SIRT2 regulates macrophage chemotaxis by de-modifying histone H4K8 lactylation

doi:10.3969/j.issn.1674-8115.2023.08.008

Abstract

Abstract:

Objective ·To explore the regulatory role of silent information regulator 2 (SIRT2) in modulating the immune phenotype of macrophages after infection by removing the lactylation at H4K8 site of histone and the corresponding mechanism. Methods ·Human THP-1 leukemia cells were induced by phorbol 12-myristate 13-acetate (PMA) and stimulated by lipopolysaccharide (LPS) to establish a macrophage infection model. Macrophages without LPS treatment (pTHP-1) were set as the control (CTRL) group, and macrophages with LPS treatment were set as the infected (LPS) group. Western blotting was used to detect the level of histone modification and SIRT2 protein in macrophages. RT-qPCR was used to detect the expression level of glycolytic key enzymes [phosphofructokinase liver type (PFKL), lactate dehydrogenase A (LDHA)] and modulators genes hypoxia inducible factor 1α (HIF-1α), and the expression level of Sirtuin genes and HDAC genes between the two groups. Transwell was used to detect the ability of macrophage chemotaxis. Lentivirus packaging and cell infection were used to construct SIRT2 overexpression cell line. The interaction analysis method of RNA sequencing (RNA-seq) and chromatin immunoprecipitation sequencing (ChIP-seq) was used to analyze the difference and pathway enrichment of the genes specifically bound to H4K8 lactylation (H4K8la). Results ·Compared to the CTRL group, macrophage glycolysis was upregulated and the level of H4K8la was significantly increased in the LPS group (P<0.05), while the level of lactylation in other sites remained unchanged. Among all known enzymes with deacetylation modification function, only SIRT2 showed a significant decrease after LPS treatment (P<0.05), and overexpression of SIRT2 could significantly inhibit the level of H4K8la modification, while the level of H4K8ac remained unchanged (P>0.05). The interactive analysis of ChIP-seq and RNA-seq revealed that chemotaxis-related genes were regulated by H4K8la, and macrophage chemotaxis ability significantly decreased after the overexpression of SIRT2 and downregulation of H4K8la (P<0.05). Conclusion ·SIRT2 can change the expression of target genes related to chemotaxis by removing H4K8la modification, thereby reducing the chemotaxis ability of macrophages. Targeting SIRT2 and H4K8la modification may help control inflammation mediated by macrophages.

Key words: macrophage, chemotaxis, lactylation, post-translational modification, silent information regulator 2 (SIRT2)

CLC Number:

R725.1

SONG Wenting, TAO Yue, PAN Yi, MO Xi, CAO Qing. SIRT2 regulates macrophage chemotaxis by de-modifying histone H4K8 lactylation[J]. Journal of Shanghai Jiao Tong University (Medical Science), 2023, 43(8): 1008-1016.

Figures/Tables 8

TrendMD

References 30

1	GALLI G, SALEH M. Immunometabolism of macrophages in bacterial infections[J]. Front Cell Infect Microbiol, 2021, 10: 607650.
2	VIOLA A, MUNARI F, SÁNCHEZ-RODRÍGUEZ R, et al. The metabolic signature of macrophage responses[J]. Front Immunol, 2019, 10: 1462.
3	PETER K, REHLI M, SINGER K, et al. Lactic acid delays the inflammatory response of human monocytes[J]. Biochem Biophys Res Commun, 2015, 457(3): 412-418.
4	ERREA A, CAYET D, MARCHETTI P, et al. Lactate inhibits the pro-inflammatory response and metabolic reprogramming in murine macrophages in a GPR81-independent manner[J]. PLoS One, 2016, 11(11): e0163694.
5	NAREIKA A, HE L, GAME B A, et al. Sodium lactate increases LPS-stimulated MMP and cytokine expression in U937 histiocytes by enhancing AP-1 and NF-κB transcriptional activities[J]. Am J Physiol Endocrinol Metab, 2005, 289(4): E534-E542.
6	SAMUVEL D J, SUNDARARAJ K P, NAREIKA A, et al. Lactate boosts TLR4 signaling and NF-κB pathway-mediated gene transcription in macrophages via monocarboxylate transporters and MD-2 up-regulation[J]. J Immunol, 2009, 182(4): 2476-2484.
7	XU H W, WU M Y, MA X M, et al. Function and mechanism of novel histone posttranslational modifications in health and disease[J]. Biomed Res Int, 2021, 2021: 6635225.
8	MOHAMMADI A, SHARIFI A, POURPAKNIA R, et al. Manipulating macrophage polarization and function using classical HDAC inhibitors: implications for autoimmunity and inflammation[J]. Crit Rev Oncol, 2018, 128: 1-18.
9	ZHANG D, TANG Z Y, HUANG H, et al. Metabolic regulation of gene expression by histone lactylation[J]. Nature, 2019, 574(7779): 575-580.
10	MURRAY P J, WYNN T A. Protective and pathogenic functions of macrophage subsets[J]. Nat Rev Immunol, 2011, 11(11): 723-737.
11	NEWSHOLME P, GORDON S, NEWSHOLME E A. Rates of utilization and fates of glucose, glutamine, pyruvate, fatty acids and ketone bodies by mouse macrophages[J]. Biochem J, 1987, 242(3): 631-636.
12	PARK D, LIM G, YOON S J, et al. The role of immunomodulatory metabolites in shaping the inflammatory response of macrophages[J]. BMB Rep, 2022, 55(11): 519-527.
13	COLEGIO O R, CHU N Q, SZABO A L, et al. Functional polarization of tumour-associated macrophages by tumour-derived lactic acid[J]. Nature, 2014, 513(7519): 559-563.
14	DIETL K, RENNER K, DETTMER K, et al. Lactic acid and acidification inhibit TNF secretion and glycolysis of human monocytes[J]. J Immunol, 2010, 184(3): 1200-1209.
15	AWASTHI D, NAGARKOTI S, SADAF S, et al. Glycolysis dependent lactate formation in neutrophils: a metabolic link between NOX-dependent and independent NETosis[J]. Biochim Biophys Acta Mol Basis Dis, 2019, 1865(12): 165542.
16	ALARCÓN P, MANOSALVA C, CONEJEROS I, et al. D (-) lactic acid-induced adhesion of bovine neutrophils onto endothelial cells is dependent on neutrophils extracellular traps formation and CD11b expression[J]. Front Immunol, 2017, 8: 975.
17	WU D, SHI Y X, ZHANG H, et al. Epigenetic mechanisms of Immune remodeling in sepsis: targeting histone modification[J]. Cell Death Dis, 2023, 14(2): 112.
18	CHEN L H, HUANG L X, GU Y, et al. Lactate-lactylation hands between metabolic reprogramming and immunosuppression[J]. Int J Mol Sci, 2022, 23(19): 11943.
19	IRIZARRY-CARO R A, MCDANIEL M M, OVERCAST G R, et al. TLR signaling adapter BCAP regulates inflammatory to reparatory macrophage transition by promoting histone lactylation[J]. Proc Natl Acad Sci USA, 2020, 117(48): 30628-30638.
20	KOVACS L, CAO Y P, HAN W H, et al. PFKFB3 in smooth muscle promotes vascular remodeling in pulmonary arterial hypertension[J]. Am J Respir Crit Care Med, 2019, 200(5): 617-627.
21	WANG N X, WANG W W, WANG X Q, et al. Histone lactylation boosts reparative gene activation post-myocardial infarction[J]. Circ Res, 2022, 131(11): 893-908.
22	MA W, AO S, ZHOU J, et al. Methylsulfonylmethane protects against lethal dose MRSA-induced sepsis through promoting M2 macrophage polarization[J]. Mol Immunol, 2022, 146: 69-77.
23	CHU X, DI C Y, CHANG P P, et al. Lactylated histone H3K18 as a potential biomarker for the diagnosis and predicting the severity of septic shock[J]. Front Immunol, 2022, 12: 786666.
24	MORENO-YRUELA C, ZHANG D, WEI W, et al. Class Ⅰ histone deacetylases (HDAC1-3) are histone lysine delactylases[J]. Sci Adv, 2022, 8(3): eabi6696.
25	DAI H, SINCLAIR D A, ELLIS J L, et al. Sirtuin activators and inhibitors: promises, achievements, and challenges[J]. Pharmacol Ther, 2018, 188: 140-154.
26	WANG Y, YANG J, HONG T, et al. SIRT2: controversy and multiple roles in disease and physiology[J]. Ageing Res Rev, 2019, 55: 100961.
27	XU H, YU X, WANG B, et al. The clinical significance of the SIRT2 expression level in the early stage of sepsis patients[J]. Ann Palliat Med, 2020, 9(4): 1413-1419.
28	SASSO G L, MENZIES K J, MOTTIS A, et al. SIRT2 deficiency modulates macrophage polarization and susceptibility to experimental colitis[J]. PLoS One, 2014, 9(7): e103573.
29	ZU H X, LI C, DAI C R, et al. SIRT2 functions as a histone delactylase and inhibits the proliferation and migration of neuroblastoma cells[J]. Cell Discov, 2022, 8(1): 54.
30	TU Q Q, YU X Y, XIE W, et al. Prokineticin 2 promotes macrophages-mediated antibacterial host defense against bacterial pneumonia[J]. Int J Infect Dis, 2022, 125: 103-113.

Gene	Forward primer (5´→3´)	Reverse primer (5´→3´)
HIF-1α	GAACGTCGAAAAGAAAAGTCTCG	CCTTATCAAGATGCGAACTCACA
PFKL	GTACCTGGCGCTGGTATCTG	CCTCTCACACATGAAGTTCTCC
LDHA	ATGGCAACTCTAAAFFATCAGC	CCAACCCCAACAACTGTAATCT
SIRT1	AGGCCACGGATAGGTCCATA	GTGGAGGTATTGTTTCCGGC
SIRT2	TGCGGAACTTATTCTCCCAGA	GAGAGCGAAAGTCGGGGAT
SIRT3	TGCTCATCAACCGGGACTTG	TTGTCTGGTCCATCAAGCCTA
SIRT5	CTCAAGATGCCAGCATCCCA	AGGAAGTGCCCACCACTAGA
HDAC1	CATCGCTGTGAATTGGGCTG	ACCCTCTGGTGATACTTTAGCAG
HDAC2	TCTGCTACTACTACGACGGTGA	TCATTTCTTCGGCAGTGGCT
HDAC3	CATGACGGTGTCCTTCCACA	CAGAGTCAGCTCCACACTGG
GAPDH	TCTCCTCTGACTTCAACAGCGACA	CCCTGTTGCTGTAGCCAAATTCGT

Gene	Forward primer (5´→3´)	Reverse primer (5´→3´)
HIF-1α	GAACGTCGAAAAGAAAAGTCTCG	CCTTATCAAGATGCGAACTCACA
PFKL	GTACCTGGCGCTGGTATCTG	CCTCTCACACATGAAGTTCTCC
LDHA	ATGGCAACTCTAAAFFATCAGC	CCAACCCCAACAACTGTAATCT
SIRT1	AGGCCACGGATAGGTCCATA	GTGGAGGTATTGTTTCCGGC
SIRT2	TGCGGAACTTATTCTCCCAGA	GAGAGCGAAAGTCGGGGAT
SIRT3	TGCTCATCAACCGGGACTTG	TTGTCTGGTCCATCAAGCCTA
SIRT5	CTCAAGATGCCAGCATCCCA	AGGAAGTGCCCACCACTAGA
HDAC1	CATCGCTGTGAATTGGGCTG	ACCCTCTGGTGATACTTTAGCAG
HDAC2	TCTGCTACTACTACGACGGTGA	TCATTTCTTCGGCAGTGGCT
HDAC3	CATGACGGTGTCCTTCCACA	CAGAGTCAGCTCCACACTGG
GAPDH	TCTCCTCTGACTTCAACAGCGACA	CCCTGTTGCTGTAGCCAAATTCGT

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