MOTS-c

Research Overview

MOTS-c serves as a valuable research tool for investigating mitochondrial-nuclear communication and metabolic regulation mechanisms in laboratory settings. This mitochondrial-derived peptide represents a paradigm shift in understanding how mitochondria actively communicate with the nucleus and regulate systemic metabolism through encoded peptide signals. MOTS-c research has expanded rapidly since its initial characterization, encompassing metabolic disease research, aging studies, exercise physiology investigations, and mitochondrial stress response pathway analysis.

The peptide’s designation reflects its genomic origin: Mitochondrial Open reading frame of the 12S rRNA-c. MOTS-c is encoded within the mitochondrial 12S ribosomal RNA gene, a region previously considered non-coding. This discovery revealed that mitochondrial DNA encodes functional regulatory peptides beyond the 13 traditional oxidative phosphorylation proteins, opening new research directions in mitochondrial biology and metabolic regulation.

Laboratory studies investigate MOTS-c’s effects on glucose metabolism, insulin sensitivity, lipid homeostasis, mitochondrial function, and cellular stress responses. Research protocols examine these effects through in vitro cell culture systems (myocytes, adipocytes, hepatocytes), isolated mitochondrial preparations, and in vivo metabolic challenge models. MOTS-c demonstrates particular research interest for its exercise-mimetic properties and potential role in age-related metabolic decline.

Molecular Characteristics

Complete Specifications:

• Molecular Weight: 1,675.9 Da
• Molecular Formula: C₁₀₁H₁₅₂N₂₈O₂₂
• Amino Acid Sequence: Met-Arg-Trp-Gln-Glu-Met-Gly-Tyr-Ile-Phe-Tyr-Pro-Arg-Lys-Leu-Arg
• Single-Letter Sequence: MRWQEMGYIFYPRKLR
• Genomic Origin: Mitochondrial 12S rRNA gene (position varies by species)
• Peptide Classification: Mitochondrial-derived peptide (MDP)
• Net Charge: +4 at physiological pH (highly basic)
• Isoelectric Point: ~11.5
• Appearance: White to off-white lyophilized powder
• Solubility: Water, bacteriostatic water, phosphate buffered saline

The peptide’s 16-amino acid structure contains multiple positively charged residues (3 arginine, 1 lysine) contributing to its highly basic character. The sequence includes two methionine residues, two tyrosine residues providing UV absorbance, and a single tryptophan residue enabling fluorescence-based detection methods. The peptide’s charge distribution and hydrophobic regions facilitate cellular uptake and potential membrane interactions relevant to its biological activities.

MOTS-c demonstrates conservation across mammalian species with some sequence variations. The human sequence shown above represents the primary form investigated in most research. Single nucleotide polymorphisms (SNPs) in the encoding region have been identified in human populations, with the m.1382A>C polymorphism (K14Q variant) showing associations with longevity in certain populations, representing an active research area in aging studies.

Discovery and Mitochondrial-Derived Peptide Biology

MOTS-c belongs to a novel class of bioactive peptides encoded within mitochondrial DNA. The discovery of mitochondrial-derived peptides challenged traditional understanding of mitochondrial genome function and revealed new mechanisms of mitochondrial-nuclear communication:

Historical Context:

• Mitochondrial DNA traditionally considered to encode only 13 oxidative phosphorylation proteins, 22 tRNAs, and 2 rRNAs
• 2012: Discovery of Humanin, the first characterized mitochondrial-derived peptide with cytoprotective properties
• 2015: Identification of MOTS-c as a metabolic regulatory peptide encoded in mitochondrial 12S rRNA
• Subsequent discoveries of additional MDPs including SHLP-1 through SHLP-6 (small humanin-like peptides)

Mitochondrial-Nuclear Communication:
MOTS-c represents a retrograde signaling mechanism where mitochondria communicate functional status to nuclear gene regulation. Research investigates how:

• Mitochondrial genome encodes regulatory peptides responding to metabolic stress
• MDPs translocate from mitochondria to cytoplasm and nucleus
• These peptides influence nuclear gene expression and systemic metabolism
• Polymorphisms in MDP sequences affect individual metabolic phenotypes

This research area provides new perspectives on mitochondrial biology beyond energy production, positioning mitochondria as active signaling organelles regulating cellular and systemic metabolism.

Pharmacokinetic Profile in Research Models

MOTS-c pharmacokinetic characterization informs experimental protocol design:

Administration and Bioavailability:

• Intraperitoneal administration demonstrates systemic distribution in rodent models
• Subcutaneous administration shows bioavailability with sustained effects
• Intravenous administration enables acute pharmacokinetic studies
• Tissue distribution studies show accumulation in metabolically active tissues (muscle, liver, adipose)

Distribution and Elimination:

• Plasma half-life: Approximately 30-45 minutes following IV administration in rodent models
• Rapid clearance from circulation with tissue uptake
• Preferential accumulation in skeletal muscle and metabolically active organs
• Nuclear translocation observed in response to metabolic stress
• Biological effects persist beyond detectable plasma presence

Metabolic Stress Effects:

• Glucose restriction increases endogenous MOTS-c expression
• Exercise increases circulating MOTS-c levels
• Aging associated with decreased MOTS-c levels
• Metabolic challenge modulates MOTS-c tissue distribution

These pharmacokinetic characteristics influence research protocol design, particularly regarding dosing regimens, timing of metabolic assessments, and tissue collection for molecular studies. The observation that metabolic stress regulates endogenous MOTS-c represents an important consideration for mechanism investigations.

Research Applications

Metabolic Regulation and Insulin Sensitivity Studies

MOTS-c serves as a primary research tool for investigating metabolic regulation mechanisms:

Glucose Metabolism Research:

• Investigation of glucose uptake mechanisms in skeletal muscle, adipocytes, and hepatocytes
• Analysis of insulin-independent glucose uptake pathways
• Studies on glucose transporter (GLUT) expression and translocation
• Examination of hepatic glucose production and gluconeogenesis regulation
• Research on glycolysis and glucose oxidation pathway modulation

Insulin Sensitivity Studies:

• Investigation of insulin signaling pathway components (IRS, PI3K/Akt, AS160)
• Analysis of insulin receptor expression and phosphorylation
• Studies on insulin-independent metabolic effects
• Examination of tissue-specific insulin sensitivity enhancement
• Research on insulin resistance reversal mechanisms in diet-induced obesity models

Lipid Metabolism Research:

• Investigation of fatty acid oxidation pathway activation
• Studies on lipid droplet formation and lipolysis regulation
• Analysis of triglyceride accumulation in metabolic tissues
• Examination of adipogenesis and adipocyte differentiation
• Research on brown adipose tissue activation and thermogenesis

Research protocols typically employ:

• Primary cell cultures (myocytes, adipocytes, hepatocytes)
• Glucose uptake assays using radioactive or fluorescent tracers
• Insulin signaling Western blot analysis
• Metabolic flux measurements using Seahorse technology
• In vivo glucose tolerance and insulin sensitivity testing
• Euglycemic-hyperinsulinemic clamp studies in rodent models

Mitochondrial Function and Bioenergetics Research

Given MOTS-c’s mitochondrial origin, substantial research focuses on mitochondrial function:

Mitochondrial Respiration Studies:

• Investigation of oxygen consumption rates and respiratory capacity
• Analysis of individual electron transport chain complex activities
• Studies on ATP production efficiency and coupling
• Examination of proton leak and uncoupling mechanisms
• Research on reserve respiratory capacity under stress conditions

Mitochondrial Biogenesis Research:

• Investigation of PGC-1α pathway activation
• Studies on mitochondrial DNA content and transcription
• Analysis of mitochondrial protein import and assembly
• Examination of mitochondrial network morphology and dynamics
• Research on fission/fusion balance regulation

Oxidative Stress and Antioxidant Defense:

• Investigation of reactive oxygen species (ROS) production
• Studies on antioxidant enzyme expression and activity
• Analysis of oxidative damage markers
• Examination of mitochondrial quality control mechanisms
• Research on mitophagy induction and selective mitochondrial clearance

Laboratory approaches include isolated mitochondrial preparations, high-resolution respirometry, fluorescent ROS detection, transmission electron microscopy for ultrastructural analysis, and mitochondrial DNA quantification.

Exercise Mimetic Effects and Physical Performance Research

MOTS-c demonstrates exercise-mimetic properties making it valuable for exercise physiology research:

Exercise Response Studies:

• Investigation of endogenous MOTS-c regulation by acute and chronic exercise
• Analysis of exercise-induced metabolic adaptations
• Studies on endurance capacity and fatigue resistance
• Examination of muscle fiber type composition changes
• Research on exercise-independent metabolic benefits

AMPK Pathway Research:

• Investigation of AMP-activated protein kinase activation mechanisms
• Studies on upstream AMPK kinases (LKB1, CaMKK)
• Analysis of downstream AMPK targets (ACC, mTOR, PGC-1α)
• Examination of AMPK-independent metabolic effects
• Research on cellular energy sensing mechanisms

Metabolic Adaptation Studies:

• Investigation of metabolic flexibility and substrate utilization
• Studies on training adaptation molecular mechanisms
• Analysis of muscle glycogen content and regulation
• Examination of lactate production and clearance
• Research on VO2max and aerobic capacity determinants

Research protocols employ exercise training models (treadmill running, swimming), exercise capacity testing (time to exhaustion, grip strength), metabolic measurements during and post-exercise, and molecular analysis of exercise-responsive signaling pathways.

Aging and Longevity Research

MOTS-c investigations extend to aging biology and longevity studies:

Age-Related Metabolic Decline:

• Investigation of MOTS-c expression changes across lifespan
• Studies on age-related insulin resistance mechanisms
• Analysis of mitochondrial dysfunction in aging tissues
• Examination of metabolic disease susceptibility with age
• Research on healthspan extension interventions

Genetic Polymorphism Studies:

• Investigation of m.1382A>C SNP (K14Q variant) associations with longevity
• Studies on population-specific genetic variations
• Analysis of polymorphism effects on metabolic phenotypes
• Examination of gene-environment interactions in aging
• Research on personalized metabolic medicine approaches

Cellular Senescence Research:

• Investigation of senescence-associated metabolic dysfunction
• Studies on senolytic and senomorphic effects
• Analysis of inflammatory mediator production (SASP)
• Examination of tissue regeneration capacity with age
• Research on age-related mitochondrial quality decline

Research in this area utilizes aged animal models, caloric restriction paradigms, genetic longevity models, and cellular senescence induction systems to investigate MOTS-c’s role in aging biology.

Obesity and Metabolic Disease Research

MOTS-c serves as a research tool in metabolic disease models:

Diet-Induced Obesity Models:

• Investigation of high-fat diet metabolic consequences
• Studies on adipose tissue expansion and inflammation
• Analysis of ectopic lipid accumulation (liver, muscle)
• Examination of obesity prevention and reversal mechanisms
• Research on weight regulation and energy expenditure

Type 2 Diabetes Research:

• Investigation of diabetic phenotype prevention or reversal
• Studies on beta-cell function and insulin secretion
• Analysis of peripheral insulin resistance mechanisms
• Examination of diabetic complications (nephropathy, neuropathy)
• Research on glucose homeostasis restoration

Metabolic Syndrome Studies:

• Investigation of multi-organ metabolic dysfunction
• Studies on systemic inflammation and metabolic stress
• Analysis of cardiovascular risk factor modulation
• Examination of hepatic steatosis and NAFLD mechanisms
• Research on integrated metabolic regulation

Experimental approaches include high-fat diet feeding protocols, genetic obesity models (ob/ob, db/db mice), metabolic phenotyping (indirect calorimetry, body composition analysis), and comprehensive metabolic profiling (glucose tolerance, insulin sensitivity, lipid panels).

Molecular Mechanism and Gene Expression Studies

Research investigates MOTS-c’s molecular mechanisms of action:

Nuclear Translocation Research:

• Investigation of cellular uptake and localization mechanisms
• Studies on nuclear localization under metabolic stress conditions
• Analysis of nuclear receptor interactions
• Examination of chromatin binding sites
• Research on gene regulatory mechanisms

Transcriptional Regulation Studies:

• Investigation of gene expression changes (RNA-seq, microarray)
• Studies on metabolic gene network coordination
• Analysis of transcription factor activation and recruitment
• Examination of enhancer and promoter region interactions
• Research on epigenetic modifications

Signaling Pathway Analysis:

• Investigation of AMPK activation mechanisms (direct vs. indirect)
• Studies on mTOR pathway interactions
• Analysis of STAT3 pathway activation
• Examination of folate-dependent one-carbon metabolism
• Research on metabolite-gene expression interactions

Molecular approaches include ChIP-seq for chromatin binding analysis, RNA-seq for transcriptome profiling, proteomics for protein expression changes, metabolomics for metabolite profiling, and phosphoproteomics for signaling pathway analysis.

Laboratory Handling and Storage Protocols

Lyophilized Powder Storage:

• Store at -20°C to -80°C in original sealed vial
• Protect from light exposure and moisture
• Desiccated storage environment essential
• Stability data available for 12+ months at -20°C
• Avoid repeated temperature cycling

Reconstitution Guidelines:

• Reconstitute with sterile water, bacteriostatic water (0.9% benzyl alcohol), or appropriate buffer
• Add solvent slowly down vial side to minimize foaming
• Gentle swirling motion recommended (avoid vigorous shaking)
• Allow complete dissolution before use (typically 1-3 minutes)
• Final concentration depends on experimental design (typical range: 0.1-1 mg/mL)
• Final pH should be 6.5-8.0 for optimal stability
• Filter sterilization (0.22 μm) recommended if not using sterile reconstitution

Reconstituted Solution Storage:

• Short-term storage: 4°C for up to 7 days in sterile conditions
• Long-term storage: -20°C or -80°C in single-use aliquots
• Aliquoting strongly recommended to prevent repeated freeze-thaw cycles
• Avoid more than 2-3 freeze-thaw cycles
• Store in polypropylene tubes (peptide can bind to glass surfaces)
• Consider adding carrier protein (0.1% BSA) for dilute solutions

Handling Considerations:

• MOTS-c contains methionine residues susceptible to oxidation
• Minimize air exposure during handling
• Consider adding reducing agents (DTT, BME) for long-term storage experiments if appropriate
• Work in sterile conditions for cell culture applications
• Peptide is relatively stable but follow general peptide handling best practices

Quality Assurance and Analytical Testing

Each MOTS-c batch undergoes comprehensive analytical characterization:

Purity Analysis:

• High-Performance Liquid Chromatography (HPLC): ≥98% purity
• Analytical method: Reversed-phase HPLC with UV detection at 280nm (tyrosine/tryptophan absorption)
• Multiple peak integration ensures accurate purity determination
• Related substance identification and quantification

Structural Verification:

• Electrospray Ionization Mass Spectrometry (ESI-MS): Confirms molecular weight 1,675.9 Da
• MALDI-TOF mass spectrometry: Alternative confirmatory method
• Amino acid analysis: Verifies sequence composition and ratios
• Peptide content determination: Quantifies actual peptide content by weight (typically ≥80%)
• N-terminal sequencing: Confirms sequence accuracy

Contaminant Testing:

• Bacterial endotoxin: <5 EU/mg via Limulus Amebocyte Lysate (LAL) method
• Heavy metals: Below detection limits per USP standards
• Residual solvents: TFA and acetonitrile within ICH guideline limits
• Water content: Karl Fischer titration (<8% for lyophilized powder)
• Bioburden testing: For sterile applications if specified

Stability Testing:

• Accelerated stability studies at elevated temperatures
• Real-time stability data at recommended storage conditions
• Freeze-thaw stability assessment
• Solution stability in various buffers and pH conditions
• Oxidation stability (methionine residues)

Documentation:

• Certificate of Analysis (COA) provided with each batch
• Batch-specific analytical chromatograms and spectra
• Third-party analytical verification available upon request
• Material Safety Data Sheet (MSDS/SDS) available
• Batch-specific QC results traceable by lot number
• Stability data documented for recommended storage conditions

Research Considerations

Experimental Design Factors:

Researchers should consider several factors when designing MOTS-c experiments:

1. Concentration Selection: Published research reports effective concentrations ranging from nanomolar to micromolar depending on experimental model and readout. In vitro cell culture studies typically use 5-50 μM, while in vivo rodent studies employ 0.5-5 mg/kg body weight. Dose-response studies recommended to determine optimal concentrations for specific applications.

2. Temporal Considerations: MOTS-c demonstrates both acute signaling effects (minutes to hours) and chronic adaptive responses (days to weeks). Experimental timing should align with specific research questions. Acute metabolic effects may be observed within 1-4 hours, while transcriptional changes and metabolic adaptations require longer exposure periods.

3. Metabolic State Considerations: MOTS-c effects may be enhanced or modified by metabolic status. Consider conducting experiments under fed vs. fasted conditions, glucose restriction, exercise challenge, or other metabolic perturbations relevant to research questions.

4. Model Selection: Choose appropriate experimental systems based on research objectives:

• In vitro: Primary cells, immortalized cell lines, co-culture systems
• Ex vivo: Isolated tissues, organotypic cultures
• In vivo: Young vs. aged animals, lean vs. obese models, genetic variants

5. Control Groups: Include appropriate controls:

• Vehicle controls (buffer/saline)
• Positive controls (insulin, metformin, exercise training)
• Time-matched controls for chronic studies
• Metabolic challenge without peptide intervention

Mechanism Investigation:

MOTS-c mechanisms of action represent active research areas. Multiple pathways have been implicated:

• AMPK Activation: Primary mechanism proposed, though debate exists regarding direct vs. indirect activation
• Nuclear Translocation: Metabolic stress induces nuclear entry and gene regulation
• Folate Metabolism: Interactions with one-carbon metabolism and ATIC enzyme
• Metabolite Signaling: Effects on cellular metabolite pools (AMP/ATP ratios, NAD+/NADH)
• Mitochondrial Function: Direct effects on respiratory chain efficiency and biogenesis
• Gene Expression: Transcriptional regulation of metabolic gene networks

Careful experimental design with appropriate inhibitors, genetic models, and complementary approaches is essential to establish mechanistic relationships.

Species and Variant Considerations:

MOTS-c sequence shows conservation across mammals with some variations. The m.1382A>C polymorphism (K14Q variant) occurs in human populations and affects function. Consider:

• Species differences when translating findings
• Genetic background effects in human studies
• Polymorphism-specific effects in population studies
• Potential for personalized metabolic medicine approaches

Compliance and Safety Information

Regulatory Status:
MOTS-c is provided as a research chemical for in-vitro laboratory studies and preclinical research only. This product has not been approved by the FDA or other regulatory agencies for human therapeutic use, dietary supplementation, or medical applications. MOTS-c is not listed as a controlled substance.

Intended Use:

• In-vitro cell culture studies and biochemical assays
• In-vivo preclinical research in approved animal models
• Laboratory investigation of metabolic mechanisms
• Academic and institutional research applications
• Mechanistic studies of mitochondrial-nuclear communication

NOT Intended For:

• Human consumption or administration
• Therapeutic treatment or diagnosis
• Dietary supplementation or performance enhancement
• Veterinary therapeutic applications without appropriate oversight
• Clinical applications of any kind

Safety Protocols:
Researchers should follow standard laboratory safety practices when handling MOTS-c:

• Use appropriate personal protective equipment (lab coat, nitrile gloves, safety glasses)
• Handle in well-ventilated areas or biosafety cabinet as appropriate
• Follow institutional biosafety guidelines and chemical hygiene plans
• Dispose of waste according to institutional and local regulations for biological/chemical waste
• Consult Material Safety Data Sheet (MSDS/SDS) for additional safety information
• Follow institutional animal care and use committee (IACUC) approved protocols for in vivo studies

Institutional Requirements:

• Appropriate institutional review board (IRB) or ethics committee approval for human-derived samples
• IACUC protocol approval for animal studies
• Biosafety committee approval if applicable
• Chemical hygiene and laboratory safety training
• Documentation of training and safety protocols

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