Caspase-3

Caspase-3 is encoded by the CASP3 gene in humans, located on chromosome 4q35.1. The zymogen consists of a prodomain, a large subunit (~17 kDa), and a small subunit (~12 kDa), with the active site containing a catalytic dyad of cysteine (Cys163) and histidine (His121). Activation occurs through proteolytic cleavage at Asp175, separating the subunits, which then assemble into a heterotetramer (two large and two small subunits) with two active sites. This cleavage is performed by initiator caspases, such as caspase-9 in the intrinsic pathway and caspase-8 in the extrinsic pathway. The active enzyme recognizes substrates with a consensus sequence of Asp-X-X-Asp (DXXD) and cleaves after the aspartate residue. Structural studies have revealed that the active site is a shallow groove that accommodates the tetrapeptide, and the enzyme's specificity is determined by the S1–S4 subsites. Unlike some other caspases, caspase-3 lacks a prodomain and is not involved in inflammatory signaling, focusing exclusively on apoptotic execution.

As an executioner caspase, caspase-3 amplifies the death signal by cleaving a broad array of substrates, including poly(ADP-ribose) polymerase (PARP), inhibitor of caspase-activated DNase (ICAD), and cytoskeletal proteins such as fodrin and gelsolin. Cleavage of ICAD releases CAD (caspase-activated DNase), which enters the nucleus and fragments DNA into nucleosomal units, a hallmark of apoptosis. Caspase-3 also cleaves lamins, leading to nuclear envelope breakdown, and activates other proteases that dismantle the cell. Its activity is essential for developmental processes such as interdigital web regression and neuronal pruning, as demonstrated in Casp3 knockout mice, which exhibit perinatal lethality and brain hyperplasia. In the immune system, caspase-3 is required for the elimination of autoreactive T cells and for the cytotoxic effects of natural killer cells and cytotoxic T lymphocytes. Furthermore, caspase-3 participates in non-apoptotic functions, including cell differentiation, proliferation, and synaptic plasticity, though these roles are less well understood.

Caspase-3 activity is negatively regulated by inhibitor-of-apoptosis proteins (IAPs), particularly XIAP, which binds to the active site and inhibits its activity. IAPs themselves are antagonized by Smac/DIABLO, which is released from mitochondria during apoptosis. Dysregulation of caspase-3 is implicated in numerous diseases: reduced activity contributes to cancer progression and chemoresistance, while excessive activation leads to neurodegeneration in conditions such as Alzheimer's, Parkinson's, and Huntington's diseases. In Alzheimer's, caspase-3 cleaves amyloid precursor protein and tau, potentially promoting plaque formation and neurofibrillary tangles. In ischemic stroke, caspase-3 activation in the penumbra exacerbates neuronal death, making it a therapeutic target. Conversely, some viruses, such as baculoviruses, encode caspase inhibitors (e.g., p35) to evade host apoptosis. Caspase-3 is also a target for drug development; small-molecule inhibitors like Z-DEVD-FMK are used in research, and activators are being explored as anticancer agents. Biomarkers of caspase-3 activity, such as cleaved PARP, are used in diagnostics and drug efficacy studies.

Beyond its canonical role, caspase-3 has several lesser-known facets. It is involved in the differentiation of lens epithelial cells, where its activity is required for enucleation of the lens, yet the cells survive. In erythroid maturation, caspase-3 cleaves proteins to remodel the cell, but the process is not fully apoptotic. Caspase-3 also plays a role in T-cell activation and proliferation, with transient activation observed in proliferating lymphocytes. In the nervous system, it contributes to synaptic scaling and long-term depression, independent of cell death. Additionally, caspase-3 is implicated in the regulation of stem cell pluripotency and reprogramming; its inhibition can enhance induced pluripotent stem cell generation. In plants, a caspase-3-like activity is involved in programmed cell death during development and pathogen response, though the enzymes are structurally distinct. Caspase-3 has also been detected in extracellular vesicles, suggesting a role in intercellular communication. Furthermore, certain bacterial pathogens, such as Shigella, manipulate host caspase-3 to promote their spread, and some viruses encode decoy substrates to inhibit apoptosis.

Apoptosis

Programmed cell death characterized by cell shrinkage, DNA fragmentation, and membrane blebbing.

Zymogen

An inactive precursor of an enzyme that requires proteolytic cleavage to become active.

Caspase

Cysteine-aspartic protease that cleaves proteins after aspartate residues.

IAP

Inhibitor of apoptosis protein that binds and inhibits caspases.

ICAD

Inhibitor of caspase-activated DNase, which when cleaved releases CAD to fragment DNA.