Hypoxia is a common stressor for aerobic cells that can lead to cell acidification, oxidative stress, cell cycle arrest and death (17). Using transmission electron microscopy, recent studies have revealed the presence of abundant electron-dense deposits, which represent aggregates of unfolded and misfolded proteins in neurons exposed to ischemic-hypoxic brain injury (18,19). During hypoxic stress, the obstruction of protein folding serves as the primary cause of protein aggregation, prompting eukaryotes to develop unfolded protein responses as a regulatory mechanism (20,21). In the current study, a comprehensive review of the mechanisms involved in hypoxia-induced aggregation of unfolded and misfolded proteins, and the cellular strategies relying to this phenomenon is presented.

The number of large multidomain proteins notably increases from prokaryotes to eukaryotes. These proteins exhibit diverse conformations, and as their protein configurations become more complex, the possibility of misfolding increases (22). Hydrophobic amino acid residues, unstructured regions in folding intermediates and misfolded proteins are often exposed to solvents, leading to aggregation (23). Aggregates are primarily driven by liquid-liquid phase separation (LLPS) or hydrophobic forces, depending on the concentration (24). While most aggregates are amorphous, the aggregation of certain proteins leads to the formation of amyloid fibers characterized by β strands normal to the long fibril axis (cross-β structure) (25). Before fiber formation, amyloid often exists in an oligomeric state, and both types of aggregates play crucial roles in diseases (26). For instance, cerebral blood flow decreased in patients with early Alzheimer's disease (AD) (27). Increased binding of oligomeric β-amyloid protein (Aβ) to ROS leads to vasoconstriction around brain cells, contributing to decreased cerebral blood flow, which may initiate a cascade reaction involving amyloid Aβ itself or the fibrous Aβ, which is important for driving cognitive decline (27,28). Thus, it is necessary to understand the mechanisms underlying hypoxia-induced protein aggregation for elucidating the pathogenesis of neurodegenerative disease and developing intervention strategies.

Molecular chaperones play an important role in maintaining protein homeostasis, and assist other proteins in acquiring functionally active conformations without affecting their final structure. Different types of molecular chaperones receive newly synthesized protein chains from ribosomes to ensure effective folding and minimize aggregate formation by guiding them through appropriate folding pathways (26). As proteins are structurally dynamic, proteostasis occurs via a network of chaperones and protein degradation mechanisms that continuously monitor the proteome (29,30). Chaperones help prevent chain compaction and misfolding, and facilitate the removal of protein aggregates through lysosomal-autophagy degradation (31). Before degradation, the depolymerization of aggregates is cooperatively carried out by heat shock proteins (Hsps) such as Hsp70, Hsp110 and Hsp40 (32,33). The clearance pathways involving proteasomes and lysosomes are intricately linked to the Hsp70 and Hsp90 chaperone systems through specialized ubiquitin ligases such as the co-chaperone C-terminus of the Hsc70-interacting protein and the BAG domain (34,35).

However, under hypoxia conditions, the regulatory network of protein homeostasis is disrupted, and numerous molecular chaperones are affected by hypoxic stress. Nguyen et al (36) observed notable global reductions of ATP-dependent Hsp70 and Hsp90 (83 and 78%, respectively) after 24 h of hypoxia treatment. Conversely, the protein expression of the ATP-independent Hsp27 and Hsp40 in the brain, heart and muscle remained constant throughout the 24-h hypoxia treatment. However, with prolonged hypoxia, the expression of the Hsp27 and Hsp40 genes in these tissues was also reduced, suggesting that the protein expression of these chaperones may also eventually decrease under hypoxia. These results suggest that energy conservation is prioritized over cytoprotective protein chaperoning in naked mole-rat tissues during acute hypoxia. Although ATP-independent partners do not require ATP to regulate their functional cycle passive histone aggregation (37), aggregate bursts under low oxygen stress also suggest that these ATP-independent partners cannot remedy the homeostatic imbalance caused by the energy gap. In fact, the effects of hypoxic stress on protein chaperones are not machine-made, for example, C2C12 cells induce Hsp70 gene expression through a similar mechanism to heat stress during acute hypoxia (38). However, macrophages exposed to 5% oxygen for 24 h notably reduced Hsp70 expression and recovered after reoxygenation (39). Proteomics indicated that Hsp72 downregulation in the cerebral cortex of rats after 5 days of hypoxia reached its lowest level (40). In addition, the Hsp90 chaperone family TRAP1 has been found to be frequently induced in tumors and regulate energy metabolism after HIF-1 stabilization (41), and hypoxia can also reduce the transcription of cyclin B1 in liver cancer cells through Hsp90 (42). These contradictory results may be due to differences in the function and distribution of molecular chaperons, and the crosstalk between hypoxia stress and chaperons may need further exploration.

Disulfide bonds are commonly found in protein domains located in the cytoplasmic membrane and enhance protein stability. The cleavage of disulfide bonds triggers the function of some secreted soluble proteins and cell-surface receptors (43). Oxidative protein folding refers to the restorative process through which proteins containing disulfide bonds transit from fully reduced and unfolded states to their original bioactive forms (44-46). Koritzinsky et al (47) used 35S labeling and suggested that the production of disulfide bonds was limited by hypoxic surroundings and that protein folding recovered upon oxygen restoration (48,49). This evidence suggested that oxygen depletion may seriously impede disulfide bonding leading to protein misfolding.

In brief, oxygen deprivation disrupts protein folding through multiple mechanisms, including inhibiting disulfide bond formation, inactivation of molecular chaperones and elevation of ROS levels (50,51). Prolonged accumulation of misfolded proteins may eventually result in the formation of pathological protein aggregates (Fig. 1). This shift contributes to the development of neurodegenerative diseases, such as AD, Huntington's disease (HD) and Parkinson's disease (PD) (3,52,53). Moreover, hypoxic-ischemic encephalopathy (HIE) occurs when the brain is exposed to oxygen deprivation and ischemia. Newborns often experience HIE due to birth asphyxia, causing an unfavorable prognosis owing to cerebral dysfunction, neuronal cell death and neurological deficits. Notably, marked molecular and subcellular changes observed in the brain cells of patients with HIE include protein misfolding, aggregation and organelle damage (54). The disruption of protein homeostasis is also closely related to cardiac hypertrophy, cardiomyopathy and heart failure caused by cardiovascular hypoxia (55). Soluble protein oligomers have been observed in the myocardial cells of patients with idiopathic dilated cardiomyopathy, non-ischemic cardiomyopathy, or hypertrophic heart disease (56). Similarly, aggregation of abnormal and ubiquitinated proteins has been detected in the heart of individuals with dilated cardiomyopathy or ischemic heart disease (57). Pattison et al (58) previously demonstrated that the expression of ectopic gene that containing 83 glutamine repeats in cardiomyocytes promoted the cohesive accumulation and aggregation of pre-glutamine amyloid oligomers, increasing protein deposition, cardiac muscle cell death and heart failure.

Cellular proteostasis is tightly controlled by a network of molecular chaperones. In addition to counteracting abnormal folding and aggregation by directly binding to misfolded proteins (59), chaperones also assist the ubiquitin-proteasome system (UPS) (60) and the autophagy-lysosome system in degrading aggregators for proteostasis (61).

The lysosomal-mediated autophagy degradation pathway is a major hunter for clearing protein aggregates, especially in neurodegenerative diseases (62). Most neurodegenerative diseases involve pathological abnormal protein aggregates, developing neurofibrillary tangles. For example, Aβ and C-terminal fragments of the amyloid precursor protein in AD, mutant α-synuclein in PD, polyglutamine-expanded huntingtin in HD, and mutant superoxide dismutase 1 and TAR DNA-binding protein 43 (TDP-43) in ALS (63-65). These protein aggregates mainly target the autophagy lysosomal degradation pathway, and chaperone proteins play a key role in this process. Specific aggrephagy receptors have been reported in yeast S. cerevisiae (Atg19) and C. elegans (SEPA-1 and EPG-7) (66-68). Recently, Ma et al (69) reported the function of the TRiC subunit chaperonin-containing TCP-1 subunit 2 (CCT2) in aggrephagy in mammals and yeast. CCT2 promotes autophagosome incorporation and clearance of protein aggregates with little liquidity by interacting with ATG8s and aggregation-prone proteins independent of cargo ubiquitination. The dual function of CCT2, as a chaperone and an aggrephagy receptor, enables double-layer maintenance of proteostasis.

Cellular stress and aging can lead to a decrease in protein homeostasis. In addition to the inhibition of protein chaperone activity by hypoxia metabolism, notably, hypoxia-reoxygenation treatment dysregulates key molecules that maintain autophagy-lysosomal flux in primary human trophoblasts, notably reduced autophagosomes and autolysosomes (70). The expression of ubiquitin 26S-proteasome E3 ligase, autophagolysosomal degradation related mRNA transcripts and proteins, and integrated stress response markers were also decreased after 12 days of hypoxic feeding (71).

The UPS system is strongly associated with regulating biomolecular condensation (60). More specifically, ubiquitin and other post-translational modifications act as agents of phase separation, and are essential for the formation of condensates and ubiquitin-proteasome system activity (5). It is noteworthy that previous studies demonstrated that polyubiquitin chains can function as multivalent molecules that can drive either the assembly or the disassembly of condensates via interactions with various ubiquitin-binding proteins (72,73).

Cellular responses to hypoxia primarily aim to enhance cell survival and restore oxygen equilibrium. In the context of uncontrolled protein folding, the accumulation of unfolded or misfolded proteins within the ER or mitochondrial space leads to activation of UPR (50,53). Through its distinct signalling network, the UPR pathway restores protein homeostasis, alleviates the burden of protein aggregation and maintains cell viability (74-78). The heavy-chain-binding protein (BIP), a member of the Hsp70 family, is a crucial chaperone that triggers UPR activation. BIP enters the ER by binding to hydrophobic amino acids to prevent incorrect folding and polymerization of the polypeptide chains. This is followed by ATP binding and subsequent release of the bound polypeptides through ATP hydrolysis (79). Environmental stress leads to misfolded proteins accumulating, causing the release of BIPs (80). The released BIPs undergo phosphorylation and polymerization, triggering the activation of protein kinase R (PKR)-like ER kinases (PERKs) and inositol-requiring enzyme-1 (IRE1) (81). Additionally, activating transcription factor (ATF) 6 is switched to the Golgi apparatus and convered to soluble and active cytoplasmic ATF6 (82-84). These PERK, IRE1 and ATF6 sensors constitute three distinct signalling pathways within the UPR (80,85). Hypoxia induces BIP expression in both cancer and endothelial cells (86-88). Hypoxia can activate the PERK signalling pathway in various models (89-91), and the phosphorylation of eukaryotic initiation factor 2 (eIF2) mediated by PERK was observed within minutes of hypoxic exposure, with a reduced response rate as the oxygen concentration increased (92). To alleviate ER stress, UPR signalling inhibits protein aggregation by reducing protein synthesis flux and activating the transcriptional program of molecular chaperones.

The hypoxia-mediated UPR has been well demonstrated in the tumor microenvironment, and exposure of solid tumors to intermittent hypoxia may lead to high ROS levels and UPR activation (93-95). For example, increased ATF4 expression has been shown in numerous hypoxic and nutrient-deprived tumors (96) and can mediate autophagy under hypoxia (97). Immunohistochemical staining demonstrated increased expression of ATF4 in hypoxic, perinecrotic regions distal to the tumour vasculature, consistent with a nutrient-deprived mechanism of translational activation. In addition, the distribution of p-eIF2a and p-GCN2 signal demonstrated considerable association in serial sections, consistently, spontaneous mouse tumours also contain greater levels of p-eIF2a and ATF4 than corresponding normal tissue (98). PERK and ATF4 protect glioblastoma cells exposed to cyclic hypoxia or radiotherapy from oxidative damage (99,100). In human cervical cancer, PERK activation leads to the accumulation of oncogenic lysosomal-associated membrane protein 3, thus increasing the aggressiveness of these cells (99).

Mitochondria are the primary consumers of oxygen within cells. Early mitochondrial dysfunction is implicated in numerous hypoxic diseases such as cancer and neurodegenerative diseases (17,101,102). The efficiency of mitochondrial oxidative phosphorylation is markedly reduced under hypoxic conditions due to mitochondrial perinuclear localization and fragmentation mediated by CHCHD4 (103-105). Mitochondria contain their inherent genetic information and rely on stress response systems to translate and fold encoded proteins, and refold nuclear-encoded proteins (106). Maintenance of protein homeostasis in this organelle involves unique molecules such as Hsp60 and the peptidase lon peptidase 1 (106,107). Under hypoxic conditions, mitochondria can also experience unfolded or misfolded proteins aggregating. For example, using C. elegans, Kaufman et al (9) identified 65 preferentially insoluble mitochondrial proteins and 110 generally insoluble mitochondrial proteins during hypoxia, and reported that the abundance of hypoxia-induced mitochondrial protein aggregates (HIMPA) increased notably with the severity of hypoxia. Additionally, Yan et al (108) reported that disruption of mitochondrial proteostasis and mitochondrial protein aggregation are early processes involved in hypoxia in C. elegans. Like in the ER, mitochondria also activate their own UPR, which is known as the mitochondrial UPR (UPRmt). The UPRmt is classically considered as a transcriptional response that increases the expression of mitochondrial chaperones to protein misfolding and aggregation in mitochondria (109-111). In C. elegans, the UPRmt was found to be regulated by sensitizing transcription factor associated with stress 1 (ATFS-1), which is a transcription factor within mitochondrial and nuclear localization sequences, and dual subcellular localization. ATFS-1 is transported into the mitochondrial matrix and then degraded by LON proteases under steady-state conditions. The transport of ATFS-1 is downregulated in mitochondrial dysfunction, and ATFS-1 is subsequently transported to the nucleus to stimulate transcriptional responses (111,112). Additional regulatory mechanisms may exist in mammalian cells, with ATF5 acting as a functional ortholog of ATFS-1 (113). In addition, ATF4 and the C/EBP homologous protein activating are important in the activation of UPRmt (114,115). Activation of UPRmt to mitochondrial stress in cancer could maintain mitochondrial integrity and tumor growth (116). A recent study by Sutandy et al (117) showed that UPRmt signaling is prompted by the release of two individual signals in the cytosolmitochondrial ROS (mtROS) and mitochondrial protein precursors in the cytosol, leading to the release of HSF1 by Hsp70, which results in nuclear translocation and transcription of UPRmt genes (117).

The expression of these transcription factors is mediated by eIF2α kinase phosphorylation (118). Recently, Guo et al (119) delineated the relationship between mitochondrial stress and the relay of ATF4. Heme-regulated initiation factor 2 α kinase (HRI) is a necessary eIF2 kinase for this relay. A genome-wide CRISPRi screen identified two upstream signaling factors for HRI: The OMA1 zinc metallopeptidase (OMA1), as a mitochondrial stress-activated protease, and the DAP3 binding cell death enhancer 1 (DELE1) associating with the inner mitochondrial membrane. Mitochondrial stress results in DELE1 cleavage by OMA1 and its accumulation in the cytosol, which interacts with HRI and increases eIF2 kinase activity. These results indicated that the UPRmt and UPR signaling pathways can been interlinked via eIF2α (Fig. 2) (109-111).

HIMPA consistently alleviates hypoxia-induced cell death, and UPRmt activation had the same effect. However, UPRmt is not necessarily protective against hypoxia-induced cell death (108). It is the overactivation of UPRmt that can induce cell death as in the case of UPR (118), and the relationship of HIMPA with UPRmt and its crosstalk with UPR needs to be explored further.

SGs are assemblies of non-translating messenger ribonucleoprotein granules, various non-membrane-bound cellular compartments that contain high concentrations of proteins and RNA (138), and are close to UPR (139). The formation of SGs is facilitated by interactions between mRNAs and mRNA-binding proteins, translation initiation factors, the 40S ribosomal subunit (a myriad of RNA-binding proteins) and translationally stalled mRNAs (139,140). Once the cells return to a normal and non-stressful environment, SGs disperse and protein translation is reinstated (141). Eukaryotic cells use SGs to redirect limited resources from protein synthesis to survival and stress resistance.

The core of the SG central node of this network incorporates the G3BP SG assembly factor 1 (G3BP1), which serves as a molecular switch instigating RNA-dependent LLPS in response to elevated concentrations of free RNA in cells. G3BP1 is also capable of modulating LLPS propensity via three different inherently disordered regions. The core SG network can be simultaneously reinforced or weakened by altering G3BP1-binding factors (142). The assembly activation cues of SGs coalesce with UPR signals to create networks that maintain protein homeostasis (139,143). The conventional assembly process of SGs is mediated by eIF2 phosphorylation. The eIF2 kinase family includes PERK, PKR, general control non-depressible 2 (GCN2), and HRI (144,145). In a hypoxic environment, eIF2 phosphorylation is induced by PERK and activated through UPR signaling or the OMA1-DELE1-HRI pathway, which is initiated by UPRmt (119). The phosphorylation state of eIF2 is regulated by its interaction with eIF2β, and this interaction inhibits the conversion of GDP to GTP by eIF2β, resulting in a decrease in the concentration of ternary complex eIF2-GTP-tRNA Met (146,147). Consequently, the RNA-binding protein TIA1 and T-cell-restricted intracellular antigen-associated protein (TIAR) stimulate the formation of the noncanonical 48S preinitiation complex (148). This complex, unable to recruit the 60S ribosomal subunit translation, can be used for SG assembly (148-150).

In addition to being mediated by UPR activation, hypoxia causes the assembly of SGs through several other pathways. Hypoxia is frequently associated with nutrient scarcity, and mammalian cells can sense alterations in amino acid levels through the GCN2 and mTORC1 pathways (151). Amino acid deprivation inhibits mTORC1-mediated protein translation but stimulates angiogenesis via the GCN2-ATF4 amino acid starvation response pathway, that is independent of HIF-1 (152,153). GCN2 also promotes eIF2 stimulation and collaborates with PERK to shield hypoxic cells from apoptosis (154). Furthermore, hypoxia generally triggers type I interferon (IFN) pathway inhibition and reduces IFN secretion, which could lead to uncontrolled double-stranded RNA (dsRNA) expression (155). As a stressor, dsRNA can incite the phosphorylation of eIF2α via PKR. This phosphorylation results in the formation of SGs, which serve as an antiviral core (156). To summarize, hypoxia instigates the activation of SG assemblies (Fig. 3).

Hypoxia-induced SG assembly effectively improves cell viability, which has been well demonstrated in the hypoxic microenvironment of cancer. Apoptosis-related molecules that accumulate within SGs assembled by cancer cells manifested antiapoptotic effects (157), and the development of hypoxia-induced SGs causes drug resistance in cancer (158). By pharmaceutically impeding hypoxia-induced SG formation in HeLa cells, Timalsina et al (159) managed to decrease drug resistance in hypoxic microenvironments. A study by Attwood et al (160) showed that hypoxia increased the number of late apoptotic/necrotic glioblastoma cells during the raloxifene-induced delay in SG dissolution. Liu et al (161) provided that hypoxic conditions could result in FUS-circTBC1D14-associated SG formation in the cytoplasm after PRMT1 modification, thus contributing to the maintenance of cellular homeostasis and promoting tumor progression in triple-negative breast cancer.

In rodent models, SGs were found to protect hepatocytes against hypoxia-induced damage by reducing apoptosis. With the increased expression of the SG marker proteins G3BP1 and TIA-1, the degree of liver injury, HIF-1α and apoptosis induced by acute liver failure decreases (162). In addition, Hu et al (163) found that impaired SGs are important in the pathogenesis of spinal muscular atrophy.

It is noteworthy that nematodes and rat cardiomyocytes produced characteristic SGs in mitochondria stimulated by sublethal hypoxia. Mitochondrial SGs are involved in early mitochondrial pathology and are closely associated with UPRmt (14).

P-bodies are also a type of biomolecules participating in phase separation (164). The structure of P-body is similar to that of SGs, and P-body shuttle RNA binding proteins and mRNAs between the two condensates (165,166). Usually, SGs uniquely house certain translation initiation factors, while P-body specifically abound to factors associated with mRNA degradation and decay, leading to functional differences (164,167). In the presence of hypoxic stress, SGs can maintain cell survival (168), while P-bodies seemed to be more inclined to regulate hypoxia-related signaling molecules.

Past research has demonstrated that hypoxia can induce RNP granule formation in C. elegans oocytes, and RNP foci are similar to the RNA-related functions of P-bodies (169). Saito et al (16) reported that HIF-1α was upregulated by the microRNA (miR)-130 family during hypoxia. The miR-130 family was increased under hypoxia, and their target was DDX6 mRNA, a component of the P-bodies. These results reveal a new translational mechanism of HIF-1α and P-bodies in hypoxic stress (16). The USP52 protein and HIF1A mRNA were found to colocalize with cytoplasmic P-bodies, suggesting that P-bodies recruit HIF1A mRNA for assembly through LLPS. The P-body component USP52/PAN2 can enhance the stability of HIF-1α mRNA, which is crucial under hypoxic conditions (170). Moreover, HIF1A mRNA localizes to P-bodies following microtubule disruption for a short period of translational repression (171). These findings suggest that P-bodies contribute to the regulation of HIF1A mRNA stabilization and protein translation, which are critical for hypoxic signaling and cellular hypoxic response (17).

Metabolic flux is an important intracellular change that occurs during hypoxic stress. When cellular oxidative phosphorylation is impaired by hypoxia, glycolysis becomes the primary source of energy (172,173). Although the glycolytic pathway has a shorter energy supply pathway, the total amount of ATP produced is lower than that produced during oxidative phosphorylation (174). To meet the ATP required for survival and speed up the flow of glycolysis, cells integrate the enzymes required for glycolysis and other scaffold proteins through LLPS to form a special biomolecular condensate, called a glycolytic body (G-body) (15,175).

Under hypoxic conditions, glycolytic enzymes are compartmentalized into cytoplasmic structures (176), and analogous condensates form were also found in C. elegans neurons (177). Therefore, Jin et al (15) demonstrated that under hypoxic conditions, cells assemble non-membrane organelles that include glycolytic enzymes, called G-bodies. They also found that glucose consumption increased, and that the level of glycolytic intermediates decreased in cells with G bodies. It is noteworthy that the formation of G-bodies increases the glycolytic output in hypoxia (15) via glycolytic enzymes such as phosphofructokinase, pyruvate kinase, acetyl-CoA carboxylase and yeast pyruvate kinase Cdc19 (178-181). These enzymes can catalyze the rate-limiting step in glycolysis and be utilized to increase the glycolysis rate under hypoxic conditions. While the mechanism of G-body activation has not been elucidated. Gregory et al (182) detected hundreds of RNA-binding proteins in G-bodies using genomic and proteomic methods. The failure of nonspecific endonucleases to maintain the structural integrity of G-bodies suggests that the assembly of G-bodies replying to hypoxia is likely mediated by an RNA-dependent phase separation mechanism (182). The enzymes involved in the formation of G-body aggregates follow a specific order post-nucleation, and the entry of each metabolic enzyme into the G-body is tightly regulated (183). The multiple glycolysis enzymes within phase separation may function to enhance the activity and increase the reaction rate in energy production, thereby forming 'metabolons' during hypoxic stress (184).

Notably, cells that are unable to form G-bodies undergo abnormal division and yield nonviable daughter cells during hypoxia, and the formation of G-bodies represents a conserved adaptive response that maintains the energy requirements of the cells (15).

Fatty acids consist a major fuel in various cells. The depletion of oxygen substrate severely inhibits the fatty acid β oxidative energy pathway of the cell, and the accumulated excess fatty acids are transformed into triglycerides for storage (185,186). The ER participates in synthesizing these triglycerides, which are subsequently stored in biomolecular condensate called LDs (187). LDs are dynamic lipid compartments that can effectively manage fluctuating cellular lipids. Following oxygen restoration and activation of fatty acid oxidation, LDs are broken down by neutral lipase, and the liberated fatty acids serve as substrates for mitochondrial oxidation, leading to energy production (188). LDs contain core lipid components, and are surrounded by an amphipathic lipid layer (189). Almost all organisms synthesize LDs, whose formation is initiated by the synthesis of neutral lipids (NLs) (190). Overnutrition or various stressors prompts cells to produce NLs in the ER bilayer (191,192), where the synthesized NLs mix with phospholipids on the membrane and diffuse in the ER bilayer (193). When the NL concentration exceeds the nucleation threshold, LLPS drives LD formation to prevent NL accumulation in the ER membrane (194).

Hypoxia-induced LDs were initially observed in cancer cells (195). They may require substantial lipids for biosynthesis, and lipid-derived bioactive molecules for cytomembrane formation and a high level of cell proliferation (196,197). It has been demonstrated that lipid formation via HIF-1α mediates reductive glutamine metabolism instead of pyruvate-mediated acetyl-CoA production in cancer cells (198). Thus, a high LD content was closely related to transcription driven by hypoxia in hypoxic cancer cells. LDs are associated with various malignant phenotypes (198). Mounting evidence supports the diverse roles of LDs in cancer cells responses to stress conditions, such as maintaining ER homeostasis (199), clearing ROS (200) and preventing drug resistance (201), all of which are crucial in the maintenance of homeostasis in cancer cells.

LD formation and degradation are controlled by numerous enzymes and LD-associated proteins. Hypoxia-inducible LD-associated protein (HILPDA) is a paramount LD-associated protein induced by HIF-1 and fatty acid expression. It localizes in the LDs of several cell types, and is situated near the ER and LDs within cells. HILPDA directly inhibits the activity of adipose triglyceride lipase via physical interaction and encourages LD accumulation by stimulating triglyceride synthesis (202,203). These findings suggest that under hypoxic stress, not only proteins and RNA, but also lipids can be orchestrated to assemble into specific molecular biopolymers for survival.

Hypoxic stress can induce the formation of protein condensates, which play a role in promoting basic biochemical processes. For instance, prolyl hydroxylases are involved in regulating molecular responses to oxygen availability. These proteins hydroxylate HIF-α, enabling its ubiquitination and degradation (204,205). Increased expression of HIF can lead to the generation of ROS, which modulates HIF-α stabilization in conjunction with prolyl hydroxylase domain proteins (PHD) (206,207). The PDH family has a function in regulating HIF through the condensation of PDH3, a protein expressed in response to oxygen deprivation that contributes to neural cell death. PDH3 forms subcellular condensates in the presence of oxygen, but its condensation is notably decreased under hypoxia (208,209). The formation of PDH3 condensates relies on microtubules and involves the integration of components from the 26S proteasome, chaperones and ubiquitin. The PHD2 condensates exhibit liquid characteristics similar to other condensates (210). When PHD3 is actively expressed under normoxia, it leads to the condensation of proteasome components, triggering apoptosis in HeLa cells. Apoptosis occurs in cells prone to PHD3 condensation and is observed before apoptosis (210).

Recently, Theodoridis et al (211) discovered that hypoxia-induced cell acidification could induce the aggregation of certain amyloid proteins in the nucleus. These proteins are not unfolded proteins but rather formed through phase separation of a class of long-chain non-coding RNAs derived from a specific site of stimulation within the ribosomal gene spacer (211). Local nuclear translation under stress conditions is crucial under various physiopathological conditions. Amyloid bodies enhance local nuclear translation during stress, suggesting that aggregates, similar to liquid condensates, can facilitate complex biochemical reactions (211,212). (However, a further detailed assessment is required to determine the degree to which soil-like condensate formation occurs under stress.

In conclusion, cells initiate the assembly of biomolecular condensates to sustain cell survival and regulate metabolism in response to hypoxic conditions. A concise overview of the crucial components and biological functions of hypoxic-related biomolecular condensates is presented in Table I, which can provide valuable information for future research on hypoxic-related diseases.