Mitochondrial-Derived Peptides in Energy Metabolism

1 Mitochondrial-derived peptides in energy metabolism
2 Troy L. Merry1,2,*, Alex H. Chan1, Jonathan S. T. Woodhead1,2, Joseph C. Reynolds3,

3 Hiroshi Kumagai3,4,5, Su-Jeong Kim3, Changhan Lee3,6,7,
4 1
Discipline of Nutrition, Faculty of Medical and Health Sciences, The University of
5 Auckland, Auckland, New Zealand.
6 2
Maurice Wilkins Centre for Molecular Biodiscovery, The University of Auckland,
7 Auckland, New Zealand.
8 3
Leonard Davis School of Gerontology, University of Southern California, Los
9 Angeles, CA 90089.
10 4
Japan Society for the Promotion of Science, Tokyo, Japan.
11 5
Graduate School of Health and Sports Science, Juntendo University, Chiba, Japan.
12 6
USC Norris Comprehensive Cancer Center, Los Angeles, CA 90089.
13 7
Biomedical Science, Graduate School, Ajou University, Suwon 16499, Korea.
14
15
16
17 *Corresponding author:
18 Troy Merry
19 Department of Nutrition, School of Medical Sciences
20 The University of Auckland
21 Private Bag 92019
22 Auckland, New Zealand
23 Ph: +64 9 923 9008
24 t.merry@auckland.ac.nz
25
26
27 Running title: MDPs in metabolism
1
Downloaded from journals.physiology.org/journal/ajpendo at Auckland Univ of Tech (156.062.003.011) on August 11, 2020.
28 ABSTRACT
29 Mitochondrial-derived peptides (MDPs) are small bioactive peptides encoded by short
30 open reading frames (sORF) in mitochondrial DNA that do not necessarily have
31 traditional hallmarks of protein-coding genes. To date, eight MDPs have been
32 identified, all of which have been shown to have various cyto- or metabolo-protective
33 properties. The 12S ribosomal RNA (MT-RNR1) gene harbors the sequence for
34 MOTS-c, while the other seven MDPs, [humanin and small humanin-like peptides
35 (SHLP) 1-6] are encoded by the 16S ribosomal RNA gene. Here we review the
36 evidence that endogenous MDPs are sensitive to changes in metabolism, showing that
37 metabolic conditions like obesity, diabetes and aging are associated with lower
38 circulating MDPs. Whereas, in humans, muscle MDP expression is upregulated in
39 response to stress that perturbs the mitochondria like exercise, some mtDNA
40 mutation-associated diseases, and healthy aging, which potentially suggests a tissue-
41 specific response aimed at restoring cellular or mitochondrial homeostasis. Consistent
42 with this, treatment of rodents with humanin, MOTS-c and SHLP2 can enhance
43 insulin sensitivity and offer protection against a range of age-associated metabolic
44 disorders. Further, assessing how mtDNA variants alter the functions of MDPs is
45 beginning to provide evidence that MDPs are metabolic signal transducers in humans.
46 Taken together, MDPs appear to form an important aspect of a retrograde signaling
47 network that communicates mitochondrial status with the wider cell, and to distal
48 tissues, to modulate adaptative responses to metabolic stress. It remains to be fully
49 determined whether the metabolo-protective properties of MDPs can be harnessed
50 into therapies for metabolic disease.
51
52 Key words: mitochondria, aging, mitochondrial derived peptides, MOTS-c, humanin,

53 SHLP, mitokine
2
54 MAIN TEXT
55 Traditionally the identification of protein-coding open reading frames (ORFs) within
56 genomes has focused on sequences which are initiated with an AUG start codon, have
57 conserved homologous amino acid sequences, and are >100 codons in length (2).
58 However, there is growing evidence that short ORFs that lack these classical
59 hallmarks of protein-coding genes can encode biologically active small- or micro-
60 peptides (< 100 amino acids) and can be found in transcripts annotated for larger
61 proteins or long noncoding RNAs (lncRNAs) (5). While efforts are increasing to
62 systematically identify functionally relevant nuclear short ORFs, several small
63 regulatory peptides encoded by mitochondrial genome (mtDNA) short ORFs
64 (collectively termed mitochondrial derived peptides) have been shown to have broad
65 cellular cyto- and metabolo-protective properties (6, 13, 31).
66
67 The mitochondria’s involvement in maintaining cellular homeostasis extends beyond
68 that of energy production to include a regulatory role in a range of processes
69 including immune/inflammatory responses, proteostasis, adaptive stress responses and
70 apoptosis (43). In order to achieve this, the mitochondria have developed extensive
71 retrograde signaling networks to communicate with the nuclear genome, other
72 intracellular organelles, and potentially neighboring cells or organs (43), of which
73 mitochondrial derived peptides (MDPs) appear to form a critical aspect. The first
74 described MDP, humanin, is encoded within the 16S ribosomal RNA gene (MT-
75 RNR2) (15, 30), and more recently the MT-RNR2 gene has been shown to harbor
76 sequences for several additional small-humanin-like peptides (SHLP1-6) (6). Despite
77 the mtDNA containing 10’s-100’s of potential peptide-encoding short ORFs, the only
78 other MDP to have been described is mitochondrial open reading frame of the 12S
3
79 rRNA-c (MOTS-c), a 16 amino acid peptide transcribed from the 12S ribosomal RNA
80 (MT-RNR1) gene that appears to have potent metabolic modulation properties (31). In
81 this review we argue that MDPs are metabolically active peptides by summarizing the
82 evidence that they are endogenously responsive to metabolic stress and can promote
83 an adaptive response to metabolic stressors, focusing on metabolic disease, aging, and
84 exercise (Figure 1).
85
86 Mitochondrial derived peptides in metabolic disease
87 Mitochondrial dysfunction is a key player in the pathophysiology of metabolic
88 diseases including obesity, insulin resistance, type 2 diabetes, and non-alcoholic fatty
89 liver disease (NAFLD) (3). As such, it is not surprising that there is a growing number
90 of studies which have assessed the effect of administering exogenous MDPs (or their
91 analogues) on glucose and lipid metabolism of rodents under metabolically
92 challenging conditions (Table 1). Of the known native MDPs, MOTS-c has most
93 consistently been reported to have metabolo-protective properties in multiple models
94 of metabolic dysfunction (Table 1). Initial reports by Lee et al. (31) showed that the
95 stable overexpression of MOTS-c in cultured cells promotes glucose clearance and
96 lactate accumulation in an AMP-activated protein kinase (AMPK) and sirtuin (SIRT)
97 1 dependent manner. Both of these proteins are nutrient sensors that have the ability
98 to control cellular substrate utilization and energy metabolism in response to
99 metabolic perturbations, and, consistent with this, daily administration of MOTS-c
100 increased glucose tolerance and insulin sensitivity of aged and diet-induced obese
101 mice (31). Whether the enhanced skeletal muscle insulin sensitivity in mice was the
102 direct result of MOTS-c-induced muscle AMPK activation and/or increased energy
103 expenditure, in part, by increased lipid oxidation is not clear (31); two studies suggest
4
104 that MOTS-c may increase the thermogenic capacity of white and brown fat in an
105 AMPK-dependent manner, leading to weight loss (35, 36).
106
107 Systemic MOTS-c treatment has beneficial effects in multiple rodent models of
108 metabolic stress, including attenuating ovariectomy-induced fat accumulation, insulin
109 resistance (36) and bone loss (42), reducing D-galactose-induced peripheral lipid
110 accumulation and mitochondrial dysfunction (32), and downregulating circulating
111 metabolite profiles that are associated with type 2 diabetes and obesity (24).
112 Consistent with these metabolo-protective effects of MOTS-c in rodents, circulating
113 levels in humans have been reported to be reduced with obesity (9), insulin resistance
114 (4), type 2 diabetes (48), chronic kidney disease (33), and endothelial function (47)
115 (Table 2). Therefore, it will be interesting to determine whether restoring circulating
116 MOTS-c levels in patients with metabolic dysfunction can improve clinical outcomes.
117 Indeed, clinical trials on MOTS-c and a MOTS-c analogue are underway with
118 indications for coronary artery disease in patients with type 2 diabetes
119 (NCT04027712) and non-alcoholic hepatic steatosis and obesity (NCT03998514).
120
121 Humanin was first identified in a cDNA library screen derived from a surviving brain
122 fraction of an individual with Alzheimer’s disease. Consistently, humanin is best
123 known for its neuroprotective effects against, in part, by regulating pro-apoptotic
124 pathways including Bax-related proteins (13) and insulin-like growth factor binding
125 protein-3 (IGFBP-3) (17). Circulating levels of humanin have been shown to be
126 reduced in several metabolic disorders (Table 2), including cardiovascular disease
127 (53, 60) and diabetes (48). Interestingly, however, muscles of patients with the
128 mitochondrial mutations that lead to MELAS (mitochondrial encephalomyopathy
5
129 with lactic acidosis and stroke-like episodes) and CPEO (chronic progressive external
130 ophthalmoplegia) have elevated humanin expression (19, 25), which could be a
131 tissue-specific stress response aimed at restoring/repairing mitochondrial homeostasis,
132 as humanin can enhance mitochondrial metabolism under metabolic stress (20, 51),
133 and the expression of humanin in the muscle can increase in response to acute
134 exercise stress (54).
135
136 Several humanin analogues have been manufactured with improved potency and
137 stability, including HNG, which has a glycine-to-serine substitution at position 14,
138 F6AHN with an alanine-to-phenylalanine substitution at position 6 (abrogates IGFBP-
139 3 binding), and HNGF6A, which contains both substitutions. The infusion of
140 humanin, its analogues or SHLP2 centrally (intra-cerebro-ventricularly) enhances
141 insulin sensitivity in rodents as determined by euglycemic-hyperinsulinemic clamp
142 studies (44). Similar effects were observed when F6AHN and HNGF6A were infused
143 peripherally (44), while intraperitoneal injections attenuate high-fat diet-induced
144 increases in fat mass and lipid accumulation, particularly in the liver (12). Humanin
145 may partially act via the activation of hypothalamic STAT3 to promote the
146 suppression of hepatic glucose production. However, this appears to occur in concert
147 with improved peripheral tissue glucose uptake (44). Indeed, under cellular stress,
148 humanin can activate the insulin-stimulated glucose transport pathway, including
149 insulin receptor substrate 1 and Akt (14, 55).
150
151 The observation that a single dose of humanin can lower blood glucose in Zucker
152 diabetic fatty (ZDF) rats by maintaining high insulin levels has led to the hypothesis
153 that humanin may also be insulinotropic (44). Indeed, HNGF6A infusion can increase
6
154 insulin levels during hyperglycaemic clamps in mice, and promote glucose-stimulated
155 insulin secretion of murine β-cells by enhancing the sensitivity of the β-cells to
156 glucose (27). Furthermore, humanin treatment can delay the onset of type 1 diabetes
157 in non-obese diabetic (NOD) mouse (16). Taken together, this suggests that the MDPs
158 MOTS-c, humanin and SHLP2 are metabolically active peptides that respond to
159 metabolic stress (Table 2) and have the potential to modulate insulin sensitivity,
160 secretion, and energy utilisation pathways. The precise molecular networks that
161 underlie these functions are currently active topics of investigation.
162
163 Mitochondrial derived peptides in aging
164 Aging is associated with a progressive loss of cellular homeostasis and resilience
165 increasing susceptibility to multiple chronic diseases, and is at least partially
166 dependent on metabolism at multiple levels as demonstrated by dietary (e.g. dietary
167 restriction), genetic (e.g. insulin/insulin-like signaling), and pharmacological (e.g.
168 rapamycin, metformin) interventions that extend healthy lifespan (34). Mitochondrial-
169 nuclear communication is considered key to cellular fitness and organismal
170 healthspan (43), and while traditionally thought to be primarily mediated by nuclear-
171 encoded proteins, transient molecules, and mitochondrial metabolites, mitochondrial-
172 encoded factors are now also emerging as key players. Notably, under stress
173 conditions, MOTS-c can translocate to the nucleus and regulate adaptive gene
174 expression through interactions with stress-responsive transcription factors and
175 chromatin binding (22). When delivered via intraperitoneal injection in rodents,
176 MOTS-c and other MDPs can exert tissue specific effects, suggesting that they can
177 also cross the extracellular space (31, and Table 1).
178
7
179 The level of MDPs have been reported to be age-related (Table 2). In cross-sectional
180 studies, plasma humanin levels were lower in aged mice (2 mo. vs 13 mo.) and
181 humans [(45-65 vs 65-80 vs 81-110 yrs.)(44); (39 vs 60 yrs.)(1)]. Further, humanin
182 and SHLP2 systemic and tissue levels were lower in older rodents compared to young
183 (6, 44). In contrast, however, within a large cohort of 693 individuals of varying
184 health status and age (21-113 years), Conte et al. (7) reported a strong positive
185 association between age and plasma humanin levels. These differences may be
186 attributed to the larger sample size, greater age range and differing individual
187 characteristics, however the latter cannot be assessed due to limited participant data
188 being reported in the earlier studies (1, 44). Because humanin can extend metabolic
189 healthspan (56), an increase in plasma humanin was interpreted as a hormetic
190 response aimed at improving the ability of cells/tissues to cope with stress (7) and
191 postulated to have beneficial or detrimental effects depending on the levels of
192 response (39). This is compatible with the finding that patients with chronic kidney
193 disease have higher circulating humanin levels, and lower skeletal muscle levels
194 compared to healthy controls (33). As for MOTS-c, in cross-sectional studies,
195 circulating levels were found to be reduced in aged mice (4 mo. vs 32 mo.)(31) and in
196 humans [(18-30 vs 45-55 vs 70-81 yrs.)(8); significant negative correlation (48)].
197 However, while MOTS-c levels in older skeletal muscle from mice are lower (4 mo.
198 vs 32 mo.)(31), levels are increased in aged men (18-30 vs 45-55 vs 70-81 yrs.)(8).
199 This discrepancy may be because 32 mo. old mice from the NIA aged rodent colony
200 represents an end-of-life morbid condition, whereas the human tissue donors were
201 still active and healthy.
202
8
203 Humanin is heavily involved in the GH/IGF-1 axis, one of the most prominent
204 endocrine regulators of aging. GH-deficient Ames mice are long-lived and show
205 higher circulating humanin levels, while short-lived GH-transgenic mice had lover
206 humanin levels compared to their wild-type mice (29). Furthermore, intermittent
207 MOTS-c treatment initiated later in life reversed age-dependent loss of physical
208 capacity and improved aging metabolism in mice (50). Because aging is linked with a
209 progressive decline in mitochondrial function and disruption of metabolic
210 homeostasis (28), MDPs may play a key role in slowing down the rate of aging and
211 delaying the onset of age-related diseases. Therefore, we suggest that an age-
212 dependent decline in MDP expression and/or function may dampen mitochondrial
213 communication, and consequently reduce the cellular capacity to dynamically adapt to
214 insults and the ever-shifting conditions.
215
216
217 Mitochondrial derived peptides in exercise
218 Regular exercise is a therapeutic and preventative measure for most metabolic
219 diseases and increases the activity of the mitochondria to provide energy for the
220 sustained contractile activity of muscle. Mitochondrial-derived peptides, particularly
221 MOTS-c, activate similar signaling pathways to exercise and when administered
222 exogenously promote exercise-like adaptations (22, 31), leading speculation that
223 MOTS-c may be an exercise mitokine. Investigations into exercise and MDPs were
224 relatively scarce. In a pre- vs post-training study design, Gidlund et al. (11) reported
225 that 12-weeks of resistance training in middle-aged pre-diabetic men resulted in an
226 increase in intramuscular humanin expression, however, similar responses were not
227 seen following Nordic walking (11), 8-weeks of aerobic training (plasma measures
9
228 only) (49) or 2-weeks of high intensity interval training (54) in participants that range
229 from young, healthy males (54) to middle-aged individuals with indications of
230 metabolic disease (11, 49). While it is difficult to discern the reasons for the
231 inconsistent effect of exercise training on humanin levels, it is possible that muscle
232 humanin is more responsive to resistance training (11).
233
234 More recently, MOTS-c and humanin have been observed to increase in muscle
235 (11.9-fold) and plasma (1.5 fold) following acute high-intensity cycling exercise in
236 healthy young men (50)(54), while plasma SHLP6 but not SHLP2 concentration also
237 respond to exercise. In support of the hypothesis that skeletal muscle is a source of
238 circulating MDPs during exercise, contraction of isolated mouse muscle rapidly
239 (within 10 min) increases intramuscular humanin and MOTS-c expression (54, and
240 unpublished observations). While it remains to be determined what intracellular
241 signals regulate MDPs during exercise, the rapid increase in levels following the onset
242 of exercise/contraction suggest a suppression of MDP degradation rather than an
243 upregulation of transcription (54). Reactive oxygen species (ROS) increase during
244 exercise and regulate adaptive responses to exercise training (39), and oxidative stress
245 in cell culture promotes MOTS-c translocation to the nucleus and expression (22).
246 The thiols in both humanin and MOTS-c sequences provide a potential mechanism
247 through which oxidative stress may directly alter the stability of the peptides.
248 However, the effect of oxidative and other metabolic stress (metformin and
249 serum/glucose restriction) on MOTS-c levels appear to be AMPK-dependent (22).
250 Since AMPK is a multifunctional and exercise-sensitive cellular energy sensor, it is
251 possible that MDPs may be responsive to multiple signals associated with the change
252 in energy status that occurs with exercise/contraction. While the role and molecular
10
253 targets of endogenously produced MDPs during exercise are yet to be identified,
254 higher doses (15 mg/kg/day as opposed to 5 mg/kg/day used in obesity interventions
255 (31)) of MOTS-c for as little as 2 weeks can increase running capacity of both young
256 and old mice (50). Mechanistically, MOTS-c treatment upregulated glycolytic and
257 protein metabolism markers following exercise, and led to an enrichment of genes
258 associated with protein regulation/metabolism, cellular metabolism, and oxidative
259 stress response, which are largely under the control of heat-shock proteins (50).
260 Therefore, it is tempting to speculate that MOTS-c may regulate adaptive responses to
261 exercise related stress conditions by forming an integral part of the mitochondrial
262 retrograde signaling network activated during exercise (40). In support for MOTS-c
263 potentially acting to facilitate nuclear genomic interactions that lead to enhanced
264 metabolic flexibility (defined by the overall adaptive capacity to a shift in metabolic
265 supply-demand equilibrium in response to a perturbation) (22, 50) as part of the
266 exercise training response, a loss-of-function MOTS-c polymorphism (K14Q, MT-
267 1382A>C) has been linked to increased type 2 diabetes susceptibility in those with
268 low physical activity levels (58).
269
270
271 Genetic variations in mitochondrial DNA and their functional implication for
272 MDPs
273 The human mitochondrial genome encodes 37 genes that are involved in oxidative
274 phosphorylation, ribosomes, and translation within the mitochondria. As such,
275 alteration of mtDNA copy number and mtDNA polymorphisms can affect
276 mitochondrial function and cell metabolism to modify metabolic disease risk (58).
277 Since mtDNA sequences are more varied by ethnicity compared to nuclear DNA
11
278 sequences (41), studying mtDNA polymorphisms provides a unique insight into
279 ethnicity-specific disease risk and functional significance of mtDNA regions. Indeed,
280 mitochondrial genomic association studies are beginning to reveal the influence of
281 mtDNA on metabolic disease, with mitochondrial variants MT-16320 being
282 associated with blood glucose levels, MT-8706 and MT-8898 with waist-hip-ratio
283 (26), and MT-8414 and MT-16189 increasing the risk of type 2 diabetes in an
284 ethnicity specific manner (18, 46), however, the functional effects of the mtDNA
285 polymorphisms remain relatively underexplored.
286
287 Sequence variation in mtDNA may result in differences in the function of classically
288 recognized mtDNA encoded proteins leading to alterations in mitochondrial
289 respiration, reactive oxygen species (ROS) production, mitochondrial matrix pH, and
290 intracellular calcium levels (18, 21). Equally, mtDNA polymorphisms could also
291 impact the function of MDPs. Consistent with this, the MT-2706 variant in the
292 humanin coding short ORF is associated with a decrease in circulating humanin levels
293 and is associated with accelerated cognitive aging (57). More recently it has been
294 recognized that 5-10% of people with East Asian ancestry have a variant (MT-1382)
295 in the MOTS-c coding region which causes a K14Q amino acid replacement in the
296 MOTS-c peptide (10). A meta-analysis of three independent Japanese cohorts (n =
297 27,527) demonstrated that the C allele of MT-1382 variant is associated with an
298 increased risk of type 2 diabetes in males (when adjusted by for age and BMI), but not
299 in females, and that this effect was dependent on physical activity levels (58).
300 Importantly, treating high-fat fed mice with K14Q MOTS-c does not confer the
301 metabolic benefits associated with native MOTS-c administration (31, 58), suggesting
302 that the MT-1382 variant results in inactive endogenous MOTS-c, which contributes
12
303 to increased metabolic disease risk. Therefore, MDP treatment could potentially be a
304 means of preventing metabolic dysfunction associated with this mtDNA variant that
305 inactivates endogenous MDPs.
306
307 Conclusions and future perspectives
308 There is growing evidence that many MDPs are responsive to changes in metabolism
309 in a tissue- and stress-specific manner, and that exogenous humanin and MOTS-c
310 protect against metabolic dysfunction associated with aging and energy imbalance
311 through improving insulin sensitivity and activating energy consuming pathways.
312 However, the underlying mechanisms driving these responses are just beginning to be
313 investigated, and there are still many open questions including how MDP
314 transcription is regulated and how transcripts from the mitochondria are exported and
315 translated. While CNTFR/GP130/WSX-1 and FPRL-1 are putative receptors for
316 humanin (30), establishing whether other MDPs also have extracellular receptors or
317 primarily exert their effects on energy metabolism by intra-cellular interactions (22)
318 will increase our understanding of the biological significance of these peptides. The
319 difficulty of editing the mitochondrial genome has slowed progress in this field,
320 however, naturally occurring genetic variants within the mitochondrial genome
321 (mitochondrial haplotypes) will likely be able to provide causative mechanistic
322 insight into currently identified MDPs and perhaps guide which of the many
323 additional short open reading frames of mtDNA also encode biologically active
324 MDPs. In addition, antagonists such as blocking antibodies to be developed against
325 MDPs will further aid to define their endogenous role in health and disease.
326 Understanding these aspects of MDP regulation will help elucidate how these new
327 players in the mitochondrial retrograde signaling network modulate adaptative
13
328 responses to metabolic stress, and whether they can be harnessed to treat metabolic
329 disease.
330
14
331 ACKNOWLEDGEMENTS
332 This work was funded by a Marsden Fast-start grant (TLM) and TLM is supported by
333 a Rutherford Discovery Fellowship. CL was supported by the NIA (R01AG052258),
334 Ellison Medical Foundation (EMF), AFAR, and the Hanson-Thorell Family.
335
336 AUTHOR CONTRIBUTIONS
337 TLM, AHC, JSTW, JR, HK, SK and CL co-wrote the manuscript.
338
339 AUTHOR CONFLICTS OF INTEREST
340 CL is a consultant and shareholder of CohBar, Inc. All other authors declare no
341 competing interests.
342
343
15
344 REFERENCES
345 1. Bachar AR, Scheffer L, Schroeder AS, Nakamura HK, Cobb LJ, Oh YK,
346 Lerman LO, Pagano RE, Cohen P, and Lerman A. Humanin is expressed in
347 human vascular walls and has a cytoprotective effect against oxidized LDL-induced
348 oxidative stress. Cardiovasc Res 88: 360-366, 2010.
349 2. Basrai MA, Hieter P, and Boeke JD. Small open reading frames: beautiful
350 needles in the haystack. Genome Res 7: 768-771, 1997.
351 3. Bhatti JS, Bhatti GK, and Reddy PH. Mitochondrial dysfunction and
352 oxidative stress in metabolic disorders - A step towards mitochondria based
353 therapeutic strategies. Biochim Biophys Acta Mol Basis Dis 1863: 1066-1077, 2017.
354 4. Cataldo LR, Fernandez-Verdejo R, Santos JL, and Galgani JE. Plasma
355 MOTS-c levels are associated with insulin sensitivity in lean but not in obese
356 individuals. J Investig Med 66: 1019-1022, 2018.
357 5. Chen J, Brunner AD, Cogan JZ, Nunez JK, Fields AP, Adamson B,
358 Itzhak DN, Li JY, Mann M, Leonetti MD, and Weissman JS. Pervasive functional
359 translation of noncanonical human open reading frames. Science 367: 1140-1146,
360 2020.
361 6. Cobb LJ, Lee C, Xiao J, Yen K, Wong RG, Nakamura HK, Mehta HH,
362 Gao Q, Ashur C, Huffman DM, Wan J, Muzumdar R, Barzilai N, and Cohen P.
363 Naturally occurring mitochondrial-derived peptides are age-dependent regulators of
364 apoptosis, insulin sensitivity, and inflammatory markers. Aging (Albany NY) 8: 796-
365 809, 2016.
366 7. Conte M, Ostan R, Fabbri C, Santoro A, Guidarelli G, Vitale G, Mari D,
367 Sevini F, Capri M, Sandri M, Monti D, Franceschi C, and Salvioli S. Human
368 Aging and Longevity Are Characterized by High Levels of Mitokines. J Gerontol A
369 Biol Sci Med Sci 74: 600-607, 2019.
370 8. D'Souza RF, Woodhead JST, Hedges CP, Zeng N, Wan J, Kumagai H,
371 Lee C, Cohen P, Cameron-Smith D, Mitchell CJ, and Merry TL. Increased
372 expression of the mitochondrial derived peptide, MOTS-c, in skeletal muscle of
373 healthy aging men is associated with myofiber composition. Aging (Albany NY) 12:
374 5244-5258, 2020.
375 9. Du C, Zhang C, Wu W, Liang Y, Wang A, Wu S, Zhao Y, Hou L, Ning Q,
376 and Luo X. Circulating MOTS-c levels are decreased in obese male children and
377 adolescents and associated with insulin resistance. Pediatr Diabetes, 2018.
378 10. Fuku N, Pareja-Galeano H, Zempo H, Alis R, Arai Y, Lucia A, and
379 Hirose N. The mitochondrial-derived peptide MOTS-c: a player in exceptional
380 longevity? Aging cell 14: 921-923, 2015.
381 11. Gidlund EK, von Walden F, Venojarvi M, Riserus U, Heinonen OJ,
382 Norrbom J, and Sundberg CJ. Humanin skeletal muscle protein levels increase
383 after resistance training in men with impaired glucose metabolism. Physiol Rep 4,
384 2016.
385 12. Gong Z, Su K, Cui L, Tas E, Zhang T, Dong HH, Yakar S, and
386 Muzumdar RH. Central effects of humanin on hepatic triglyceride secretion. Am J
387 Physiol Endocrinol Metab 309: E283-292, 2015.
388 13. Guo B, Zhai D, Cabezas E, Welsh K, Nouraini S, Satterthwait AC, and
389 Reed JC. Humanin peptide suppresses apoptosis by interfering with Bax activation.
390 Nature 423: 456-461, 2003.
16
391 14. Han K, Jia N, Zhong Y, and Shang X. S14G-humanin alleviates insulin
392 resistance and increases autophagy in neurons of APP/PS1 transgenic mouse. J Cell
393 Biochem 119: 3111-3117, 2018.
394 15. Hashimoto Y, Niikura T, Tajima H, Yasukawa T, Sudo H, Ito Y, Kita Y,
395 Kawasumi M, Kouyama K, Doyu M, Sobue G, Koide T, Tsuji S, Lang J,
396 Kurokawa K, and Nishimoto I. A rescue factor abolishing neuronal cell death by a
397 wide spectrum of familial Alzheimer's disease genes and Abeta. Proceedings of the
398 National Academy of Sciences of the United States of America 98: 6336-6341, 2001.
399 16. Hoang PT, Park P, Cobb LJ, Paharkova-Vatchkova V, Hakimi M, Cohen
400 P, and Lee KW. The neurosurvival factor Humanin inhibits beta-cell apoptosis via
401 signal transducer and activator of transcription 3 activation and delays and
402 ameliorates diabetes in nonobese diabetic mice. Metabolism: clinical and
403 experimental 59: 343-349, 2010.
404 17. Ikonen M, Liu B, Hashimoto Y, Ma L, Lee KW, Niikura T, Nishimoto I,
405 and Cohen P. Interaction between the Alzheimer's survival peptide humanin and
406 insulin-like growth factor-binding protein 3 regulates cell survival and apoptosis.
407 Proceedings of the National Academy of Sciences of the United States of America
408 100: 13042-13047, 2003.
409 18. Jiang W, Li R, Zhang Y, Wang P, Wu T, Lin J, Yu J, and Gu M.
410 Mitochondrial DNA Mutations Associated with Type 2 Diabetes Mellitus in Chinese
411 Uyghur Population. Sci Rep 7: 16989, 2017.
412 19. Kariya S, Hirano M, Furiya Y, Sugie K, and Ueno S. Humanin detected in
413 skeletal muscles of MELAS patients: a possible new therapeutic agent. Acta
414 Neuropathol 109: 367-372, 2005.
415 20. Kariya S, Hirano M, Furiya Y, and Ueno S. Effect of humanin on decreased
416 ATP levels of human lymphocytes harboring A3243G mutant mitochondrial DNA.
417 Neuropeptides 39: 97-101, 2005.
418 21. Kazuno AA, Munakata K, Nagai T, Shimozono S, Tanaka M, Yoneda M,
419 Kato N, Miyawaki A, and Kato T. Identification of mitochondrial DNA
420 polymorphisms that alter mitochondrial matrix pH and intracellular calcium
421 dynamics. PLoS Genet 2: e128, 2006.
422 22. Kim KH, Son JM, Benayoun BA, and Lee C. The Mitochondrial-Encoded
423 Peptide MOTS-c Translocates to the Nucleus to Regulate Nuclear Gene Expression in
424 Response to Metabolic Stress. Cell metabolism 28: 516-524 e517, 2018.
425 23. Kim SJ, Mehta HH, Wan J, Kuehnemann C, Chen J, Hu JF, Hoffman
426 AR, and Cohen P. Mitochondrial peptides modulate mitochondrial function during
427 cellular senescence. Aging (Albany NY) 10: 1239-1256, 2018.
428 24. Kim SJ, Miller B, Mehta HH, Xiao J, Wan J, Arpawong TE, Yen K, and
429 Cohen P. The mitochondrial-derived peptide MOTS-c is a regulator of plasma
430 metabolites and enhances insulin sensitivity. Physiol Rep 7: e14171, 2019.
431 25. Kin T, Sugie K, Hirano M, Goto YI, Nishino I, and Ueno S. Humanin
432 expression in skeletal muscles of patients with chronic progressive external
433 ophthalmoplegia. J Hum Genet 51: 555-558, 2006.
434 26. Kraja AT, Liu C, Fetterman JL, Graff M, Have CT, Gu C, Yanek LR,
435 Feitosa MF, Arking DE, Chasman DI, Young K, Ligthart S, Hill WD, Weiss S,
436 Luan J, Giulianini F, Li-Gao R, Hartwig FP, Lin SJ, Wang L, Richardson TG,
437 Yao J, Fernandez EP, Ghanbari M, Wojczynski MK, Lee WJ, Argos M, Armasu
438 SM, Barve RA, Ryan KA, An P, Baranski TJ, Bielinski SJ, Bowden DW,
439 Broeckel U, Christensen K, Chu AY, Corley J, Cox SR, Uitterlinden AG,
440 Rivadeneira F, Cropp CD, Daw EW, van Heemst D, de Las Fuentes L, Gao H,
17
441 Tzoulaki I, Ahluwalia TS, de Mutsert R, Emery LS, Erzurumluoglu AM, Perry
442 JA, Fu M, Forouhi NG, Gu Z, Hai Y, Harris SE, Hemani G, Hunt SC, Irvin MR,
443 Jonsson AE, Justice AE, Kerrison ND, Larson NB, Lin KH, Love-Gregory LD,
444 Mathias RA, Lee JH, Nauck M, Noordam R, Ong KK, Pankow J, Patki A, Pattie
445 A, Petersmann A, Qi Q, Ribel-Madsen R, Rohde R, Sandow K, Schnurr TM,
446 Sofer T, Starr JM, Taylor AM, Teumer A, Timpson NJ, de Haan HG, Wang Y,
447 Weeke PE, Williams C, Wu H, Yang W, Zeng D, Witte DR, Weir BS, Wareham
448 NJ, Vestergaard H, Turner ST, Torp-Pedersen C, Stergiakouli E, Sheu WH, et
449 al. Associations of Mitochondrial and Nuclear Mitochondrial Variants and Genes
450 with Seven Metabolic Traits. Am J Hum Genet 104: 112-138, 2019.
451 27. Kuliawat R, Klein L, Gong Z, Nicoletta-Gentile M, Nemkal A, Cui L,
452 Bastie C, Su K, Huffman D, Surana M, Barzilai N, Fleischer N, and Muzumdar
453 R. Potent humanin analog increases glucose-stimulated insulin secretion through
454 enhanced metabolism in the beta cell. FASEB J 27: 4890-4898, 2013.
455 28. Lane RK, Hilsabeck T, and Rea SL. The role of mitochondrial dysfunction
456 in age-related diseases. Biochim Biophys Acta 1847: 1387-1400, 2015.
457 29. Lee C, Wan J, Miyazaki B, Fang Y, Guevara-Aguirre J, Yen K, Longo V,
458 Bartke A, and Cohen P. IGF-I regulates the age-dependent signaling peptide
459 humanin. Aging cell 13: 958-961, 2014.
460 30. Lee C, Yen K, and Cohen P. Humanin: a harbinger of mitochondrial-derived
461 peptides? Trends Endocrinol Metab 24: 222-228, 2013.
462 31. Lee C, Zeng J, Drew BG, Sallam T, Martin-Montalvo A, Wan J, Kim SJ,
463 Mehta H, Hevener AL, de Cabo R, and Cohen P. The mitochondrial-derived
464 peptide MOTS-c promotes metabolic homeostasis and reduces obesity and insulin
465 resistance. Cell Metab 21: 443-454, 2015.
466 32. Li Q, Lu H, Hu G, Ye Z, Zhai D, Yan Z, Wang L, Xiang A, and Lu Z.
467 Earlier changes in mice after D-galactose treatment were improved by mitochondria
468 derived small peptide MOTS-c. Biochem Biophys Res Commun 513: 439-445, 2019.
469 33. Liu C, Gidlund EK, Witasp A, Qureshi AR, Soderberg M, Thorell A,
470 Nader GA, Barany P, Stenvinkel P, and von Walden F. Reduced skeletal muscle
471 expression of mitochondrial-derived peptides humanin and MOTS-C and Nrf2 in
472 chronic kidney disease. Am J Physiol Renal Physiol 317: F1122-F1131, 2019.
473 34. Lopez-Otin C, Galluzzi L, Freije JMP, Madeo F, and Kroemer G.
474 Metabolic Control of Longevity. Cell 166: 802-821, 2016.
475 35. Lu H, Tang S, Xue C, Liu Y, Wang J, Zhang W, Luo W, and Chen J.
476 Mitochondrial-Derived Peptide MOTS-c Increases Adipose Thermogenic Activation
477 to Promote Cold Adaptation. Int J Mol Sci 20, 2019.
478 36. Lu H, Wei M, Zhai Y, Li Q, Ye Z, Wang L, Luo W, Chen J, and Lu Z.
479 MOTS-c peptide regulates adipose homeostasis to prevent ovariectomy-induced
480 metabolic dysfunction. J Mol Med (Berl) 97: 473-485, 2019.
481 37. Mangkhang K, Punyapornwithaya V, Tankaew P, Pongkan W,
482 Chattipakorn N, and Boonyapakorn C. Plasma humanin as a prognostic biomarker
483 for canine myxomatous mitral valve disease: a comparison with plasma NT-roBNP.
484 Pol J Vet Sci 21: 673-680, 2018.
485 38. Mehta HH, Xiao J, Ramirez R, Miller B, Kim SJ, Cohen P, and Yen K.
486 Metabolomic profile of diet-induced obesity mice in response to humanin and small
487 humanin-like peptide 2 treatment. Metabolomics 15: 88, 2019.
488 39. Merry TL and Ristow M. Mitohormesis in exercise training. Free Radic Biol
489 Med 98: 123-130, 2016.
18
490 40. Merry TL and Ristow M. Nuclear factor erythroid-derived 2-like 2
491 (NFE2L2, Nrf2) mediates exercise-induced mitochondrial biogenesis and antioxidant
492 response in mice. J Physiol, 2016.
493 41. Miller B, Arpawong TE, Jiao H, Kim SJ, Yen K, Mehta HH, Wan J,
494 Carpten JC, and Cohen P. Comparing the Utility of Mitochondrial and Nuclear
495 DNA to Adjust for Genetic Ancestry in Association Studies. Cells 8, 2019.
496 42. Ming W, Lu G, Xin S, Huanyu L, Yinghao J, Xiaoying L, Chengming X,
497 Banjun R, Li W, and Zifan L. Mitochondria related peptide MOTS-c suppresses
498 ovariectomy-induced bone loss via AMPK activation. Biochemical and biophysical
499 research communications 476: 412-419, 2016.
500 43. Mottis A, Herzig S, and Auwerx J. Mitocellular communication: Shaping
501 health and disease. Science 366: 827-832, 2019.
502 44. Muzumdar RH, Huffman DM, Atzmon G, Buettner C, Cobb LJ, Fishman
503 S, Budagov T, Cui L, Einstein FH, Poduval A, Hwang D, Barzilai N, and Cohen
504 P. Humanin: a novel central regulator of peripheral insulin action. PloS one 4: e6334,
505 2009.
506 45. Oh YK, Bachar AR, Zacharias DG, Kim SG, Wan J, Cobb LJ, Lerman
507 LO, Cohen P, and Lerman A. Humanin preserves endothelial function and prevents
508 atherosclerotic plaque progression in hypercholesterolemic ApoE deficient mice.
509 Atherosclerosis 219: 65-73, 2011.
510 46. Poulton J, Luan J, Macaulay V, Hennings S, Mitchell J, and Wareham
511 NJ. Type 2 diabetes is associated with a common mitochondrial variant: evidence
512 from a population-based case-control study. Hum Mol Genet 11: 1581-1583, 2002.
513 47. Qin Q, Delrio S, Wan J, Jay Widmer R, Cohen P, Lerman LO, and
514 Lerman A. Downregulation of circulating MOTS-c levels in patients with coronary
515 endothelial dysfunction. International journal of cardiology 254: 23-27, 2018.
516 48. Ramanjaneya M, Bettahi I, Jerobin J, Chandra P, Abi Khalil C, Skarulis
517 M, Atkin SL, and Abou-Samra AB. Mitochondrial-Derived Peptides Are Down
518 Regulated in Diabetes Subjects. Front Endocrinol (Lausanne) 10: 331, 2019.
519 49. Ramanjaneya M, Jerobin J, Bettahi I, Bensila M, Aye M, Siveen KS,
520 Sathyapalan T, Skarulis M, Abou-Samra AB, and Atkin SL. Lipids and insulin
521 regulate mitochondrial-derived peptide (MOTS-c) in PCOS and healthy subjects. Clin
522 Endocrinol (Oxf) 91: 278-287, 2019.
523 50. Reynolds J, Lai RW, Woodhead JST, Joly JH, Mitchell CJ, Cameron-
524 Smith D, Lu R, Cohen P, Graham NA, Benayoun BA, Merry TL, and Lee C.
525 MOTS-c is an Exercise-Induced Mitochondrial-Encoded Regulator of Age-Dependent
526 Physical Decline and Muscle Homeostasis. bioRxiv: 2019.2012.2022.886432, 2019.
527 51. Thummasorn S, Apaijai N, Kerdphoo S, Shinlapawittayatorn K,
528 Chattipakorn SC, and Chattipakorn N. Humanin exerts cardioprotection against
529 cardiac ischemia/reperfusion injury through attenuation of mitochondrial dysfunction.
530 Cardiovasc Ther 34: 404-414, 2016.
531 52. Wei M, Gan L, Liu Z, Liu L, Chang JR, Yin DC, Cao HL, Su XL, and
532 Smith WW. Mitochondrial-Derived Peptide MOTS-c Attenuates Vascular
533 Calcification and Secondary Myocardial Remodeling via Adenosine Monophosphate-
534 Activated Protein Kinase Signaling Pathway. Cardiorenal Med 10: 42-50, 2020.
535 53. Widmer RJ, Flammer AJ, Herrmann J, Rodriguez-Porcel M, Wan J,
536 Cohen P, Lerman LO, and Lerman A. Circulating humanin levels are associated
537 with preserved coronary endothelial function. Am J Physiol Heart Circ Physiol 304:
538 H393-397, 2013.
19
539 54. Woodhead JST, D'Souza RF, Hedges CP, Wan J, Berridge MV,
540 Cameron-Smith D, Cohen P, Hickey AJR, Mitchell CJ, and Merry TL. High-
541 intensity interval exercise increases humanin, a mitochondrial encoded peptide, in the
542 plasma and muscle of men. J Appl Physiol (1985), 2020.
543 55. Xu X, Chua CC, Gao J, Chua KW, Wang H, Hamdy RC, and Chua BH.
544 Neuroprotective effect of humanin on cerebral ischemia/reperfusion injury is
545 mediated by a PI3K/Akt pathway. Brain research 1227: 12-18, 2008.
546 56. Yen K, Mehta HH, Kim SJ, Lue Y, Hoang J, Guerrero N, Port J, Bi Q,
547 Navarrete G, Brandhorst S, Lewis KN, Wan J, Swerdloff R, Mattison JA,
548 Buffenstein R, Breton CV, Wang C, Longo V, Atzmon G, Wallace D, Barzilai N,
549 and Cohen P. The mitochondrial derived peptide humanin is a regulator of lifespan
550 and healthspan. Aging (Albany NY) 12: 11185-11199, 2020.
551 57. Yen K, Wan J, Mehta HH, Miller B, Christensen A, Levine ME, Salomon
552 MP, Brandhorst S, Xiao J, Kim SJ, Navarrete G, Campo D, Harry GJ, Longo V,
553 Pike CJ, Mack WJ, Hodis HN, Crimmins EM, and Cohen P. Humanin Prevents
554 Age-Related Cognitive Decline in Mice and is Associated with Improved Cognitive
555 Age in Humans. Sci Rep 8: 14212, 2018.
556 58. Zempo H, Kim S-J, Fuku N, Nishida Y, Higaki Y, Wan J, Yen K, Miller
557 B, Vicinanza R, Miyamoto-Mikami E, Kumagai H, Naito H, Xiao J, Mehta HH,
558 Lee C, Hara M, Patel YM, Setiawan VW, Moore TM, Hevener AL, Sutoh Y,
559 Shimizu A, Kojima K, Kinoshita K, Tanaka K, and Cohen P. A Pro-Diabetogenic
560 mtDNA Polymorphism in the Mitochondrial-Derived Peptide, MOTS-c. bioRxiv:
561 695585, 2019.
562 59. Zhang X, Urbieta-Caceres VH, Eirin A, Bell CC, Crane JA, Tang H,
563 Jordan KL, Oh YK, Zhu XY, Korsmo MJ, Bachar AR, Cohen P, Lerman A, and
564 Lerman LO. Humanin prevents intra-renal microvascular remodeling and
565 inflammation in hypercholesterolemic ApoE deficient mice. Life Sci 91: 199-206,
566 2012.
567 60. Zhloba AA, Subbotina TF, Molchan NS, and Polushin YS. [The level of
568 circulating humanin in patients with ischemic heart disease.]. Klin Lab Diagn 63:
569 466-470, 2018.
570
571
20
572
573 FIGURE LEGENDS
574 Figure 1. Summary of metabolic stressor that modulate MDP expression and in vivo
575 metabolic effects of MDP treatment in rodents.
576
21
Table 1. Metabolic outcomes of exogenous mitochondrial derived peptide treatment
in vivo
Reference Model MDP Dose Response

Aging
Kim et al. (23) Aged mice HNG 5 mg/kg/d ↑ hippocampus AKT and
ERK phosphorylation
Qin et al. (47) Aged mice HNG 4 mg/kg/2x wk ↓ reduced myocardial fibrosis
Yen et al. (56) Aged mice HNG 4 mg/kg/2x wk ↑ metabolic healthspan
↔ lifespan
Lee et al. (31) Aged mice MOTS-c 5 mg/kg/d ↑ insulin sensitivity
Reynolds et al. (50) Aged mice MOTS-c 15 mg/kg/3x wk ↑ lifespan, ↓ aging markers
Exercise
Reynolds et al. (50) Treadmill run MOTS-c 5-15 mg/kg/d ↑ performance
(mice)
Cardiovascular
Oh et al. (45), Zhang ApoE-deficient HNGF6A 0.4 mg/kg/d ↑aortic function, ↓
et al. (59) mice atherogenesis
Wei et al. (52) Rat; vit D3 + MOTS-c 5 mg/kg/d ↓ vascular calcification
nicotine
Metabolic
Han et al. (14) APP/PS1 mice HNG 50-100 ug/kg/d ↓ IRS-1, ↑ AKT
phosphorylation in brain
Lu et al. (35) Cold exposure MOTS-c 5 mg/kg/d ↑ cold adaptation (browning
(mice) WAT and BAT response)
Li et al. (32) D-galactose MOTS-c 10 mg/kg/d ↓ hepatic lipid accumulation
treated mice
Gong et al. (12) DIO mice HNG 2 mg/kg/d ↓ fat mass and tissue lipid, ↑
glucose homeostasis
Mehta et al. (38), DIO mice HNG, SHLP2, 2.5 mg/kg/d ↓ metabolic disease
Kim et al. (24) MOTS-c metabolite signatures in blood
Lee et al. (31) DIO mice MOTS-c 0.5-5 mg/kg/d ↑ insulin sensitivity, ↓ fat
mass and tissue lipid
Kuliawat et al. (27) Rat HNGF6A 0.07 mg/kg/h ↑ GSIS
Cobb et al. (6) Rats and mice SHLP2, SHLP3 ICV 0.16 SHLP2 ↑ insulin sensitivity
ug/kg/min SHLP3 ↑ IL-6 and MCP-1
2 mg/kg/BID
Muzumdar et al. Rat; ICV and HNGF6A 20ug ICV; ↑ insulin sensitivity
(44) IV 0.05 mg/kg/h IV
Diabetic rat HNGF6A 100 ug ↓ blood glucose
Lu et al. (36) Ovariectomy MOTS-c 5 mg/kg/d ↓ fat mass, ↓ lipid, ↑ glucose
mice homeostasis
1
APP/PS1, amyloid precursor protein/presenilin-1; BAT, brown adipose tissue; DIO,
diet induced obesity; GSIS, glucose stimulated insulin secretion; ICV,
intracerebroventricular; HNG/HNGF6A, humanin analogs; IV, intravenous; WAT,
white adipose tissue; ↑, increase; ↓, decrease.
2
Table 2. Endogenous mitochondrial derived peptide (MDP) response to metabolic
stressors in vivo.
Reference Metabolic stress MDP Tissue Effect

Aging
Muzumdar et al. (44), Aging (human) Humanin, MOTS-c Blood 
Bachar et al.(1), D'Souza
et al. (8)
Conte et al. (7) Aging (human) Humanin Blood 
D'Souza et al. (8) Aging (human) MOTS-c Muscle 
Muzumdar et al. (44), Aging (rodent) Humanin, MOTS-c, Blood, muscle, 
Cobb et al. (6), Lee et al. SHLP2 hypothalamus
(31)
Exercise
Woodhead et al. (54) AEx (human) Humanin Muscle, blood ,
TRx (human) Muscle, blood , 
AEx, TRx (human) SHLP2 Blood 
AEx, TRx (human) SHLP6 Blood , 
Reynolds et al. (50) AEx (human) MOTS-c Muscle, blood ,
Gidlund et al. (11) TRx (human) Humanin Muscle, blood ,
Ramanjenaya et al. (49) TRx (human) MOTS-c Blood 
Cardiovascular
Widmer et al. (53), Zhloba Heart disease (human) Humanin Blood 
et al. (60), Qin et al. (47)
Mangkhang et al. (37) Mitral valve disease Humanin Blood 
(canine)
Metabolic disorders
Ramanjaneya et al. (48) Type 2 diabetes Humanin, MOTS-c Blood 
(human)
Cataldo et al. (4) IR (human) MOTS-c Blood 
Obesity (human) MOTS-c Blood 
Du et al. (9) Obesity (human) MOTS-c Blood 
Liu et al. (33) Kidney disease MOTS-c Muscle, blood 
(human) Humanin Muscle, blood ,
Muscle #
Other
Lu et al. (35) Cold exposure (rodent) MOTS-c Blood 
Ramanjaneya et al. (49) Intralipid infusion MOTS-c Blood 

(human)
Kariya et al. (19), Kin et mtDNA-related Humanin Muscle 
al. (25) diseases (human)
AEx, acute exercise bout; Blood, serum or plasma; IR, insulin resistance; TRx,
exercise training; , increase; , decrease; , no change; #, relative to mtDNA.
1

Mitochondrial-Derived Peptides in Energy Metabolism

Uploaded by

Document Information

Original Title

Copyright

Available Formats

Share this document

Share or Embed Document

Sharing Options

Did you find this document useful?

Is this content inappropriate?

Copyright:

Available Formats

Mitochondrial-Derived Peptides in Energy Metabolism

Uploaded by

Copyright:

Available Formats

1 Mitochondrial-derived peptides in energy metabolism

2 Troy L. Merry1,2,*, Alex H. Chan1, Jonathan S. T. Woodhead1,2, Joseph C. Reynolds3,

29 Mitochondrial-derived peptides (MDPs) are small bioactive peptides encoded by short

31 traditional hallmarks of protein-coding genes. To date, eight MDPs have been

38 circulating MDPs. Whereas, in humans, muscle MDP expression is upregulated in

40 mutation-associated diseases, and healthy aging, which potentially suggests a tissue-

41 specific response aimed at restoring cellular or mitochondrial homeostasis. Consistent

43 insulin sensitivity and offer protection against a range of age-associated metabolic

46 Taken together, MDPs appear to form an important aspect of a retrograde signaling

48 tissues, to modulate adaptative responses to metabolic stress. It remains to be fully

49 determined whether the metabolo-protective properties of MDPs can be harnessed

50 into therapies for metabolic disease.

52 Key words: mitochondria, aging, mitochondrial derived peptides, MOTS-c, humanin,

55 Traditionally the identification of protein-coding open reading frames (ORFs) within

59 hallmarks of protein-coding genes can encode biologically active small- or micro-

62 systematically identify functionally relevant nuclear short ORFs, several small

63 regulatory peptides encoded by mitochondrial genome (mtDNA) short ORFs

65 cellular cyto- and metabolo-protective properties (6, 13, 31).

67 The mitochondria’s involvement in maintaining cellular homeostasis extends beyond

68 that of energy production to include a regulatory role in a range of processes

69 including immune/inflammatory responses, proteostasis, adaptive stress responses and

71 retrograde signaling networks to communicate with the nuclear genome, other

72 intracellular organelles, and potentially neighboring cells or organs (43), of which

76 sequences for several additional small-humanin-like peptides (SHLP1-6) (6). Despite

83 an adaptive response to metabolic stressors, focusing on metabolic disease, aging, and

84 exercise (Figure 1).

86 Mitochondrial derived peptides in metabolic disease

87 Mitochondrial dysfunction is a key player in the pathophysiology of metabolic

91 analogues) on glucose and lipid metabolism of rodents under metabolically

93 consistently been reported to have metabolo-protective properties in multiple models

95 stable overexpression of MOTS-c in cultured cells promotes glucose clearance and

96 lactate accumulation in an AMP-activated protein kinase (AMPK) and sirtuin (SIRT)

98 to control cellular substrate utilization and energy metabolism in response to

99 metabolic perturbations, and, consistent with this, daily administration of MOTS-c

105 AMPK-dependent manner, leading to weight loss (35, 36).

108 metabolic stress, including attenuating ovariectomy-induced fat accumulation, insulin

110 accumulation and mitochondrial dysfunction (32), and downregulating circulating

112 Consistent with these metabolo-protective effects of MOTS-c in rodents, circulating

119 (NCT04027712) and non-alcoholic hepatic steatosis and obesity (NCT03998514).

122 fraction of an individual with Alzheimer’s disease. Consistently, humanin is best

128 mitochondrial mutations that lead to MELAS (mitochondrial encephalomyopathy

131 tissue-specific stress response aimed at restoring/repairing mitochondrial homeostasis,

134 exercise stress (54).

138 F6AHN with an alanine-to-phenylalanine substitution at position 6 (abrogates IGFBP-

140 humanin, its analogues or SHLP2 centrally (intra-cerebro-ventricularly) enhances

141 insulin sensitivity in rodents as determined by euglycemic-hyperinsulinemic clamp

143 peripherally (44), while intraperitoneal injections attenuate high-fat diet-induced

149 insulin receptor substrate 1 and Akt (14, 55).

161 underlie these functions are currently active topics of investigation.

163 Mitochondrial derived peptides in aging

165 increasing susceptibility to multiple chronic diseases, and is at least partially

166 dependent on metabolism at multiple levels as demonstrated by dietary (e.g. dietary

167 restriction), genetic (e.g. insulin/insulin-like signaling), and pharmacological (e.g.

169 nuclear communication is considered key to cellular fitness and organismal

171 encoded proteins, transient molecules, and mitochondrial metabolites, mitochondrial-

174 expression through interactions with stress-responsive transcription factors and

189 healthspan (56), an increase in plasma humanin was interpreted as a hormetic

191 postulated to have beneficial or detrimental effects depending on the levels of

194 compared to healthy controls (33). As for MOTS-c, in cross-sectional studies,

201 still active and healthy.

209 progressive decline in mitochondrial function and disruption of metabolic

214 insults and the ever-shifting conditions.

217 Mitochondrial derived peptides in exercise