Cell Corpses Research Articles

Innate immune sensing of cell death in disease and therapeutics.

Innate immunity, cell death and inflammation underpin many aspects of health and disease. Upon sensing pathogens, pathogen-associated molecular patterns or damage-associated molecular patterns, the innate immune system activates lytic, inflammatory cell death, such as pyroptosis and PANoptosis. These genetically defined, regulated cell death pathways not only contribute to the host defence against infectious disease, but also promote pathological manifestations leading to cancer and inflammatory diseases. Our understanding of the underlying mechanisms has grown rapidly in recent years. However, how dying cells, cell corpses and their liberated cytokines, chemokines and inflammatory signalling molecules are further sensed by innate immune cells, and their contribution to further amplify inflammation, trigger antigen presentation and activate adaptive immunity, is less clear. Here, we discuss how pattern-recognition and PANoptosome sensors in innate immune cells recognize and respond to cell-death signatures. We also highlight molecular targets of the innate immune response for potential therapeutic development.

Transgenerational reproductive toxicity induced by carboxyl and amino charged microplastics at environmental concentrations in Caenorhabditis elegans: Involvement of histone methylation

Microplastics, recognized as emerging contaminants, are commonly observed to be charged in the environment, potentially exerting toxic effects on various organisms. However, the transgenerational reproductive toxicity and underlying mechanisms of polystyrene (PS), particularly carboxyl-modified PS (PS-COOH) and amino-modified PS (PS-NH2), remain largely unexplored. In this study, the parental generation (P0) of Caenorhabditis elegans was subjected to environmental concentrations (0.1–100 μg/L) of PS, PS-COOH, and PS-NH2, with subsequent generations (F1–F4) cultured under normal conditions. Exposure to PS-NH2 at concentrations of 10–100 μg/L exhibited more pronounced reproductive toxicity compared to PS or PS-COOH, resulting in decreased brood size, egg ejection rate, number of fertilized eggs, and cell corpses per gonad. Similarly, maternal exposure to 100 μg/L of PS-NH2 induced more severe transgenerational reproductive effects in C. elegans. Significant increases in H3 on lysine 4 dimethylation (H3K4me2) and H3 on lysine 9 trimethylation (H3K9me3) levels were observed in the subsequent generation, concurrent with the transgenerational upregulation of set-30 and met-2 following parental exposure to PS, PS-COOH, and PS-NH2. Correlation analyses revealed significant associations between the expression of these genes with the reproductive ability. Molecular docking studies suggested that PS-NH2 exhibited higher affinity for SET-30 and MET-2. Further analysis demonstrated that transgenerational effects on reproduction were absent in set-30(gk315) and met-2(n4256) mutants, highlighting the pivotal role of set-30 and met-2 in mediating the transgenerational effect. This study provides novel insights into the environmental risks associated with negatively and positively charged microplastics.

Phagolysosomes break down the membrane of a non-apoptotic corpse independent of macroautophagy.

Cell corpses must be cleared in an efficient manner to maintain tissue homeostasis and regulate immune responses. Ubiquitin-like Atg8/LC3 family proteins promote the degradation of membranes and internal cargo during both macroautophagy and corpse clearance, raising the question how macroautophagy contributes to corpse clearance. Studying the clearance of non-apoptotic dying polar bodies in Caenorhabditis elegans embryos, we show that the LC3 ortholog LGG-2 is enriched in the polar body phagolysosome independent of membrane association or autophagosome formation. We demonstrate that ATG-16.1 and ATG-16.2, which promote membrane association of lipidated Atg8/LC3 proteins, redundantly promote polar body membrane breakdown in phagolysosomes independent of their role in macroautophagy. We also show that the lipid scramblase ATG-9 is needed for autophagosome formation in early embryos but is dispensable for timely polar body membrane breakdown or protein cargo degradation. These findings demonstrate that macroautophagy is not required to promote polar body degradation, in contrast to recent findings with apoptotic corpse clearance in C. elegans embryos. Determining how membrane association of Atg8/LC3 promotes the breakdown of different types of cell corpses in distinct cell types or metabolic states is likely to give insights into the mechanisms of immunoregulation during normal development, physiology, and disease.

CED-6/GULP and components of the clathrin-mediated endocytosis machinery act redundantly to correctly display CED-1 on the cell membrane in Caenorhabditis elegans.

CED-1 (cell death abnormal) is a transmembrane receptor involved in the recognition of "eat-me" signals displayed on the surface of apoptotic cells and thus central for the subsequent engulfment of the cell corpse in Caenorhabditis elegans. The roles of CED-1 in engulfment are well established, as are its downstream effectors. The latter include the adapter protein CED-6/GULP and the ATP-binding cassette family homolog CED-7. However, how CED-1 is maintained on the plasma membrane in the absence of engulfment is currently unknown. Here, we show that CED-6 and CED-7 have a novel role in maintaining CED-1 correctly on the plasma membrane. We propose that the underlying mechanism is via endocytosis as CED-6 and CED-7 act redundantly with clathrin and its adaptor, the Adaptor protein 2 complex, in ensuring correct CED-1 localization. In conclusion, CED-6 and CED-7 impact other cellular processes than engulfment of apoptotic cells.

Type of stomata and peel structure associated with programmed cell death of senescent spotting in banana

Senescent spotting or brown spot on bananas, is regarded as a form of programmed cell death (PCD). These spots appear in areas where stomata are present on the banana peel, there is no report describing the relationship between these spots and stomata on peel. This research aims to investigate whether senescent spotting in banana is involved in the PCD of stomata on peel. Three banana cultivars, ‘Khai-Kasetsart 2’ (KK2), ‘Dwarf Cavendish’ (CVD), and ‘Hom Thong’ (HT) bananas were examined for senescent spotting. KK2 banana exhibited more severe senescent spotting than CVD banana, while HT bananas did not display brown spot. Electrolyte leakage and DNA degradation increased before the spotting was observed in KK2 and CVD bananas. The peel structure revealed compact cells in CVD and HT bananas, whereas KK2 banana had a loose cell structure. Notably, KK2 banana had a high stomata density correlate with weight loss. Interestingly, the stomata type of KK2 banana was the same as CVD banana, being the typical type, while HT had a sunken type. The spotting zone of KK2 and CVD bananas revealed that stomata were more open than in the healthy zone. Additionally, the outer ground tissue of spotting zone flattened and collapsed, while the healthy zone did not exhibit this flattening and collapsing. Observations of cell morphology in the spotting zone of KK2 banana showed a detached plasma membrane, organelle destruction, and the presence of cell corpse with an absence of cytoplasm clearance. In conclusion, the result provides an evidence that stomata type on the banana peel involve in PCD, resulting in senescent spotting.

Allogeneic "Zombie Cell" as Off-The-Shelf Vaccine for Postsurgical Cancer Immunotherapy.

Allogeneic tumor cell vaccines provide off-the-shelf convenience but lack patient specificity due to heterogeneity in tumor antigens. Here, allogeneic tumor cell corpses are converted into "zombie cells" capable of assimilating heterogeneous tumor by seizing cancer cells and spreading adjuvant infection. This causes pseudo-oncolysis of tumors, transforming them into immunogenic targets for enhanced phagocytosis. It is shown that in postoperative tumor models, localized delivery of premade "zombie cells" through stepwise gelation in resection cavity consolidates tumor surgery. Compared to analogous vaccines lacking "seizing" or "assimilating" capability, "zombie cell" platform effectively mobilizes T cell response against residual tumors, and establishes immunological memory against tumor re-challenge, showing less susceptibility to immune evasion. Despite using allogeneic sources, "zombie cell" platform functions as generalizable framework to produce long-term antitumor immunity in different tumor models, showing comparable effect to autologous vaccine. Together, with the potential of off-the-shelf availability and personalized relevance to heterogenous tumor antigens, this study suggests an alternative strategy for timely therapy after tumor surgery.

Chemokine-like Orion is involved in the transformation of glial cells into phagocytes in different developmental neuronal remodeling paradigms.

During animal development, neurons often form exuberant or inappropriate axons and dendrites at early stages, followed by the refinement of neuronal circuits at late stages. Neural circuit refinement leads to the production of neuronal debris in the form of neuronal cell corpses, fragmented axons and dendrites, and pruned synapses requiring disposal. Glial cells act as predominant phagocytes during neuronal remodeling and degeneration, and critical signaling pathways between neurons and glia are necessary for the execution of phagocytosis. Chemokine-like mushroom body neuron-secreted Orion is essential for astrocyte infiltration into the γ axon bundle leading to γ axon pruning. Here we show a role of orion in debris engulfment and phagocytosis. Interestingly, orion is involved in the overall transformation of astrocytes into phagocytes. In addition, analysis of several neuronal paradigms demonstrates the role of orion in eliminating both peptidergic vCrz+ and PDF-Tri neurons via additional phagocytic glial cells like cortex and/or ensheathing glia. Our results suggest that Orion is essential for phagocytic activation of astrocytes, cortex, and ensheathing glia, and point to Orion as a trigger of glial infiltration, engulfment and phagocytosis.

Defective phagocytosis leads to neurodegeneration through systemic increased innate immune signaling

In nervous system development, disease, and injury, neurons undergo programmed cell death, leaving behind cell corpses that are removed by phagocytic glia. Altered glial phagocytosis has been implicated in several neurological diseases including Alzheimer's disease. To untangle the links between glial phagocytosis and neurodegeneration, we investigated Drosophila mutants lacking the phagocytic receptor Draper. Loss of Draper leads to persistent neuronal cell corpses and age-dependent neurodegeneration. Here we investigate whether the phagocytic defects observed in draper mutants lead to chronic increased immune activation that promotes neurodegeneration. We found that the antimicrobial peptide Attacin-A is highly upregulated in the fat body of aged draper mutants and that the inhibition of the Immune deficiency (Imd) pathway in the glia and fat body of draper mutants led to reduced neurodegeneration. Taken together, these findings indicate that phagocytic defects lead to neurodegeneration via increased immune signaling, both systemically and locally in the brain.

Evaluation of the Reproductive Toxicity of Fluopimomide in Meloidogyne incognita and Caenorhabditis elegans

Fluopimomide is a new pesticide that is widely applied in agriculture; however, the effects and molecular mechanisms of fluopimomide in inhibiting nematode reproduction remain unknown. In this study, the effects of fluopimomide on the development and infection of Meloidogyne incognita and the reproductive toxicity in Caenorhabditis elegans were evaluated. Results showed that, in comparison to inoculated control, fluopimomide at 0.33, 0.67, and 1.0 mg/kg soil significantly (p < 0.05) delayed M. incognita development and decreased the reproduction in pot experiments. Fluopimomide notably reduced the galls index with a control effect of 78.6%, 67.9%, and 50.0%, respectively. In addition, a dose–response relationship existed between the brood size and germ cell number of C. elegans and fluopimomide concentrations. Compared with the control group, fluopimomide at 1.0 and 5.0 mg/L notably (p < 0.001) increased the number of cell corpses per gonad in the N2 strain of C. elegans by 8.8- and 14.4-fold, respectively. The number of cell corpses per gonad was similar between the fluopimomide treated worms and the control group in mutants of ced-3, ced-4, and ced-9. Further evidence revealed fluopimomide significantly enhanced the expression of cep-1, egl-1, and clk-2, while no obvious effects were observed in their mutants. Taken together, these results indicated that fluopimomide inflicted DNA damage and induced the core apoptosis pathway caused by germ-cell apoptosis, leading to the reduction of the brood size of C. elegans.

Endosperm cell death promoted by NAC transcription factors facilitates embryo invasion in Arabidopsis

In flowering plants, two fertilization products develop within the limited space of the seed: the embryo and the surrounding nutritive endosperm. The final size of the endosperm is modulated by the degree of embryo growth. In Arabidopsis thaliana, the endosperm expands rapidly after fertilization, but later gets invaded by the embryo that occupies most of the seed volume at maturity, surrounded by a single remaining aleurone-like endosperm layer.1,2,3,4 Embryo invasion is facilitated by the endosperm-expressed bHLH-type transcription factor ZHOUPI, which promotes weakening of endosperm cell walls.5,6 Endosperm elimination in zou mutants is delayed, and embryo growth is severely affected; the endosperm finally collapses around the dwarf embryo, causing the shriveled appearance of mature zou seeds.5,6,7 However, whether ZHOUPI facilitates mechanical endosperm destruction by the invading embryo or whether an active programmed cell death (PCD) process causes endosperm elimination has been subject to debate.2,8 Here we show that developmental PCD controlled by multiple NAC transcription factors in the embryo-adjacent endosperm promotes gradual endosperm elimination. Misexpressing the NAC transcription factor KIRA1 in the entire endosperm caused total endosperm elimination, generating aleurone-less mature seeds. Conversely, dominant and recessive higher-order NAC mutants led to delayed endosperm elimination and impaired cell corpse clearance. Promoting PCD in the zhoupi mutant partially rescued its embryo growth defects, while the endosperm in a zhoupi nac higher-order mutant persisted until seed desiccation. These data suggest that a combination of cell wall weakening and PCD jointly facilitates embryo invasion by an active auto-elimination of endosperm cells.

Macrophage Nrf 2 the rescue.

The exuberant phagocytosis of apoptotic cell corpses by macrophages in Drosophila embryos creates highly oxidative environments. Stow and Sweet discuss work from Clemente and Weavers (2023. J. Cell Biol.https://doi.org/10.1083/jcb.202203062) showing for the first time how macrophage Nrf2 is primed to help sustain immune function and mitigate bystander oxidative damage.

In vivo imaging of the phagocytic dynamics underlying efficient clearance of adult-born hippocampal granule cells by ramified microglia.

The phagocytosis of dead cells by microglia is essential in brain development and homeostasis. However, the mechanism underlying the efficient removal of cell corpses by ramified microglia remains poorly understood. Here, we investigated the phagocytosis of dead cells by ramified microglia in the hippocampal dentate gyrus, where adult neurogenesis and homeostatic cell clearance occur. Two-color imaging of microglia and apoptotic newborn neurons revealed two important characteristics. Firstly, frequent environmental surveillance and rapid engulfment reduced the time required for dead cell clearance. The motile microglial processes frequently contacted and enwrapped apoptotic neurons at the protrusion tips and completely digested them within 3-6 h of the initial contact. Secondly, while a single microglial process engaged in phagocytosis, the remaining processes continued environmental surveillance and initiated the removal of other dead cells. The simultaneous removal of multiple dead cells increases the clearance capacity of a single microglial cell. These two characteristics of ramified microglia contributed to their phagocytic speed and capacity, respectively. Consistently, the cell clearance rate was estimated to be 8-20 dead cells/microglia/day, supporting the efficiency of removing apoptotic newborn neurons. We concluded that ramified microglia specialize in utilizing individual motile processes to detect stochastic cell death events and execute parallel phagocytoses.

Long-term exposure to 6-PPD quinone reduces reproductive capacity by enhancing germline apoptosis associated with activation of both DNA damage and cell corpse engulfment in Caenorhabditis elegans

Recently, 6-PPD quinone (6-PPDQ), a derivative of tire antioxidant 6-PPD, was reported to have acute toxicity for organisms. However, the possible reproductive toxicity of 6-PPDQ is still largely unclear. In this study, the reproductive toxicity of 6-PPDQ after long-term exposure was further investigated in Caenorhabditis elegans. Exposure to 1 and 10 μg/L 6-PPDQ reduced the reproductive capacity. Meanwhile, exposure to 1 and 10 μg/L 6-PPDQ enhanced the germline apoptosis, which was accompanied by upregulation of ced-3, ced-4, and egl-1 expressions and downregulation of ced-9 expression. The observed increase in germline apoptosis in 1 and 10 μg/L 6-PPDQ exposed nematodes was associated with the enhancement in DNA damage and increase in expressions of related genes of cep-1, clk-2, hus-1, and mrt-2. The detected enhancement in germline apoptosis in 1 and 10 μg/L 6-PPDQ exposed nematodes was further associated with the increase in expressions of ced-1 and ced-6 governing the cell corpse engulfment process. Molecular docking analysis indicated the binding potentials of 6-PPDQ with three DNA damage checkpoints (CLK-2, HUS-1, and MRT-2) and corpse-recognizing phagocytic receptor CED-1. Therefore, our data suggested the toxicity on reproductive capacity by 6-PPDQ at environmentally relevant concentrations by enhancing DNA damage- and cell corpse engulfment-induced germline apoptosis in organisms.

SCAV-3 affects apoptotic cell degradation in Caenorhabditis elegans.

As the final step in apoptosis, apoptotic cells (ACs) are swiftly removed by specialized phagocytes, such as macrophages, or nonprofessional phagocytes, such as epidermal cells. Genetic studies of model organisms such as Caenorhabditis elegans have helped to elucidate the mechanisms of AC clearance and the underlying causes of disorders related to the dysregulation of these pathways. C. elegans possesses six class B scavenger receptor homologs, but whether they affect apoptosis is unknown. Here, we show that only the loss of function of scav-3, the C. elegans homolog of human lysosomal integral membrane protein-2, resulted in a considerable accumulation of cell corpses, which was caused by a failure in degradation rather than engulfment. SCAV-3 was found to be widely distributed and localized in lysosomes to maintain the integrity of the lysosomal membrane. Further study revealed that loss of scav-3 had no effect on phagosome maturation or the recruitment of lysosomes to phagosomes carrying cell corpses. Moreover, we discovered that the hydrolytic enzymes contained in the lysosomes were reduced in phagosomes in scav-3 mutants. Thus, hydrolases may leak from the damaged lysosome during phagolysosome formation due to the loss of scav-3 function, which reduces lysosome digestion activity and thus directly contributes to the elimination of ACs.

Phagocytosis: Phagolysosome vesiculation promotes cell corpse degradation

Cutting up food into small pieces is well known to improve digestion. New work now shows that this concept also applies in the cellular world, by demonstrating that phagolysosome vesiculation promotes cell corpse degradation in Caenorhabditis elegans blastomeres.

A BORC-dependent molecular pathway for vesiculation of cell corpse phagolysosomes

Phagocytic clearance is important to provide cells with metabolites and regulate immune responses, but little is known about how phagolysosomes finally resolve their phagocytic cargo of cell corpses, cell debris, and pathogens. While studying the phagocytic clearance of non-apoptotic polar bodies in C.elegans, we previously discovered that phagolysosomes tubulate into small vesicles to facilitate corpse clearance within 1.5 h. Here, we show that phagolysosome vesiculation depends on amino acid export by the solute transporter SLC-36.1 and the activation of TORC1. We demonstrate that downstream of TORC1, BLOC-1-related complex (BORC) is de-repressed by Ragulator through the BORC subunit BLOS-7. In addition, the BORC subunit SAM-4 is needed continuously to recruit the small GTPase ARL-8 to the phagolysosome for tubulation. We find that disrupting the regulated GTP-GDP cycle of ARL-8 reduces tubulation by kinesin-1, delays corpse clearance, and mislocalizes ARL-8 away from lysosomes. We also demonstrate that mammalian phagocytes use BORC to promote phagolysosomal degradation, confirming the conserved importance of TOR and BORC. Finally, we show that HOPS is required after tubulation for the rapid degradation of cargo in small phagolysosomal vesicles, suggesting that additional rounds of lysosome fusion occur. Thus, by observing single phagolysosomes over time, we identified the molecular pathway regulating phagolysosome vesiculation that promotes efficient resolution of phagocytosed cargos.

Reproductive toxicity and underlying mechanisms of fine particulate matter (PM2.5) on Caenorhabditis elegans in different seasons

Although numerous studies have investigated that atmospheric fine particulate matter (PM2.5) can be toxic to environmental organisms, the research on the reproductive toxicity of PM2.5 is limited, and the key toxic components and underlying mechanisms remain unknown. In this work, PM2.5 samples of four seasons in Nanjing from March 1, 2021, to February 28, 2022 were collected and the chemical components were analyzed. Caenorhabditis elegans (C. elegans) was employed to conduct the toxicological testing. The reproductive toxicity of PM2.5 to C. elegans in different seasons was evaluated by multiple reproductive endpoints. Exposure to high concentrations of PM2.5 significantly decreased the brood size and the number of fertilized eggs in utero. PM2.5 exposure also increased the number of germ cell corpses and caused abnormal expression of apoptosis-related genes (ced-9, ced-4, and ced-3), which confirmed that PM2.5 induced germline apoptosis. In addition, PM2.5 exposure significantly increased the production of reactive oxygen species (ROS) in C. elegans and the fluorescence intensity of HUS-1 protein in of transgenic strain WS1433. Meanwhile, the expression of genes related to DNA damage (cep-1, clk-2, egl-1, and hus-1) and oxidative stress (mev-1, isp-1, and gas-1) also significantly altered in C. elegans, suggesting induction of DNA damage and oxidative stress. According to Pearson correlation analyses, DNA damage and oxidative stress were significantly correlated with multiple reproductive endpoints in C. elegans. Thus, it was speculated that PM2.5 caused reproductive dysfunction and germ cell apoptosis in C. elegans may be by inducing ROS and DNA damage. In addition, heavy metals in PM2.5 were significantly correlated with multiple endpoints at physiological and biochemical, suggesting that the heavy metals might be an important contributor to the reproductive toxicity induced by PM2.5.

Nanoplastic exposure in soil compromises the energy budget of the soil nematode C. elegans and decreases reproductive fitness

Environmental nanoplastics (NPs) can accumulate in soils, posing a potential risk to soil ecosystems. However, the ecotoxicity of NPs for soil organisms has received little research attention. This study investigated whether NP exposure in soil leads to reproductive decline in the soil nematode Caenorhabditis elegans and sought to determine the mechanisms by which it may occur. Wild-type N2 C. elegans L1 larvae were exposed to various concentrations of nano-sized polystyrene (100 nm) in soil (0, 1, 10, 100, and 1000 mg/kg dry weight) for 96 h. We show that nano-sized polystyrene (100 nm) labeled with red fluorescence significantly accumulated in the intestine of C. elegans in a dose-dependent fashion via soil exposure (8%–47% increase). In addition, NP soil exposure led to 7%–33% decline in the number of eggs in utero and 2.6%–4.4% decline in the egg hatching percentage. We also find that the number of germ cell corpses (31%–55% increase) and the mRNA levels of germline apoptosis marker gene ced-3 (14%–31% increase) were significantly higher with greater NP soil exposure (10, 100, and 1000 mg/kg), while intracellular ATP levels were significantly reduced. Finally, the DEBtox model, which is based on the dynamic energy budget theory, was applied to show that the increased reproductive costs for C. elegans caused by NPs in soil are associated with energy depletion and reproductive decline. The threshold value (4.18 × 10−6 mg/kg) for the energy budget also highlighted the potential high reproductive risk posed by NPs in terrestrial ecosystems. Our study provides new insights into how soil organisms interact with NPs in soil ecosystems.

Tissue-specific modifier alleles determine Mertk loss-of-function traits.

Knockout (KO) mouse models play critical roles in elucidating biological processes behind disease-associated or disease-resistant traits. As a presumed consequence of gene KO, mice display certain phenotypes. Based on insight into the molecular role of said gene in a biological process, it is inferred that the particular biological process causally underlies the trait. This approach has been crucial towards understanding the basis of pathological and/or advantageous traits associated with Mertk KO mice. Mertk KO mice suffer from severe, early-onset retinal degeneration. MERTK, expressed in retinal pigment epithelia, is a receptor tyrosine kinase with a critical role in phagocytosis of apoptotic cells or cellular debris. Therefore, early-onset, severe retinal degeneration was described to be a direct consequence of failed MERTK-mediated phagocytosis of photoreceptor outer segments by retinal pigment epithelia. Here, we report that the loss of Mertk alone is not sufficient for retinal degeneration. The widely used Mertk KO mouse carries multiple coincidental changes in its genome that affect the expression of a number of genes, including the Mertk paralog Tyro3. Retinal degeneration manifests only when the function of Tyro3 is concomitantly lost. Furthermore, Mertk KO mice display improved anti-tumor immunity. MERTK is expressed in macrophages. Therefore, enhanced anti-tumor immunity was inferred to result from the failure of macrophages to dispose of cancer cell corpses, resulting in a pro-inflammatory tumor microenvironment. The resistance against two syngeneic mouse tumor models observed in Mertk KO mice is not, however, phenocopied by the loss of Mertk alone. Neither Tyro3 nor macrophage phagocytosis by alternate genetic redundancy accounts for the absence of anti-tumor immunity. Collectively, our results indicate that context-dependent epistasis of independent modifier alleles determines Mertk KO traits.

Trim-21 promotes proteasomal degradation of CED-1 for apoptotic cell clearance in C. elegans.

The phagocytic receptor CED-1 mediates apoptotic cell recognition by phagocytic cells, enabling cell corpse clearance in Caenorhabditis elegans. Whether appropriate levels of CED-1 are maintained for executing the engulfment function remains unknown. Here, we identified the C. elegans E3 ubiquitin ligase tripartite motif containing-21 (TRIM-21) as a component of the CED-1 pathway for apoptotic cell clearance. When the NPXY motif of CED-1 was bound to the adaptor protein CED-6 or the YXXL motif of CED-1 was phosphorylated by tyrosine kinase SRC-1 and subsequently bound to the adaptor protein NCK-1 containing the SH2 domain, TRIM-21 functioned in conjunction with UBC-21 to catalyze K48-linked poly-ubiquitination on CED-1, targeting it for proteasomal degradation. In the absence of TRIM-21, CED-1 accumulated post-translationally and drove cell corpse degradation defects, as evidenced by direct binding to VHA-10. These findings reveal a unique mechanism for the maintenance of appropriate levels of CED-1 to regulate apoptotic cell clearance.