El CYP2J2 epoxigenasa tiene un nuevo papel en la inflamación vascular y biología

Inducible CYP2J2 and its product 11,12-EET promotes bacterial phagocytosis: a role for CYP2J2 deficiency in the pathogenesis of Crohn's disease?

Bystrom J, Thomson SJ, Johansson J, Edin ML, Zeldin DC, Gilroy DW, Smith AM, Bishop-Bailey D.

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Abstract

The epoxygenase CYP2J2 has an emerging role in inflammation and vascular biology. The role of CYP2J2 in phagocytosis is not known and its regulation in human inflammatory diseases is poorly understood. Here we investigated the role of CYP2J2 in bacterial phagocytosis and its expression in monocytes from healthy controls and Crohns disease patients. CYP2J2 is anti-inflammatory in human peripheral blood monocytes. Bacterial LPS induced CYP2J2 mRNA and protein. The CYP2J2 arachidonic acid products 11,12-EET and 14,15-EET inhibited LPS induced TNFα release. THP-1 monocytes were transformed into macrophages by 48h incubation with phorbol 12-myristate 13-acetate. Epoxygenase inhibition using a non-selective inhibitor SKF525A or a selective CYP2J2 inhibitor Compound 4, inhibited E. coli particle phagocytosis, which could be specifically reversed by 11,12-EET. Moreover, epoxygenase inhibition reduced the expression of phagocytosis receptorsCD11b and CD68. CD11b also mediates L. monocytogenes phagocytosis. Similar, to E. coli bioparticle phagocytosis, epoxygenase inhibition also reduced intracellular levels of L. monocytogenes, which could be reversed by co-incubation with 11,12-EET. Disrupted bacterial clearance is a hallmark of Crohn's disease. Unlike macrophages from control donors, macrophages from Crohn's disease patients showed no induction of CYP2J2 in response to E. coli. These results demonstrate that CYP2J2 mediates bacterial phagocytosis in macrophages, and implicates a defect in the CYP2J2 pathway may regulate bacterial clearance in Crohn's disease.

Inducible CYP2J2 y su producto de 11,12 -EET promueve la fagocitosis bacteriana : un papel para la deficiencia de CYP2J2 en la patogénesis de la enfermedad de Crohn ?

Bystrom J , Thomson SJ, Johansson J , Edin ML , Zeldin DC, DW Gilroy , Smith AM , Bishop- Bailey D.

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abstracto

El CYP2J2 epoxigenasa tiene un nuevo papel en la inflamación vascular y biología. El papel de CYP2J2 en la fagocitosis no es conocido y su regulación en enfermedades inflamatorias humanas es poco conocida. Aquí se investigó el papel de CYP2J2 en la fagocitosis de bacterias y su expresión en monocitos de los controles sanos y los pacientes con enfermedad de Crohn. CYP2J2 es anti - inflamatoria en monocitos de sangre periférica humanos . LPS bacterianos inducidos ARNm y la proteína CYP2J2 . El CYP2J2 productos del ácido araquidónico 11,12 -EET y 14,15 -EET inhibidos LPS liberación TNFa inducida. Monocitos THP-1 se transforman en macrófagos mediante la incubación 48h con forbol 12 - miristato 13 -acetato . Epoxigenasa inhibición utilizando un SKF525A inhibidor no selectivo o un compuesto inhibidor selectivo CYP2J2 4 , inhibe la fagocitosis de partículas de E. coli , lo que podría ser revertido específicamente por 11,12 -EET . Por otra parte , la inhibición epoxigenasa reduce la expresión de receptores de fagocitosis CD11b y CD68 . CD11b también media la fagocitosis L. monocytogenes . Similares , para E. coli biopartícula fagocitosis , la inhibición epoxigenasa también reduce los niveles intracelulares de L. monocytogenes , que podrían ser revertidos por la co -incubación con 11,12 -EET . Interrumpió el aclaramiento de bacterias es un sello distintivo de la enfermedad de Crohn . A diferencia de los macrófagos de donantes de control , los macrófagos de pacientes con enfermedad de Crohn no mostraron inducción de CYP2J2 en respuesta a E. coli . Estos resultados demuestran que CYP2J2 media la fagocitosis bacteriana en macrófagos , e implica un defecto en la vía CYP2J2 puede regular el aclaramiento de bacterias en la enfermedad de Crohn

Removal of misincorporated ribonucleotides from prokaryotic genomes: an unexpected role for nucleotide excision repair.

Vaisman A, McDonald JP, Huston D, Kuban W, Liu L, Van Houten B, Woodgate R.

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Abstract

Stringent steric exclusion mechanisms limit the misincorporation of ribonucleotides by high-fidelity DNA polymerases into genomic DNA. In contrast, low-fidelity Escherichia coli DNA polymerase V (pol V) has relatively poor sugar discrimination and frequently misincorporates ribonucleotides. Substitution of a steric gate tyrosine residue with alanine (umuC_Y11A) reduces sugar selectivity further and allows pol V to readily misincorporate ribonucleotides as easily as deoxynucleotides, whilst leaving its poor base-substitution fidelity essentially unchanged. However, the mutability of cells expressing the steric gate pol V mutant is very low due to efficient repair mechanisms that are triggered by the misincorporated rNMPs. Comparison of the mutation frequency between strains expressing wild-type and mutant pol V therefore allows us to identify pathways specifically directed at ribonucleotide excision repair (RER). We previously demonstrated that rNMPs incorporated by umuC_Y11A are efficiently removed from DNA in a repair pathway initiated by RNase HII. Using the same approach, we show here that mismatch repair and base excision repair play minimal back-up roles in RER in vivo. In contrast, in the absence of functional RNase HII, umuC_Y11A-dependent mutagenesis increases significantly in ΔuvrA, uvrB5 and ΔuvrC strains, suggesting that rNMPs misincorporated into DNA are actively repaired by nucleotide excision repair (NER) in vivo. Participation of NER in RER was confirmed by reconstituting ribonucleotide-dependent NER in vitro. We show that UvrABC nuclease-catalyzed incisions are readily made on DNA templates containing one, two, or five rNMPs and that the reactions are stimulated by the presence of mispaired bases. Similar to NER of DNA lesions, excision of rNMPs proceeds through dual incisions made at the 8(th) phosphodiester bond 5' and 4(th)-5(th) phosphodiester bonds 3' of the ribonucleotide. Ribonucleotides misinserted into DNAcan therefore be added to the broad list of helix-distorting modifications that are substrates for NER.

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