Disease relevance of RNASE2

• In addition, transferrin-targeted EDN exhibited tumor cell toxicities similar to those of hRNase [1].
• The DC chemotactic activity of EDN was inhibited by either pretreatment of DCs with pertussis toxin or by simultaneous addition of placental RNase inhibitor to inhibit the activity of EDN [2].
• EDN is also a potent helminthotoxin and can mediate antiviral activity of eosinophils against isolated virions of the respiratory syncytial virus [3].
• EDN is a distinct cationic protein of the eosinophil's large specific granule known primarily for its ability to induce ataxia, paralysis, and central nervous system cellular degeneration in experimental animals (Gordon phenomenon) [4].
• Evolution of antiviral activity in the ribonuclease A gene superfamily: evidence for a specific interaction between eosinophil-derived neurotoxin (EDN/RNase 2) and respiratory syncytial virus [5].

High impact information on RNASE2

• The EDN/ECP gene pair arose from a recent duplication event that occurred after the divergence of New World and Old World monkeys [6].
• We have traced the rapid molecular evolution of eosinophil-derived neurotoxin (EDN) and eosinophil cationic protein (ECP), two host defense proteins that are members of the mammalian ribonuclease gene family [6].
• All three patients had elevated serum and urinary levels of this protein and eosinophil-derived neurotoxin, indicative of eosinophil degranulation [7].
• These 14-d cord blood-derived cell cultures showed strong hybridization signals for eosinophil-derived neurotoxin by RNA blot analysis and contained 78 ng histamine per 10(6) cells [8].
• In contrast, mRNA for EDN was detected in uninduced HL-60 cells and was upregulated in HL-60 cells induced with DMSO [9].

Chemical compound and disease context of RNASE2

• In rats subjected to chronic hypoxia (10% O2) for up to 10 wk, the two electrophysiological abnormalities had developed by 4 wk and were very similar in degree to those seen in EDN and EGN [10].
• The cardinal electrophysiological abnormalities in experimental diabetic (EDN) and experimental galactose (EGN) neuropathy, models in which endoneurial hypoxia has been demonstrated, are a slowing in nerve conduction velocity (NCV) and a resistance to ischemic conduction block (RICB) [10].

Biological context of RNASE2

• We mutated two human ribonucleases-pancreatic RNase (hRNAse) and eosinophil-derived neurotoxin (EDN)-to incorporate cysteine residues at putative sites of close contact to RI, but distant from the catalytic sites [1].
• Amino acid sequence analyses showed that ECP-1 and ECP-2 are identical from residue 1 through residue 59 and that the sequences of EDN and ECP are highly homologous (37 of 55 residues identical) [11].
• Neurologic abnormalities of onconase-treated animals were indistinguishable from those of EDN-treated animals, and histology showed dramatic Purkinje cell loss in the brains of onconase-treated animals [12].
• In contrast, the homologous bovine pancreatic RNase A injected intraventricularly at a dose 5000 times greater than the LD50 dose of EDN or onconase is not toxic and does not cause the Gordon phenomenon [12].
• We have previously shown that optimal expression of the EDN gene is dependent on an interaction between an intronic enhancer element or elements and the 5' promoter region [13].

Anatomical context of RNASE2

• Biochemical and functional similarities between human eosinophil-derived neurotoxin and eosinophil cationic protein: homology with ribonuclease [11].
• EDN, ECP-1, and ECP-2 had neurotoxic activity, causing the Gordon phenomenon at doses down to 0.15 micrograms when injected into the cisterna magna; the proteins were comparable in their activities [11].
• EDN was not chemotactic for leukocytes other than DCs [2].
• We found that EDN was selectively chemotactic for dendritic cells (DCs) [2].
• We have found that another member of the RNase superfamily, an antitumor protein called onconase, isolated from Rana pipiens oocytes and early embryos, will also cause the Gordon phenomenon when injected into the cerebrospinal fluid of guinea pigs at a dose similar to that of EDN (LD50, 3-4 micrograms) [12].

Associations of RNASE2 with chemical compounds

• We have now determined high resolution (1.8 A) crystal structures for EDN in complex with adenosine-3',5'-diphosphate (3',5'-ADP), adenosine-2',5'-di-phosphate (2',5'-ADP), adenosine-5'-diphosphate (5'-ADP) as well as for a native structure in the presence of sulfate refined at 1.6 A [3].
• C2-alpha-Mannosyltryptophan was discovered in human RNase 2, an enzyme that occurs in eosinophils and is involved in host defense [14].
• However, A23187 at a concentration (1 microgram/ml) that caused 15% EDN release and 30% CLC protein release also produced release of the cytoplasmic enzyme lactic dehydrogenase (LDH) and loss of cell viability, both of which were calcium dependent [15].
• After trifluoromethanesulfonic acid treatment, the molecular weight of RNase UpI-2 was reduced and approached that of nonsecretory RNase, which indicated that the protein contains a significant amount of carbohydrate (approximately 50%) [16].
• EDN conformation was not greatly affected by iodoacetate treatment since interaction of the modified protein with antibodies was only slightly altered [17].

Physical interactions of RNASE2

• Simultaneous substitutions of three neighboring tryptophans (261, 263, and 318) on RI attenuate affinity even more dramatically (by 4900-fold), indicating that the interactions of this RI region also contribute a considerable amount of the binding energy for the EDN complex [18].

Regulatory relationships of RNASE2

• Finally, WAY103 inhibited VCAM-1-stimulated superoxide generation but enhanced cytokine-activated eosinophil-derived neurotoxin degranulation [19].
• Priming of eosinophils with IL-5 enhanced CCR3 ligand-induced degranulation but did not cause non-CCR3 ligands to induce eosinophil-derived neurotoxin release [20].
• Novel co-operation between eotaxin and substance-P in inducing eosinophil-derived neurotoxin release [21].
• IRp60 is constitutively expressed on mast cells but is down-regulated in vitro by the eosinophil proteins major basic protein and eosinophil-derived neurotoxin [22].
• GM-CSF also enhanced superoxide production and eosinophil-derived neurotoxin release stimulated by the lower concentrations of immobilized lactoferrin [23].

Other interactions of RNASE2

• Coupling of Cys89 of RNase and Cys87 of EDN to proteins at these sites via a thioether bond produced enzymatically active conjugates that were resistant to RI [1].
• Radioimmunoassay, using monoclonal antibodies, of fractions from the heparin-Sepharose chromatography showed one peak of EDN activity and two peaks of ECP activity (termed ECP-1 and ECP-2) [11].
• The uneventful trends observed for RNase k6 serve to spotlight the unique nature of EDN and ECP and the unusual evolutionary constraints to which these two ribonuclease genes must be responding [24].
• Expression of chimeras of RNase 2 and nonglycosylated RNase 4 and deletion mutants in HEK293 cells identified residues 1-13 to be sufficient for C-mannosylation [14].
• We conclude that ECP and EPX/EDN may be used to monitor antiinflammatory treatment in asthmatic patients, and that smoking asthmatic subjects are resistant to inhaled corticosteroids [25].

Analytical, diagnostic and therapeutic context of RNASE2

• Eosinophil-derived neurotoxin (EDN) and eosinophil cationic protein (ECP) were isolated from lysates of human eosinophil granules by gel filtration and ion exchange chromatography on heparin-Sepharose [11].
• By indirect immunofluorescence, both eosinophils and neutrophils stained with antibodies to EDN and ECP [26].
• Molecular cloning of the human eosinophil-derived neurotoxin: a member of the ribonuclease gene family [4].
• EDN mRNA was assessed by real-time PCR [27].
• CONCLUSIONS: Specific elevated posttranslationally modified urinary EDN and osteopontin COOH-terminal fragments in ovarian cancer might lead to potential noninvasive screening tests for early diagnosis [28].

References