A review of Yi et al's paper in PNAS regarding fat cell cold-activated thermogenesis for Harvard BIOS E-10 Biochemistry Journal Club

1. Authors Li Ye, et al
Date: April 17th, 2018 Presenter David Longo
Fat cells directly sense temperature to
activate thermogenesis

2. Details of the Paper
Response to Cold
Adipose Tissue
Figures
Discussion

3. 2 1 -0 -
• Since 2009, adipose tissue research with
respect to thermogenesis has increased
considerably
Released in 2013
• Dana-Farber Cancer Institute
• B.A. Biological Sciences
Tsinghua University
• Ph.D. Harvard University (HMS/DFCI)
(Bruce. M. Spiegelman)
• Post-Doctoral Stanford University
(Karl Deisseroth)
Li Ye
Published in PNAS
• Official scientific journal (Proceedings) of the
National Academy of Sciences.
• Between 1999—2009, PNAS was the second
most cited journal[1]
Details of the paper
Cited 129 times
Rosen, E. D., & Spiegelman, B. M. (2014). What we talk about when we talk about fat.
Cell, 156(1–2), 20–44. https://doi.org/10.1016/j.cell.2013.12.012
[1] "08.2.2009 - Top Ten Most-Cited Journals (All Fields), 1999-2009". Sciencewatch.com.
Thomson Reuters. 2 August 2009. Retrieved 2017-01-08.

4. - 03 331
Key Points
Brown Adipose Tissue
Reacts to norepinephrine
to stimulate
thermogenesis
Does not react to cold
directly
-0 - 0 3 - 0 -
Activates regardless of ß-
AR response
Exhibits normal function
when returned to normal
temperatures
-3
Subcutaneous fat possesses
substantial thermogenic
capacity in response to cold
or ß-AR stimulation

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What happens when we get cold?
(Peripheral Tissues)
Cold -> Sensory Nerves
(Hypothalamus)
SNS -> NE
(Brown / Beige Adipocytes)
GCPR ß-AR Receptors
Norepinephrine
cAMP
PKA CREB
mRNA
UCP1
Adenylyl
Cyclase
Phosphorylation

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How does cell signaling work again?
Cosmo Bio Company, LTD
https://www.cosmobio.com/contents/pmc_20110218.html

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What does UCP1 (thermogenin) do?
Lynes, M. D., & Tseng, Y.-H. (2015). The thermogenic circuit: Regulators of thermogenic
competency and differentiation. Genes & Diseases, 2(2), 164–172.
https://doi.org/10.1016/J.GENDIS.2015.03.001

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But what does this have to do with class?
Proton ionophore
Uncoupler of ETC
Destroys proton gradient;
No stored energy for ATP;
DOES NOT stop electron flow;
Oxygen still being consumed
Metabolic fuels are consumed
and energy is released as heat
Stored FAs are mobilized to use
as substrates for oxidation
Robin Lynn Hayes, BIOS E-10, Lecture 9, Spring 2018 (Fall 2017)

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But fat is just… fat… right?
Tran, T. T., & Kahn, C. R. (2010). Transplantation of adipose tissue and stem cells: role in metabolism and disease.
Nature Reviews Endocrinology, 6, 195. Retrieved from http://dx.doi.org/10.1038/nrendo.2010.20
“Epididymal”
“VAT”
“Intrascapular”
“BAT”
“Subcutaneous”
“Inguinal”
This Stuff is Different

10. 1 -0 -
But fat is just… fat… right?
Tran, T. T., & Kahn, C. R. (2010). Transplantation of adipose tissue and stem cells: role in metabolism and disease.
Nature Reviews Endocrinology, 6, 195. Retrieved from http://dx.doi.org/10.1038/nrendo.2010.20

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Is it all fight-or-flight?
ß-less ↷ BAT
Mice that lack all three adrenergic receptors ( less mice)
have been shown to have severely impaired thermogenesis in their
inter- scapular BAT in response to cold
DIRECT
EFFECT
What if cells could respond
autonomously to cold?
White / Beige
Interestingly, and in contrast, the induction of thermogenic genes
in the s.c. (inguinal) fat in the β-less mice was largely preserved on
cold exposure, suggesting there is a β-AR–independent pathway
that controls thermogenic gene expression in the s.c. fat.

12. 1 0 - --
Cold Treatment Protocol
Mice individually housed in 10 C for 20h
Figure 1 – In Vivo Testing (Mice)
CL316243
ß3adrenoceptor agonist
(1mg/kg twice daily)
Cold response in s.c.

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Importantly, raising the temperature to 39 C did
not have the same effect on gene expression,
suggesting that the induction of thermogenic
genes is not a nonspecific response to the stress
of non-physiological temperatures.
Double negatives, oh my!
i.e. IT’S THE COLD, WATSON!
Heat just won’t do.

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Figure 2C
Note the desensitization to cold after 4h
Figure 2 – In Vitro Testing
Figure 2F
Oxygen consumption rate increases
with cold exposure

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Figure 3 – More In Vitro Testing
Cold induces UCP1 / PGC1a
Expression
In various fat cell lines, from various
different regions of the body, cold is
shown to increase UCP1 / PGC1a mRNA
expression

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Figure 4D
DMSO–Dimethyl-Sulfide
(Cryoprotective agent)
H89 – Protein Kinase Inhibitor
Figure 4 – What would we do w/o a Western Blot?
Figure 4C
This Western Blot shows an increase
in phosphorylation of CREB at serine
133 due to NE treatment, but not due
to cold.
Figure 4A/4B
More confirmation that UCP1 is more
relatively expressed due to NE
stimulation and cold, but not NE in ß-
less adipocytes

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Methods - PCR
http://ib.bioninja.com.au/standard-level/topic-3-genetics/35-genetic-modification-and/pcr.html

18. 1 -0 -
Methods - qPCR
S. Jähnichen, Wikipedia
Total RNA from cultured cells or tissues was isolated using the TRIzol method (Invitrogen) combined with Qiagen RNAEasy minicolumns, according to
the manufacturer’s instructions. For quantitative PCR (qPCR) analysis, RNA was reverse transcribed, using the ABI high- capacity cDNA synthesis kit,
and was used in quantitative PCR reactions containing SYBR-green fluorescent dye (ABI). Relative expression of mRNAs was determined after
normalization with TATA-binding protein (TBP) levels using the ΔΔCt method.

19. References
Ye, L., Wu, J., Cohen, P., Kazak, L., Khandekar, M. J., Jedrychowski, M. P., …
Spiegelman, B. M. (2013). Fat cells directly sense temperature to activate
thermogenesis. Proceedings of the National Academy of Sciences of the
United States of America, 110(30), 12480–12485.
http://doi.org/10.1073/pnas.1310261110

20. Authors Li Ye, et al
Class BIOS E-10 Presenter David Longo
Thanks
For Listening