GO:1990442 intrinsic apoptotic signaling pathway in response to nitrosative stress: Mechanism, Genes, Functions and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:1990442 describes the intracellular signaling cascade that triggers apoptosis in response to nitrosative stress, a condition caused by high levels of nitric oxide (NO) or peroxynitrite.
• Nitrosative stress activates the intrinsic (mitochondrial) apoptotic pathway, involving BAX/BAK, cytochrome c release, and caspase-9 activation.
• This pathway is distinct from extrinsic apoptosis and is often studied in beta-cell diabetes models and muscle atrophy research.
• Key regulators include TP53, BCL-2 family proteins, and nitric oxide synthases (NOS1, NOS2, NOS3).
• Dysregulation contributes to beta-cell death in diabetes and may influence cancer cell survival under nitrosative conditions.
• CRISPR knockout, knock-in, and overexpression models are essential to dissect gene function in this pathway.
Description
The intrinsic apoptotic signaling pathway in response to nitrosative stress (GO:1990442) is a biological process that triggers programmed cell death via mitochondrial signaling when cells encounter excessive nitric oxide (NO) or peroxynitrite. Nitrosative stress arises from the reaction of NO with superoxide anions, producing peroxynitrite, a highly reactive oxidant that damages cellular components and activates apoptotic cascades. This pathway is critical for understanding how cells respond to nitrosative damage in diseases such as diabetes and neurodegeneration. Researchers study this term to identify molecular targets that modulate cell survival under nitrosative conditions, with implications for therapeutic intervention.
intrinsic apoptotic signaling pathway in response to nitrosative stress At A Glance
| GO ID | GO:1990442 |
|---|---|
| GO term | intrinsic apoptotic signaling pathway in response to nitrosative stress |
| Ontology | biological_process |
| Synonym | nitrosative stress-induced apoptosis; nitrosative stress-induced intrinsic apoptotic signaling pathway |
| Major function | Triggering mitochondrial-dependent apoptosis in response to nitric oxide or peroxynitrite |
| Definition source | QuickGO |
| Related pathways | Intrinsic apoptosis, mitochondrial apoptotic signaling, response to oxidative stress |
What Is GO:1990442?
GO:1990442 defines the series of molecular signals that convey an intracellular signal to trigger apoptotic death, specifically induced by nitrosative stress. This stress results from high levels of nitric oxide (NO) or peroxynitrite, which forms when NO interacts with superoxide anions. The pathway is intrinsic, meaning it acts through mitochondrial mechanisms rather than death receptor signaling.
Why Is intrinsic apoptotic signaling pathway in response to nitrosative stress Important in Cell Biology?
Understanding GO:1990442 is vital because nitrosative stress-induced apoptosis contributes to the pathogenesis of several human diseases, including diabetes mellitus and muscle wasting disorders. In diabetes, beta-cell apoptosis driven by nitrosative stress leads to insulin deficiency. In skeletal muscle, oxidative and nitrosative stress can activate intrinsic apoptosis, and myostatin deficiency protects against this process. Targeting this pathway may offer therapeutic strategies to preserve cell survival in degenerative conditions or to promote cancer cell death.
• Mediates beta-cell death in diabetes mellitus, contributing to insulin deficiency.
• Involved in muscle atrophy and oxidative stress-induced apoptosis in skeletal muscle.
• Nitric oxide and peroxynitrite are key inducers of mitochondrial-dependent apoptosis.
• Provides a mechanism for cellular response to nitrosative damage in inflammation.
• Potential target for protecting beta cells in diabetes.
• Relevant to cancer biology, where nitrosative stress can either promote or inhibit tumor growth.
• Studied in C2C12 myoblasts to understand myostatin regulation of apoptosis.
• Involves BCL-2 family proteins and caspases, offering druggable targets.
• Links nitrosative stress to mitochondrial dysfunction and cell death.
• Can be modeled using CRISPR gene editing to dissect gene function.
What Happens During intrinsic apoptotic signaling pathway in response to nitrosative stress?
Induction by Nitric Oxide and Peroxynitrite
In simple terms: Excess nitric oxide reacts with superoxide to form peroxynitrite, which damages cells and starts the death signal.
Nitrosative stress is induced by high levels of nitric oxide (NO) or peroxynitrite, a product of NO and superoxide anions. This stress can occur during inflammation, ischemia-reperfusion, and neurodegenerative conditions. Peroxynitrite modifies proteins and lipids, leading to mitochondrial dysfunction and activation of apoptotic signaling.
Mitochondrial Outer Membrane Permeabilization
In simple terms: The mitochondria become leaky, releasing factors that trigger cell death.
Following nitrosative stress, pro-apoptotic BCL-2 family proteins such as BAX and BAK are activated, leading to mitochondrial outer membrane permeabilization (MOMP). This allows the release of cytochrome c and other apoptogenic factors from the intermembrane space into the cytosol.
Caspase Activation and Apoptosome Formation
In simple terms: Released cytochrome c builds a death machine that activates caspases, the executioners of apoptosis.
Cytochrome c binds to APAF1, forming the apoptosome, which recruits and activates caspase-9. Active caspase-9 then cleaves effector caspases-3 and -7, leading to the biochemical and morphological hallmarks of apoptosis.
Regulation by BCL-2 Family Proteins
In simple terms: A balance between pro-death and pro-survival proteins decides whether the cell lives or dies.
The BCL-2 family includes anti-apoptotic proteins (BCL-2, BCL-XL, MCL-1) and pro-apoptotic proteins (BAX, BAK, BID, BIM, PUMA). Nitrosative stress can tip the balance toward apoptosis by activating pro-apoptotic members or inhibiting anti-apoptotic ones. For example, peroxynitrite can inactivate BCL-2 via nitration, promoting apoptosis.
Cross-talk with Other Stress Pathways
In simple terms: Nitrosative stress can also trigger other stress responses that feed into apoptosis.
Nitrosative stress can activate JNK and p38 MAPK pathways, which phosphorylate and regulate BCL-2 family proteins. Additionally, endoplasmic reticulum stress and DNA damage responses may intersect with the intrinsic apoptotic pathway under nitrosative conditions.
Key Genes Involved in GO:1990442 intrinsic apoptotic signaling pathway in response to nitrosative stress
The following genes and proteins are central to the intrinsic apoptotic signaling pathway in response to nitrosative stress, based on published literature.
| Gene | Major Role | Research Relevance |
|---|---|---|
| TP53 | Tumor suppressor, transcriptionally activates pro-apoptotic genes | Mutated in many cancers, affects nitrosative stress response |
| BAX | Pro-apoptotic BCL-2 family member, mediates MOMP | Knockout models show resistance to nitrosative apoptosis |
| BAK | Pro-apoptotic BCL-2 family member, mediates MOMP | Redundant with BAX, double KO blocks apoptosis |
| BCL2 | Anti-apoptotic, inhibits BAX/BAK | Overexpression protects against nitrosative stress |
| CASP9 | Initiator caspase, activates downstream caspases | Essential for apoptosome-mediated apoptosis |
| CASP3 | Executioner caspase, cleaves cellular substrates | Activity assays measure apoptosis |
| APAF1 | Forms apoptosome with cytochrome c | Required for caspase-9 activation |
| CYCS | Cytochrome c, released from mitochondria | Marker of MOMP |
| NOS2 | Inducible nitric oxide synthase, produces NO | Upregulated in inflammation, induces nitrosative stress |
| NOS3 | Endothelial NOS, produces NO | Regulates vascular tone, can contribute to nitrosative stress |
| NOS1 | Neuronal NOS, produces NO | Involved in neurodegeneration |
| MSTN | Myostatin, negative regulator of muscle growth | Deficiency protects C2C12 cells from oxidative stress-induced apoptosis |
| AKT1 | Survival kinase, inhibits apoptosis | Phosphorylates BAD, blocks intrinsic apoptosis |
| MAPK8 | JNK, stress-activated kinase | Phosphorylates BCL-2 family proteins |
| MAPK14 | p38 MAPK, stress-activated kinase | Regulates apoptosis under nitrosative stress |
| NFKB1 | Transcription factor, regulates survival genes | Can be pro- or anti-apoptotic depending on context |
| PPARG | Nuclear receptor, regulates metabolism | Involved in beta-cell survival |
| INS | Insulin | Beta-cell marker, loss indicates diabetes |
How Is intrinsic apoptotic signaling pathway in response to nitrosative stress Regulated?
The intrinsic apoptotic signaling pathway in response to nitrosative stress is regulated at multiple levels. Nitric oxide synthases (NOS1, NOS2, NOS3) control NO production, which determines the extent of nitrosative stress. The BCL-2 family proteins integrate signals from stress kinases such as JNK and p38 MAPK, which phosphorylate and modulate their activity. Survival pathways like PI3K/AKT can inhibit apoptosis by phosphorylating BAD and preventing MOMP. Additionally, transcription factors such as TP53 and NF-kB regulate the expression of pro- and anti-apoptotic genes in response to nitrosative stress.
intrinsic apoptotic signaling pathway in response to nitrosative stress and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| INS | Diabetes mellitus, beta-cell loss | INS-1 or MIN6 beta-cell knockout of apoptotic genes |
| MSTN | Muscle atrophy, sarcopenia | C2C12 myoblast knockout or overexpression |
| TP53 | Cancer, apoptosis resistance | HCT116 or MCF7 knockout |
| BAX | Cancer, apoptosis resistance | BAX knockout mouse embryonic fibroblasts |
| NOS2 | Inflammation, nitrosative stress | RAW264.7 macrophage knockout |
Diabetes Mellitus
In diabetes, chronic hyperglycemia and inflammation lead to increased nitric oxide production and nitrosative stress, which induces beta-cell apoptosis via the intrinsic pathway. This contributes to progressive loss of insulin secretion. Understanding GO:1990442 may help identify protective strategies for beta cells.
Muscle Wasting and Sarcopenia
Nitrosative stress can trigger apoptosis in skeletal muscle, contributing to muscle atrophy. Myostatin, a negative regulator of muscle growth, may sensitize cells to oxidative stress-induced intrinsic apoptosis, as shown in C2C12 myoblasts. Targeting this pathway could ameliorate muscle loss.
Neurodegenerative Diseases
Nitrosative stress is implicated in neuronal death in Alzheimer's and Parkinson's diseases. Peroxynitrite-mediated damage activates intrinsic apoptosis in neurons, and this pathway may be a therapeutic target.
Cancer
Nitrosative stress can have dual roles in cancer: it may promote apoptosis in tumor cells, but cancer cells often develop resistance by upregulating anti-apoptotic proteins. Modulating this pathway is a potential anticancer strategy.
From intrinsic apoptotic signaling pathway in response to nitrosative stress-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Does gene X regulate nitrosative stress-induced apoptosis? | CRISPR knockout in C2C12 or beta-cell lines |
| Does a point mutation in BAX affect MOMP? | Point mutation knock-in via CRISPR |
| Can overexpression of BCL2 protect against nitrosative stress? | CRISPR-mediated overexpression |
| Does tagging of CYCS affect its release? | Tagged knock-in (e.g., GFP-CYCS) |
| What is the role of TP53 in nitrosative apoptosis? | TP53 knockout in cancer cell lines |
| Does myostatin deficiency alter apoptosis? | MSTN knockout C2C12 cells |
How to Study the intrinsic apoptotic signaling pathway in response to nitrosative stress Process
| Method | What It Measures | Typical Application |
|---|---|---|
| Annexin V/PI flow cytometry | Phosphatidylserine externalization | Quantify apoptosis in cell populations |
| Caspase-3/7 activity assay | Caspase enzymatic activity | Measure apoptosis induction |
| JC-1 staining | Mitochondrial membrane potential | Detect MOMP |
| Cytochrome c immunostaining | Cytochrome c release | Confirm intrinsic pathway activation |
| Nitrotyrosine Western blot | Protein nitration | Assess nitrosative stress |
| RNA-seq | Global gene expression | Identify pathways altered by nitrosative stress |
| CRISPR knockout | Gene function | Test causality of candidate genes |
| Proteomics | Protein modifications and interactions | Discover nitrosative stress targets |
Measuring Apoptosis
Apoptosis can be quantified by flow cytometry using Annexin V/PI staining, caspase-3/7 activity assays, and TUNEL staining. These methods detect phosphatidylserine externalization, caspase activation, and DNA fragmentation, respectively.
Mitochondrial Function Assays
Mitochondrial membrane potential (JC-1 dye) and cytochrome c release (immunostaining or Western blot) are used to assess MOMP. These are key events in the intrinsic pathway.
Nitrosative Stress Detection
Nitric oxide and peroxynitrite levels can be measured using fluorescent probes (DAF-FM, APF) or by detecting nitrotyrosine residues via immunostaining or mass spectrometry.
Gene Expression Analysis
RNA-seq and qPCR can quantify expression of apoptotic genes (BAX, BCL2, CASP3) and NOS isoforms under nitrosative stress conditions.
How CRISPR Can Be Used to Study GO:1990442 intrinsic apoptotic signaling pathway in response to nitrosative stress
Knockout
CRISPR knockout of candidate genes (e.g., BAX, BAK, TP53) is used to determine their requirement for nitrosative stress-induced apoptosis. For example, BAX/BAK double knockout cells are resistant to MOMP and apoptosis.
Point Mutation
Point mutations can be introduced to study specific residues, such as phosphorylation sites in BCL-2 family proteins or nitration targets. This helps dissect signaling mechanisms.
Knock-in
Knock-in of tagged proteins (e.g., GFP-CYCS) allows real-time imaging of cytochrome c release during apoptosis. This provides spatial and temporal resolution.
Overexpression
CRISPR activation (CRISPRa) or cDNA overexpression can be used to test if a gene (e.g., BCL2) protects against nitrosative stress-induced apoptosis.
How EDITGENE Supports intrinsic apoptotic signaling pathway in response to nitrosative stress Research
Researchers studying intrinsic apoptotic signaling pathway in response to nitrosative stress-related genes often need to determine whether a candidate gene is causally involved in the pathway or merely a bystander. EDITGENE provides comprehensive CRISPR services to enable precise genetic manipulation and functional validation.
Contact EDITGENE today to design your custom CRISPR model for intrinsic apoptotic signaling pathway in response to nitrosative stress research.
Frequently Asked Questions About intrinsic apoptotic signaling pathway in response to nitrosative stress
What is GO:1990442?
GO:1990442 is the Gene Ontology term for the intrinsic apoptotic signaling pathway in response to nitrosative stress, a process where nitric oxide or peroxynitrite triggers mitochondrial-dependent cell death.
What genes are involved in intrinsic apoptotic signaling pathway in response to nitrosative stress?
Key genes include TP53, BAX, BAK, BCL2, CASP9, CASP3, APAF1, CYCS, and NOS isoforms.
How is nitrosative stress induced in cells?
Nitrosative stress is induced by high levels of nitric oxide (NO) or peroxynitrite, often generated by inflammatory stimuli or NOS overexpression.
What is the difference between intrinsic and extrinsic apoptosis?
Intrinsic apoptosis is mediated by mitochondria and intracellular stress signals, while extrinsic apoptosis is triggered by death receptors.
What diseases are associated with nitrosative stress-induced apoptosis?
It is linked to diabetes mellitus, muscle atrophy, neurodegenerative diseases, and cancer.
How can I study GO:1990442 in the lab?
Common methods include Annexin V staining, caspase activity assays, mitochondrial membrane potential measurements, and CRISPR knockout models.
What is the role of BCL-2 family proteins in this pathway?
BCL-2 family proteins regulate mitochondrial outer membrane permeabilization; pro-apoptotic BAX/BAK promote it, while anti-apoptotic BCL-2 inhibits it.
Can CRISPR be used to study nitrosative stress-induced apoptosis?
Yes, CRISPR knockout, knock-in, and overexpression models are powerful tools to dissect gene function in this pathway.
What is peroxynitrite and how does it cause apoptosis?
Peroxynitrite is a reactive oxidant formed from NO and superoxide; it damages mitochondria and activates intrinsic apoptosis.
What are the key markers of intrinsic apoptosis?
Cytochrome c release, caspase-9 and caspase-3 activation, and phosphatidylserine externalization are key markers.
Conclusion
GO:1990442 represents a critical cellular response to nitrosative stress, linking nitric oxide and peroxynitrite to mitochondrial apoptosis. Its dysregulation contributes to diabetes, muscle wasting, and neurodegeneration, making it a target for therapeutic intervention. Advances in CRISPR gene editing enable precise dissection of this pathway, and EDITGENE offers comprehensive services to support such research.
References
- 1. Anuradha R et al.. 2014. Apoptosis of beta cells in diabetes mellitus.. DNA Cell Biol 33(11):743-8 PMID: 25093391
- 2. Boyd CS et al.. 2002. Nitric oxide and cell signaling pathways in mitochondrial-dependent apoptosis.. Biol Chem 383(3-4):411-23 PMID: 12033432
- 3. Drysch M et al.. 2021. Myostatin Deficiency Protects C2C12 Cells from Oxidative Stress by Inhibiting Intrinsic Activation of Apoptosis.. Cells 10(7) PMID: 34359850
