Discovery of lactylation

In 2019, Zhang and colleagues employed mass spectrometry to detect a 72.021 Da mass shift on lysine residues of histones and proposed lactylation as an enzymatic PTM that uses lactyl coenzyme A (lactyl-CoA) as a substrate (Fig 1) [8]. Through isotope metabolic labeling, they demonstrated the widespread occurrence of lactylation in both human and mouse cells and identified 28 lactylation sites on the core histones [8]. They also further proved that lactylation levels correlated with lactate production, with both exogenous and endogenous lactate directly affecting this modification. Lactate that induces lactylation primarily exists in two forms: L-lactate (L-la) and D-lactate (D-la). L-la is predominantly produced through anaerobic glycolysis, particularly under hypoxic conditions, and its accumulation is markedly elevated in tumor cells due to the Warburg effect, reaching concentrations of 10–30 mM [13,14]. Elevated L-la contributes to an immunosuppressive microenvironment and promotes disease progression [15]. In contrast, D-la originates mainly from gut microbiota, diet, and the methylglyoxal pathway, and circulates at much lower levels (approximately 0.01 mM) [16,17]. It rises during conditions like intestinal injury and can impair mitochondrial respiration by disrupting the electron transport chain and increasing ROS [18,19].

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Fig 1. Lactate metabolism and the formation of lactylation.

https://doi.org/10.1371/journal.ppat.1013366.g001

Functionally, L-la serves as a substrate for enzymatic histone L-lactylation (K(L)-la), a modification primarily mediated by the histone acetyltransferase p300 and reversed by histone deacetylases HDAC1–3 [8,20]. K(L)-la plays an important role in linking glycolytic activity to transcriptional regulation. For instance, K(L)-la of cGAS suppresses cGAMP production, thereby modulating antiviral immune responses and tumor progression [21]. On the other hand, D-la promotes a distinct non-enzymatic lactylation pathway-histone D-lactylation (K(D)-la)-primarily through reaction with S-D-lactoylglutathione (LGSH), a byproduct of the glyoxalase pathway [22,23]. K(D)-la has been implicated in gene regulation; for example, it modifies histones to suppress TLR2 expression, thus inhibiting hepatocellular carcinoma (HCC) progression.

Despite their functional differences, detecting K(L)-la and K(D)-la poses substantial challenges due to their identical molecular weights and similar structural properties. Both forms are indistinguishable by high-performance liquid chromatography-mass spectrometry (HPLC-MS) [24], as peptides bearing K(L)-la and K(D)-la are not resolved by reverse-phase HPLC. Moreover, the much higher endogenous concentration of L-la relative to D-la leads to strong background signals, further complicating the detection of D-la-derived modifications [25].

Lactate, derived from glycolysis or the extracellular milieu or microenvironment, contributes to lactyl-CoA synthesis. Lactyl-CoA is a key intermediate in transferring lactyl groups to lysine residues on histones or non-histone proteins. Lactylation is catalyzed by “writers” enzymes such as p300 and KAT7/8, while “erasers” enzymes such as HDACs and SIRTs reverse the modification. Histone lactylation impacts gene expression, while non-histone lactylation influences cellular functions. BioRender supported the illustration rendering portion of this work (https://www.biorender.com/).

The “writers” and “erasers” of lactylation modification

Like other epigenetic modifications, such as methylation and acetylation, lactylation modifications involve “writer” and “eraser” enzymes. The histone acetyltransferase p300, a crucial “writer”, induces histone H3 lysine acetylation and activates gene transcription by transferring the lactyl group from lactyl-CoA to protein lysine residues (Table 1). Zhang and colleagues have validated that p300 is involved in histone lactylation modification, as its overexpression in HEK293T cells increased histone lactylation levels, and its knockdown in both HCT116 and HEK293T cells decreased H3K18la levels [8]. Liu and colleagues further discovered that p300 mediates lactylation and regulates profibrotic gene expression by affecting lactylation levels [26]. In addition, CREB-binding protein (CBP), a homologous protein of p300, performs a similar function. Research by Li and colleagues showed that inhibiting the p300/CBP or blocking its enzyme activity with C646 attenuated lactate-induced lactylation levels in the high mobility group box 1 (HMGB1) immunocomplex [27]. YiaC, another “writer”, catalyzes lactyl-CoA formation and donates the lactyl group for lactylation, impacting metabolic enzyme activity [28]. As a critical lactyltransferase, KAT2A (also known as GCN5), in conjunction with ACSS2, forms a lactyltransferase complex that catalyzes histone lactylation at H3K14 and H3K18, which plays a critical role in tumor progression [29]. Additionally, KAT2A also catalyzes lysine lactylation at K124 of RCC2, and targeting this modification has been shown to suppress the rapid proliferation of breast cancer cells [30]. Beyond histone targets, emerging evidence suggests that KAT2A may also participate in the lactylation of cytoplasmic and mitochondrial proteins, although the mechanisms and biological consequences of these modifications remain to be fully elucidated. Recent research identified KAT8, a lysine acetyltransferase that acts as a pan-lactylation “writer”, enhances protein translation efficiency by lactylating of eEF1A2 at the K408 site, thereby promoting tumor progression [31]. Similarly, KAT7 is a “writing” enzyme involved in lactylation modification [32]. Moreover, AARS1 and AARS2 may act as “writers” and primarily function in the cytoplasm and mitochondria, respectively [33,34].

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Table 1. The “writers” and “erasers” enzymes of lactylation modification.

https://doi.org/10.1371/journal.ppat.1013366.t001

In contrast to “writers”, “erasers” terminate the lactylation cycle and maintain its dynamic balance by removing lactyl modifications from lysine residues. Histone deacetylases (HDACs), comprising HDAC1–11 and categorized into classes I, II, and IV, require Zn²⁺ as a cofactor for their activity. In vitro experiments have demonstrated that HDAC1 and HDAC3 reduce L-la and D-la levels by cleaving ε-N-L-lactyl lysine [35]. Zessin and colleagues further reported that HDAC6 and HDAC8 possess potential delactylase activity, although their enzymatic activity is considerably lower than that of HDAC3 [36]. Conversely, sirtuins (SIRT1–7), classified as class III HDACs, depend on NAD⁺ as a substrate [37]. Among the sirtuins, SIRT1–3 exhibited stronger activity in removing L-la, but HDAC3’s enzymatic activity in this process is several thousand times higher than SIRT2 (Table 1) [20]. Furthermore, Zhang and colleagues discovered CobB serves as the primary lysine delactylase regulating metabolism in Escherichia coli. It specifically modulates the activity of pyruvate kinase I (PykF) by regulating K382la, thereby promoting glycolysis and bacterial growth [28].

Crosstalk between lactylation and other epigenomic events

It has been reported that many proteins contain at least one type of regulatory PTM [42]. Given that most proteins interact with other proteins to varying extents, the crosstalk between PTMs across different proteins is widespread. While research on the crosstalk between lactylation and other PTMs is still in its early stages, emerging studies indicate potential crosstalk between lactylation and acetylation [43,44]. Both commonly target lysine residues, and when occurring on histones, they often share sites such as H3K18, H3K27, and H3K23 [8]. Li and colleagues discovered that the Gli-like transcription factor 1 (Glis1) increases lactate and acetyl-CoA generation by regulating glycolytic gene expression, thereby enhancing the occurrence of lactylation and acetylation [44]. In CD4⁺ Th1 cells, glycolysis maintains acetyl-CoA levels during T-cell differentiation. Lactate dehydrogenase A (LDHA) is important in acetyl-CoA production, increasing histone acetylation at lysine 9 and 27 of H3 (H3K9Ac and H3K27Ac) and boosting IFN-γ expression in Th1 cells [45]. Moreover, p300 is the “writer” for lactylation and acetylation, providing further insights into their interrelationship. Research shows that p300 is highly enriched at the promoters of pluripotency genes (Oct4, Sall4, and c-Myc), suggesting that lactylation and acetylation may work synergistically to regulate cell fate through dynamic changes in lactate and acetyl-CoA levels (Fig 1) [46].

Besides acetylation, various other acylated modifications, including crotonylation, 2-hydroxyisobutyrylation, succinylation, and malonylation, may also co-modify target proteins alongside lysine lactylation, suggesting potential crosstalk among these modifications [47,48]. For example, Gao and colleagues identified a total of 143 sites on 83 lactylated proteins in B. cinerea that were commonly co-modified by lactylation, crotonylation and 2-hydroxyisobutyrylation, furtherly demonstrating the high conservation of acylated modifications among plant, human, and fungi [47]. Furthermore, in lipopolysaccharide (LPS)-activated macrophages, the succinylation of pyruvate kinase 2 at lysine K311 impairs glycolytic activity and drives its nuclear localization, promoting hypoxia-inducible factor (HIF)-dependent gene transcription and IL-1β production [49,50].

Additionally, significant crosstalk exists between lactylation and RNA modifications. Yu and colleagues have reported this potential connection, which between lactylation and RNA modifications, offering new perceptions of epigenetic regulation in carcinogenesis [51]. Specifically, lactate has been shown to induce methyltransferase-like 3 (METTL3) transcription in tumor-infiltrating myeloid cells (TIMs) via histone H3K18 lactylation, a marker associated with poor prognosis in colon cancer patients [52]. In addition to transcriptional regulation, Xiong and colleagues identified two lactylation sites within the zinc-finger domain (ZFD) of METTL3 that are critical for its RNA-binding ability, underscoring the role of lactylation in promoting METTL3-mediated m6A modifications, particularly in TIMs [53]. Notably, beyond its involvement in cancer, METTL3 also acts as a suppressor of antiviral immunity. Activators of the METTL3 RNA methyltransferase complex have been shown to enhance the production of viral particles in cells harboring the HIV-1 provirus [54]. During Vesicular Stomatitis Virus (VSV) infection, METTL3 translocates to the cytoplasm, promotes m6A modification of viral RNA, reduces viral dsRNA accumulation, and consequently attenuates RIG-I/MDA5-mediated antiviral signaling [55]. Given that lactylation upregulates METTL3 expression and enhances its RNA-binding capacity [52,53], it is plausible that lactylation may potentiate METTL3-driven immune suppression during viral infection. Although direct mechanistic evidence in viral contexts remains limited, this potential crosstalk may represent a novel strategy for viral immune evasion.

Beyond regulating METTL3 activity, lactylation also upregulates the expression of YTH N6-methyladenosine RNA binding protein 2 (YTHDF2), an m6A “reader” that selectively binds to m6A-modified RNAs, to the broader regulation of m6A-dependent processes. Mechanistically, lactylation increases YTHDF2 expression, enabling it to recognize m6A-modified Per1 and p53 mRNAs, promoting their degradation, thus accelerating the occurrence of ocular melanoma [51,56]. Although no literature has directly reported the involvement of lactylation in regulating YTHDF2 expression during viral infection, research has demonstrated that YTHDF2 binding to m6A sites on HIV-1 transcripts significantly enhances the stability of viral RNAs [57]. Furthermore, the downregulation of YTHDF2 appears to be linked to and potentially senses host shutoff induced by infection, thereby augmenting antiviral responses through the depression of interferon-stimulated gene (ISG) expression [58]. These findings highlight the importance of investigating whether lactylation modulates YTHDF2 expression or function during viral infection, which could reveal a novel layer of regulation in virus–host interactions.

Lactylation mediates immune response

Excessive lactate contributes to the establishment of a microenvironment that supports cell growth and is crucial for shaping immune cell function [59]. Lactylation, a product of lactate metabolism, has been identified as a key regulatory mechanism in innate immune responses and a modulator of macrophage polarization. Macrophages exhibit two predominant phenotypes, pro-inflammatory (M1) and anti-inflammatory (M2), each playing specific roles in different pathophysiological processes [60–62]. Zhang and colleagues discovered that M1 macrophages possess an endogenous “lactate clock” that drives the late-stage transition to M2-type characteristics through lactylation (Fig 1) [8]. This modification increases in a time-dependent manner, promoting the overexpression of M2-like genes, such as arginine enzyme 1 (Arg-1), correlating positively with histone lactylation levels [8]. Further research by Noe and colleagues proposed that early M1 macrophages with high glycolytic/low TCA activity undergo lactylation, using lactate from glycolysis to initiate the M1 to M2 phenotypic transition in tumor-associated macrophages (TAMs). As glucose decreases and lactate levels increase in the microenvironment, M2 macrophages undergo metabolic reprogramming and exhibit higher TCA activity, which supports their anti-inflammatory phenotype and promotes tissue repair and tumor progression [63].

Beyond macrophage polarization, lactylation is also critical in regulating adaptive immune responses. Regulatory T (Treg) cells are well known for suppressing anti-tumor T cell activity, thereby maintaining an immunosuppressive microenvironment [64]. Research has shown that lactate promotes Treg cell generation by the lactylation of Lys72 in MOESIN (membrane-organizing extension spike protein), subsequently enhancing the TGF-β signaling pathway to upregulate the expression of forkhead box P3 (FOXP3). In contrast, the loss of FOXP3 results in immune dysregulation, contributing to tumorigenesis [65].

The role of lactylation in viral infection

Viral infections trigger metabolic reprogramming in host cells, with enhanced glycolysis emerging as a hallmark feature. Elevated lactate levels, driven by glycolysis and LDH activity, suppress RLR-induced type-I IFN production in antiviral immunity, allowing pathogens to evade immune surveillance [7]. An intracellular surveillance protein is cyclic guanosine Monophosphate-Adenosine monophosphate (GMP-AMP) synthase (cGAS), a cytosolic DNA sensor. Lactylation at K21 of cGAS promotes tumor growth by accelerating cGAS protein degradation and suppressing interferon production through a ubiquitin-independent mechanism [66]. Emerging evidence further indicates that lactylation plays a significant role in viral infections, influencing the expression of viral and host genes, and modulating viral replication and host immune responses. In the following sections, we discuss the primary mechanisms that lactylation effects on viral infection and replication (Table 2).

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Table 2. Roles and mechanisms of lactylation in various types of viruses.

https://doi.org/10.1371/journal.ppat.1013366.t002

Hepatitis B virus (HBV)

HBV is a double-stranded DNA virus with a small viral genome (3.2 kb), belonging to the Hepadnaviridae family. If left untreated, chronic HBV infection progresses to end-stage liver disease, such as liver cirrhosis and HCC [77]. Elevated serum lactate levels, recognized as a hallmark of liver dysfunction, have been confirmed as a simple and accurate prognostic marker [78]. Clinical research revealed that lactate levels are significantly higher in non-surviving patients with acute-on-chronic liver failure (ACLF) compared to survivors, and these also serve as a predictor of six-month mortality of patients with HBV-related decompensated cirrhosis [79]. Moreover, adjusting for lactate levels enhances the prognostic accuracy of established scoring systems like MELD and Child-Pugh scores [80].

As well as acting as a prognostic marker, lactate plays an active role in the pathogenesis of HBV by promoting lactylation, thereby facilitating liver injury and HCC progression. As an intracellular pathogen, HBV relies on host cell metabolism for replication, with the viral protein HBx being central in this process. Recent studies have shown that HBx reshapes the metabolic profile of infected hepatocytes by altering glycolysis and glycogen metabolism, inducing the Warburg effect by activating the mitogen-activated protein kinases (MAPK) signaling pathway. This metabolic reprogramming enhances HCC cell proliferation, promotes metastasis, and inhibits apoptosis [81,82]. Moreover, HBx enhances aerobic glycolysis by activating the NF-κBp65/hexokinase 2 (HK2) pathway, producing excessive lactate [83]. This accumulation of lactate further drives HCC progression through multiple mechanisms. It stimulates HCC cell proliferation via the PI3K/Akt signaling pathway to induce lactylation, while disrupting key signaling pathways such as Wnt/β-catenin, MAPK, and Notch, thereby exacerbating liver injury and driving tumor progression [83]. Building on these findings, Liao and colleagues identified centromere protein A (CENPA) as a vital transcriptional regulator in HCC, which lactylation at lysine 124 enables CENPA to cooperate with YY1 to upregulate target gene expression, enhancing its activation and accelerating tumorigenesis. Notably, high expression levels of CENPA correlate with poor prognosis (Fig 2) [67].

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Fig 2. The mechanisms and therapeutic interventions of lactylation in human and animal viral pathogenesis.

https://doi.org/10.1371/journal.ppat.1013366.g002

Liver cancer stem cells (LCSCs) are considered the primary drivers of phenotypic and functional heterogeneity in HCC and are associated with elevated levels of lactylation. As with HCC cells, LCSCs exhibit heightened glycolytic metabolism, increased lactate accumulation, and elevated lactylation levels. Importantly, this modification, exemplified by H3K56la, enhances OCT4 gene expression, specifically promoting tumorigenesis and influencing the proliferative capacity and stem cell properties of LCSCs [68]. Besides histone modifications, lactylation is critical in regulating non-histone proteins within LCSCs. For instance, lactylation at the K230 and K322 sites of aldolase A (ALDOA) diminishes its binding to the dead box deconjugate enzyme 17 (DDX17), facilitating the nuclear translocation of DDX17. Once in the nucleus, DDX17 drives stem cell-like properties of LCSCs and aggravates HCC [68]. Similarly, SIRT3 has been shown to regulate HCC progression through its impact on non-histone protein lactylation (Fig 2). Jin and colleagues proved that SIRT3 suppresses HCC development by delactylating cyclin E2 (CCNE2), a lactylated substrate of SIRT3, thereby reducing its ability to promote tumor growth in HCC cells [69].

These findings collectively emphasize the significant role of lactylation as a regulator of histone and non-histone proteins in HCC cells and LCSCs, targeting it may be a promising therapeutic avenue for improving outcomes in HBV-related liver disease and HCC.

Human papillomaviruses (HPV)

HPV is a small, non-enveloped virus with a double-stranded circular DNA genome of 8 kb and encodes eight proteins, including the oncogenes E6 and E7. These oncogenes are essential for regulating cell cycle progression, inhibiting apoptosis, and evading immune response [84]. A hallmark of HPV infection is the reprogramming of host cell metabolism, which is driven by the E6 protein and facilitates rapid cell proliferation and viral replication [85]. Specifically, E6 promotes the degradation of p53, leads to increased glucose uptake through the overexpression of glucose transporter 1 (GLUT1), and further elevates glycolytic activity [86]. This metabolic shift not only promotes lactate accumulation but also alters cellular functions. Among its modulators is miR-34a, a tumor suppressor tightly regulated by p53. It exerts an anti-inflammatory effect by downregulating TNF-α and IL-6 [87] and suppresses lactate production by targeting LDHA [88]. However, HPV E6 degrades p53, resulting in decreased miR-34a levels and enhanced lactate production, ultimately fostering lactylation and potentially contributing to tumorigenesis [89]. These findings suggest that HPV-driven metabolic reprogramming enhances glycolysis and lactate production, potentially helping achieve an efficient viral replication.

The metabolic remodeling is also prominent in high-risk HPV types, such as HPV16, it accounts for more than half of cervical cancer cases [90]. Similar to other HPV types, HPV16 reprograms host metabolism by elevating glycolytic activity and lactate production, contributing to lactylation and tumor growth. In contrast to HPV E6, which promotes glycolysis by upregulating GLUT1, HPV16 oncoproteins E6 and E7 elevate lactate production by suppressing TP53-induced glycolysis and apoptosis regulator (TIGAR) expression through p53 degradation [91]. Subsequently, the accumulated lactate functions as a substrate for lactylation, impacting key cellular pathways related to tumor progression. Emerging evidence suggests that lactylation-modified proteins are predominantly enriched in glycolysis, the TCA cycle, and the pentose phosphate pathway (PPP), emphasizing the key role of lactylation in metabolic regulation [92]. The PPP is particularly noteworthy among these pathways, as it is often overactivated in tumor cells, supplying NADPH and metabolic intermediates that support cancer cell proliferation and tumor growth [93]. Notably, viral oncoproteins can directly influence PPP activity to promote tumor progression. For example, Meng and colleagues discovered that HPV16 E6 activates the PPP activity by inhibiting lactylation of glucose-6-phosphate dehydrogenase (G6PD) at K45, promoting its dimerization and increasing enzymatic activity (Fig 2). This activation of the PPP facilitates nucleotide synthesis, facilitating tumor cell proliferation [70]. Given the important role of lactylation in HPV16-driven metabolic reprogramming and tumor cell proliferation, developing new strategies to battle HPV16 infection by targeting lactylation is helpful.

Herpesvirus and poxviruses

KSHV, also called Human Herpesvirus-8 (HHV-8), is a γ-2 herpesvirus with a 165–170 kb double-stranded linear DNA genome. It is the causative agent of diseases such as Kaposi’s sarcoma (KS) and primary effusion lymphoma (PEL) [94]. A characteristic of KSHV infection is the reprogramming of host cell metabolism, with enhanced glycolysis, increased lactate production, and reduced oxidative phosphorylation [95]. This metabolic transition drives the Warburg effect during latent infection and is important for tumor growth. In support of this, targeting glycolysis with specific inhibitors is a novel therapeutic approach for inhibiting latent KSHV infection [95]. Moreover, this metabolic vulnerability is also closely linked to the function of KSHV microRNAs within the oncogenic cluster, as they drive metabolic reprogramming by decreasing oxygen consumption, increasing lactate secretion and glucose uptake, and stabilizing HIF-1α. Mechanistically, these microRNAs downregulate hypoxia-inducible factor prolyl hydroxylase 2 (EGLN2) and mitochondrial heat shock protein A9 (HSPA9), leading to reduced mitochondrial copy number, which is a metabolic shift crucial for maintaining latency and providing a growth advantage [96].

In addition to KSHV microRNAs, the viral interferon regulatory factor 1 (vIRF1) promotes lactate secretion by activating the heterogeneous nuclear ribonuclear protein Q1 (hnRNP Q1) via the recruitment of the E3 ubiquitin ligase Kelch-like 3 (KLHL3). Through this interaction, hnRNP Q1 binds to the mRNA of glycerophosphodiester phosphodiesterase domain containing 1 (GDPD1), stabilizing it at the transcript level. However, this stabilization reduces GDPD1 protein expression, ultimately contributing to vIRF1-induced aerobic glycolysis [97]. Furthermore, the E3 ubiquitin ligase K5 further accelerates glucose consumption and lactate generation by directly altering receptor tyrosine kinase (RTK) endocytosis rates and downstream signaling [98]. This metabolic reprogramming in KSHV-infected cells drives elevated lactate levels and modulates lactylation processes, including NAT10 modification (Fig 2). As a lactylation “writer,” α-tubulin acetyltransferase 1 (ATAT1) interacts with NAT10 both in vitro and in vivo. This interaction is mediated by the viral transcript PAN RNA, enhancing NAT10 lactylation, promoting tRNASer-CGA-1-1 ac4C modification, and boosting KSHV lytic protein translation and viral reactivation [99].

Further to promoting viral replication, lactylation also shapes the host’s antiviral immune responses, particularly modulating key proteins like AlkB homolog 5 (ALKBH5). Li and colleagues found that lactylation of the m6A demethylase ALKBH5 is essential for establishing an effective immune response against KSHV (Fig 2). This lactylation modification is regulated by increased interactions with acetyltransferase ESCO2 and decreased interactions with deacetyltransferase SIRT6 during viral infections. Specifically, ALKBH5 binds to IFN-β mRNA, promoting the demethylation of its m6A modifications. Overexpression of ESCO2 or depletion of SIRT6 further enhances ALKBH5 lactylation, strengthening the host’s antiviral immune response [72]. This dual role of lactylation facilitates viral replication while improving the host immune system, emphasizing its pivotal role as a regulatory hub in the dynamic interplay between KSHV and host defense mechanisms.

Monkeypox virus (MPXV), a double-stranded DNA (dsDNA) virus, belongs to the Orthopoxvirus (OPXV) genus within the Poxviridae family [100]. Recurrent human monkeypox outbreaks in Africa, along with its accidental introduction into the United States in 2003, underscore the importance of maintaining vigilance against this orthopoxvirus as a potential emerging infectious disease [101]. Moreover, ecological disturbances and the emergence of co-infections such as HIV may further facilitate MPXV transmission [102]. Notably, the mechanism by which MPXV induces lactylation closely parallels that of Kaposi’s sarcoma-associated KSHV. In both cases, ALKBH5-mediated m6A RNA modification is regulated via lactylation, impacting type I interferon (IFN-I) responses. MPXV infection has also been shown to suppress the expression of SIRT6 and impair its recruitment by ALKBH5, thereby enhancing ALKBH5 lactylation and further modulating host antiviral defenses [72].

The other two herpesviruses-human cytomegalovirus (HCMV), a β-herpesvirus with a double-stranded linear DNA genome of approximately 230 kb, and herpes simplex virus 1 (HSV-1), a dsDNA virus with a complex structure that has evolved to replicate within human neurons and epithelial cells-are also capable of inducing significant metabolic reprogramming in host cells. This metabolic reprogramming is characterized by enhanced glycolysis and a Warburg-like effect, resulting in elevated lactate production and consequently increased protein lactylation [103]. However, the mechanisms and functional consequences of lactylation differ between these two viruses. During HCMV infection, lactylation facilitates viral spread, primarily mediated by the alanyl-tRNA synthetase (AlaRS) lactyltransferase activity of AARS1, which promotes to efficient cell-to-cell transmission. Mechanistically, HCMV infection induces lactylation of gluconeogenesis proteins, while canonical glycolysis proteins, including pyruvate kinase M (PKM) at lysine 62, also exhibit upregulated lactylation sites. In contrast, during HSV-1 infection of fibroblasts, neither exogenous lactate nor oxamate significantly alters viral titers, highlighting a fundamental difference from HCMV [73]. Despite these differences, both HCMV and HSV-1 target a common host restriction factor: interferon-γ-inducible protein 16 (IFI16), a nuclear viral DNA sensor that undergoes virus-induced lactylation, albeit at distinct sites [104]. In HCMV-infected cells, IFI16 lactylation is increased at multiple residues within the intrinsically disordered region (IDR) of IFI16, particularly at lysine 128 within the nuclear localization signal, a site previously known to be acetylated by p300 [105]. Conversely, HSV-1 increases lactylation primarily within the HIN-200 domain #2 of IFI16, potentially disrupting its DNA-binding ability. Notably, both infections induce conserved lactylation at lysine 90 in the IDR, a site not previously associated with acetylation [74]. Collectively, findings from Matthew and colleagues suggest that virus-induced lactate production may contribute to immune evasion by modulating lactylation of key host immune regulators, thereby impairing antiviral signaling pathways [73].

Severe fever with thrombocytopenia syndrome virus (SFTSV)

SFTSV (Dabie bandavirus), a negative-stranded RNA virus from the Phenuiviridae family, causes severe clinical symptoms, including hemorrhagic fever and thrombocytopenia, leading to high fatality rates [106]. A clinical study found that the m6A demethylation gene FTO tended towards high expression after SFTSV infection, indicating the potential role of m6A modification in regulating viral replication and pathogenesis [107]. Beyond viral infections, m6A exerts profound effects in regulating RNA stability, localization, and translation, while also playing a pivotal role in tumor progression [108]. Recent research has shown that lactylation modulates the protein regulated by m6A. In ocular melanoma, elevated histone lactylation levels promote the expression of YTHDF2, binding to m6A-modified sites on PER1 and TP53 mRNAs, accelerating their degradation and facilitating tumor progression. Targeted correction of aberrant histone lactylation effectively inhibits tumorigenesis [51].

Similarly, m6A modification of SFTSV RNAs is crucial for regulating the virus’s infection and replication. Liu and colleagues found that SFTSV infection triggers m6A epigenetic modification in viral RNA, enabling recognition by the host’s YTHDF1 and leading to RNA degradation [75]. In response, the virus induces lactylation of YTHDF1 at the K517 and K521 sites, promoting its degradation and supporting viral replication (Fig 2). Mechanistically, ESCO1 and Sirt6 act as the “writer” and “eraser” of YTHDF1 lactylation, respectively, mutating the lactylation sites reduces YTHDF1 ubiquitination. Additionally, the virulence factor NSs of SFTSV competitively binds to Sirt6, inhibiting the formation of the Sirt6-YTHDF1 complex, which results in YTHDF1 degradation and counteracts the host’s degradation of m6A-modified viral RNAs [75]. Targeting viral-mediated lactylation of key m6A readers could enhance host defense.

Porcine reproductive and respiratory syndrome virus (PRRSV)

PRRSV is an enveloped, positive-sense single-stranded RNA virus belonging to the Arteriviridae family. Since its discovery in 1987, PRRSV has become one of the most destructive pathogens in the global swine industry, causing significant economic losses [109]. Research has shown that PRRSV infection enhances the cellular uptake of glucose and glutamine, which are critical for viral replication. Depriving cells of either glucose or glutamine significantly reduces PRRSV replication [110]. It is worth noting that PRRSV relies on glycolysis and TCA cycle activity for its replication, while excessively elevated glycolysis or TCA cycle activity disrupts viral proliferation, suggesting that a finely metabolic environment is necessary for optimal viral replication [110]. This delicate balance is closely linked to key glycolytic enzymes that regulate energy metabolism. As an essential enzyme in glycolysis, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) catalyzes the conversion of glyceraldehyde 3-phosphate to d-glycerate 1,3-bisphosphate, a crucial step in glycolysis. Inhibiting its activity could suppress aerobic glycolysis [111]. Beyond its metabolic role, GAPDH directly interacts with viral components during PRRSV infection. Liu and colleagues found that GAPDH binds to the PRRSV major envelope glycoprotein 5 (GP5), with the interaction occurring between 13-amino-acid (aa) region (aa 93–105) of GP5 and at Lys227 of GAPDH. This binding restricts GAPDH from translocating to the nucleus, redirecting it to the cytoplasm, where its glycolytic activity facilitates PRRSV replication [112].

In addition to glycolysis, PRRSV infection significantly enhances lactate production, a critical metabolite derived from glycolytic activity that directly promotes viral replication [113]. During PRRSV infection, elevated intracellular lactate levels induce lactylation of various proteins in both the cytoplasm and nucleus, thereby enabling the virus to manipulate host cell functions [110]. Pang and colleagues found that lactate is essential for optimal PRRSV proliferation and that its infection increases histone lactylation H3K18 (H3K18la) in a dose-dependent manner, underscoring its role as a key epigenetic mechanism facilitating viral immune evasion (Fig 2) [76]. Increased H3K18la directly activates HSPA6 expression, which inhibits the interaction between TNF receptor-associated factor 3 (TRAF3) and the IκB kinase subunit ε (IKKε), thus suppressing IFN-β production and weakening the host’s antiviral immune response [76]. This finding indicates that the lactate-lactylation-HSPA6 axis promotes viral growth by interfering with IFN-β induction, highlighting a potential therapeutic target for PRRSV infection.

White spot syndrome virus (WSSV)

WSSV is a large, enveloped, rod-or elliptical-shaped dsDNA virus that mainly infects shrimp, causing severe damage to the shrimp aquaculture industry [114]. During the viral genome replication stage (12 h postinfection), WSSV infection activates aerobic glycolysis, glutamine metabolism, and the PPP, which collectively increase glucose consumption and plasma lactate levels, playing a crucial role in supporting viral replication and pathogenesis [115]. This metabolic reprogramming is essential for viral replication and is linked to manipulating host enzymes to sustain infection. In this context, viral components, such as palmitic amide packaged in WSSV virions, bind to triosephosphate isomerase (TPI), enhancing its enzymatic activity, promoting glycolysis, and playing a pivotal role in initiating the viral life cycle within the host cytoplasm. This interaction leads to lactate accumulation, which upregulates HIF-1 expression and intensifies glycolysis, establishing a positive feedback loop that facilitates viral infection [116].

Beyond driving metabolic reprogramming, lactate regulates histone lactylation within host cells, creating an epigenetic environment to support viral replication. This shift in cellular metabolism is reflected in the alteration of key enzymes and histone modifications. Zhang and colleagues found that WSSV infection significantly increased the levels of key glycolytic enzymes, such as HK and LDH, in hemocytes and intestinal tissues of the infected shrimp, enhancing glycolysis and lactate accumulation, and increasing the lactylation levels of H3K18 and H4K12 [4]. In this context, shrimp p300 acted as the “writer” of lactylation, while HDAC1 and HDAC3 functioned as lysine delactylases to regulate lactylation levels. Further analysis revealed that H3K18la and H4K12la modifications were enriched in the promoters of 75 target genes, including ribosomal protein S6 kinases 2 (S6K2) (Fig 2). This upregulation of S6K2 promotes viral infection. Interestingly, the virus-encoded miR-N20 targets HIF-1α, inhibiting glycolysis and suppressing H3K18la and H4K12la modification [4]. This finding highlights the pivotal role of lactylation in WSSV infection, driving metabolic reprogramming for viral replication and maintaining lactylation levels to sustain infection persistence.

In summary, lactate-derived lactylation influences the replication and transmission of various viruses through histone and non-histone lactylation sites across multiple pathways (Fig 2).

Histone and/or non-histone lactylation regulate the proliferation and infection of human and animal viruses by driving the expression of various genes. The figure highlights potential therapeutic strategies targeting lactylation pathways in different viruses, such as HKL, oxamate, and C646, underscoring their role in inhibiting viral replication and modulating host metabolism. BioRender supported the illustration rendering portion of this work (https://www.biorender.com/).

Therapeutic potential of targeting lactylation in virus infection

The lactylation process involves lactate generation, transport, and regulation by “writers” or “erasers” all representing potential therapeutic targets [117]. To fully utilize this potential, a deeper understanding of lactylation mechanisms and their specific targets in viral infections is crucial, paving the way for developing novel treatment strategies.

Modulation of enzymes involved in lactylation

One key approach to targeting lactylation involves modulating its “writers” and “erasers”, the enzymes that regulate the addition and removal of lactylation marks. Lactyltransferases, as the “writers”, play a central role in this modification. Similarly, “erasers” like SIRT enzymes counterbalance this process by removing lactylation. These processes regulate intracellular lactylation levels by targeting specific enzymes, providing novel therapeutic opportunities. For instance, the p300/CBP inhibitor, C646 effectively suppresses WSSV replication in infection models by reducing H3K18la and H4K12la lactylation levels (Fig 2) [4]. Additionally, SIRT3, a key delactylase that targets CCNE2, has been identified as a potential therapeutic target for liver cancer treatment. Honokiol (HKL), a SIRT3 activator, directly enhances delactylation enzyme activity, effectively inhibiting HCC growth in vivo (Fig 2) [69]. This emphasizes the therapeutic potential of targeting delactylation pathways in disease treatment. Notably, lactylation shares “writers” and “erasers” with other PTMs, particularly acetylation, which increases the risk of off-target effects, highlighting the need for further investigation into drugs targeting p300 and SIRT regulation.

Modulating metabolic pathways associated with lactylation modification

Targeting the metabolism of virus-infected cells, particularly by inhibiting glycolysis or modulating other metabolic pathways, may offer an indirect strategy to reduce lactylation. For example, Demethylzeylasteral (DML), a triterpene antitumor compound, has been shown to effectively suppress the tumorigenesis of LCSCs by reducing lactate levels and inhibiting H3 histone lactylation [118]. Building on these findings, further metabolic interventions could focus on targeting specific enzymes and transporters that are critical to glycolysis and lactate metabolism, thereby modulating lactylation and its associated cellular processes.

LDH is a key enzyme in glycolysis that catalyzes the conversion of pyruvate to lactate while regenerating NAD⁺ from NADH, a critical step for sustaining glycolysis and ATP production [119]. Since lactate is the direct substrate for lactylation, inhibiting LDH reduces intracellular lactate concentrations and lowers lactylation levels. In WSSV-infected shrimp, silencing LDH significantly reduces viral loads and mortality [4]. Similarly, Oxamate, an LDH inhibitor, reduces lactate production and lactylation, impairing cellular glycolysis and restricting viral replication such as HBV, PRRSV, and SVA (Fig 2) [113,120–122]. Beyond intracellular lactate metabolism, extracellular lactate transport also plays a vital role in shaping the metabolic microenvironment. Monocarboxylate transporters (MCTs), primarily MCT1 and MCT4, regulate lactate flux across the plasma membrane, with MCT1 facilitating lactate import and MCT4 driving its export [123]. Although the role of MCT inhibitors in viral infections is unreported, targeting MCT1 has been shown to enhance anti-tumor immune cell infiltration, such as dendritic and natural killer cells, suppressing tumor growth. For example, the MCT1 inhibitor AZD3965 reduces lactate export, reprograms metabolism, and promotes immune cell activity to inhibit tumor progression [124]. Furthermore, virus-specific MCT1-deficient CD8⁺ T cells fail to establish memory T cells, which are crucial for controlling γ-herpesvirus reactivation during latency. This highlights that lactate export is essential for generating protective memory T cells needed to prevent viral latency from progressing into chronic infection [125]. Although significant progress has been made in utilizing lactylation to address viral infection, further research is essential to uncover its underlying mechanisms.