for telomeric
DNA silencing. It has been scientifically proven to play a key role in maintaining telomeric chromatin in the “silenced state”
(i.e. heterochromatin state). At telomeres, SIRT6 deacetylates H3K9 and allows for the association of WRN. WRN is the
protein that is mutated in Werner’s syndrome. We known how important it is to have a good copy of WRN or you will have
accelerated aging. H3K56 deacetylation must occur after the S-phase of the cell cycle. If SIRT6 does not deacetylate
H3K56, then stem cells develop cellular senescence. Thus you will have accelerated aging if you do not express
SIRT6 and have plenty of NAD in the cell nucleus to act as a co-factor for SIRT6. You can “turn on” the gene for SRIT6
by activating SIRT1 (SIRT1 “turns on” the genes for SIRT3 and SIRT6). SIRT6 has been shown to have other mechanisms
of increasing lifespan. SIRT6 also prevents H3K9 acetylation and resultant gene expression of NF-kB dependent genes.
Thus SIRT6 plays a telomere-dependent and a telomere-independent role in lifespan regulation. SIRT6 is a “bona fide”
Here is an illustration of the role of SIRT6 and telomeric DNA deacetylation as well as NF-kB deacetylation.
Here is an illustration of SIRT6 and the TPE
15.ALT mechanism – human telomeres can be lengthened by the “Alternative Lengthening of Telomeres” (ALT) pathway.
This is why 15% of cancers that are still rapidly dividing do not express telomerase. In other words, in these cells, the
ALT pathway is activated. The ALT pathway used to be a mystery. Now it is well-known and the details of its molecular
mechanisms has been elucidated. This is why telomerase activation or inhibition does not make as much of a difference
as you would expect for life span.
Here is a diagram/illustration of the ALT pathway:
There are many other proteins and epigenetic mechanisms involved in telomere length, telomere stability, etc. that I did not cover here.
I am very concerned that LA-GRG discussion groups are are not keeping up with all of the above discoveries about telomeres
that have been made since the discovery of telomerase. Yes, telomerease is very important, but TERRA, H3K9me3, H4K20me3
H3K9Ac, COMPASS, subtelomeric DNA methylation, miR-290, TRF1, TRF2, POT1, Rap-1, SIRT1, TIN1, TIN2, SIRT6, and the
ALT mechanism are very important too!
I do not understand why the LA-GRG discussion group is still focused only on the enzyme telomerase…….this is 20 years behind the times!
Only looking at the enzyme telomerease was truly a cutting edge idea about 20 years ago, but not any longer.
Now it is well known now that telomere length is regulated by the above 15 factors.
Now it is also well known that telomere length is NOT an “aging clock”.
Today, we know that there are much better “aging clocks” that “keep time” much better than any measurement of telomere length.
(For instance, Steve Horvath of UCLA has shown that measuring the differential DNA methylation of just 353 CpG residues
is about 2-3 times more accurate of an “aging clock” than mean telomere length, % short telomeres, or any other telomere metric).
Reversing telomere length does not reverse 99% of the phenotype of aging. (The men in Dean Ornish’s study whose telomeres
were 10% longer still looked 5 years older at the end of the 5 year study!)
I gave a very good lecture at one of the LA-GRG meetings on telomere shortening and how most aging-scientists
had abandoned the pursuit of replicative senescence and are now looking at a much more broader view of
cellular senescence. Cellular senescence can be triggered by many other things besides shortened
telomeres. I even showed the evidence of how cells where all of the telomeres were long
still could undergo senescence due to oxidative stress, due to oncogenic stress, and
due to radiation.
This is why it would be well if LA-GRG discussion groups “move on” and join the scientific community that is looking at more
than just the enzyme that uses the hTERC RNA template and see the bigger picture of what regulates telomere length.
On Mar 17, 2015, at 10:32 AM, Josh Mitteldorf wrote:
Is telomerase production stimulated by exercise or other moderate stressors?
This is a subject that’s just beginning to be explored. The measurements are not easy. I believe exercise has been linked positively to telomerase, Life stressors are a negative, meditation a positive. I don’t know about others. Look at some of the works of Elissa Epel at UCSF.
On Tue, Mar 17, 2015 at 7:15 AM, wrote:
I love your writings, Josh. Thanks for contributing!
Is telomerase production stimulated by exercise or other moderate stressors?