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Relationship: 1017

Title

A descriptive phrase which clearly defines the two KEs being considered and the sequential relationship between them (i.e., which is upstream, and which is downstream). More help

Interference, nuclear localization of NFAT leads to Reduction, NFAT/AP-1 complex formation

Upstream event

The causing Key Event (KE) in a Key Event Relationship (KER). More help

Interference, nuclear localization of NFAT

Downstream event

The responding Key Event (KE) in a Key Event Relationship (KER). More help

Reduction, NFAT/AP-1 complex formation

Key Event Relationship Overview

The utility of AOPs for regulatory application is defined, to a large extent, by the confidence and precision with which they facilitate extrapolation of data measured at low levels of biological organisation to predicted outcomes at higher levels of organisation and the extent to which they can link biological effect measurements to their specific causes.Within the AOP framework, the predictive relationships that facilitate extrapolation are represented by the KERs. Consequently, the overall WoE for an AOP is a reflection in part, of the level of confidence in the underlying series of KERs it encompasses. Therefore, describing the KERs in an AOP involves assembling and organising the types of information and evidence that defines the scientific basis for inferring the probable change in, or state of, a downstream KE from the known or measured state of an upstream KE. More help

AOPs Referencing Relationship

AOP Name	Adjacency	Weight of Evidence	Quantitative Understanding	Point of Contact	Author Status	OECD Status
Inhibition of Calcineurin Activity Leading to Impaired T-Cell Dependent Antibody Response	adjacent	High	High	Cataia Ives (send email)	Open for comment. Do not cite	WPHA/WNT Endorsed

Taxonomic Applicability

Latin or common names of a species or broader taxonomic grouping (e.g., class, order, family) that help to define the biological applicability domain of the KER.In general, this will be dictated by the more restrictive of the two KEs being linked together by the KER. More help

Term	Scientific Term	Evidence	Link
Homo sapiens	Homo sapiens	High	NCBI
Mus musculus	Mus musculus	High	NCBI

Sex Applicability

An indication of the the relevant sex for this KER. More help

Sex	Evidence
Unspecific	High

Life Stage Applicability

An indication of the the relevant life stage(s) for this KER. More help

Term	Evidence
All life stages	High

Key Event Relationship Description

Provides a concise overview of the information given below as well as addressing details that aren’t inherent in the description of the KEs themselves. More help

Activated (dephosphorylated) nuclear factor of activated T cells (NFAT) is translocated into the nucleus through the molecular changes of exposing nuclear localization signal (NLS) and concomitant masking of nuclear export signal (NES) due to dephosphorylation of the SP motifs of NFAT. (Matsuda and Koyasu 2000, Zhu and McKeon 1999).

In the nucleus NFAT binds with AP 1 at the IL-2 promoter region, (Schreiber and Crabtree 1992; Jain et al. 1992) and induces transcription of IL-2 (Jain et al. 1993). In addition to IL-2, NFAT localized in the nucleus of T cells also binds to the promoter region of the other classes of cytokines including IL-4 and IL-13.

Once CN phosphatase activity is inhibited, dephosphorylation of NFAT and subsequent nuclear localization of NFAT decreases, which results in a decrease of NFAT/AP-1 complex formation at the cytokine promoter sites (Rao et al. 1997).

Evidence Collection Strategy

Include a description of the approach for identification and assembly of the evidence base for the KER. For evidence identification, include, for example, a description of the sources and dates of information consulted including expert knowledge, databases searched and associated search terms/strings. Include also a description of study screening criteria and methodology, study quality assessment considerations, the data extraction strategy and links to any repositories/databases of relevant references.Tabular summaries and links to relevant supporting documentation are encouraged, wherever possible. More help

Evidence Map 2.0

ID	Experimental Design	Species	Upstream Observation	Downstream Observation	Citation (first author, year)	Notes

Evidence Map

Addresses the scientific evidence supporting KERs in an AOP setting the stage for overall assessment of the AOP. More help

Title	First Author	Biological Plausibility	Dose Concordance	Temporal Concordance	Incidence Concordance

Biological Plausibility

Dose Concordance Evidence

Temporal Concordance Evidence

Incidence Concordance Evidence

Uncertainties and Inconsistencies

Addresses inconsistencies or uncertainties in the relationship including the identification of experimental details that may explain apparent deviations from the expected patterns of concordance. More help

Nothing especially

Known modulating factors

This table captures specific information on the MF, its properties, how it affects the KER and respective references.1.) What is the modulating factor? Name the factor for which solid evidence exists that it influences this KER. Examples: age, sex, genotype, diet 2.) Details of this modulating factor. Specify which features of this MF are relevant for this KER. Examples: a specific age range or a specific biological age (defined by...); a specific gene mutation or variant, a specific nutrient (deficit or surplus); a sex-specific homone; a certain threshold value (e.g. serum levels of a chemical above...) 3.) Description of how this modulating factor affects this KER. Describe the provable modification of the KER (also quantitatively, if known). Examples: increase or decrease of the magnitude of effect (by a factor of...); change of the time-course of the effect (onset delay by...); alteration of the probability of the effect; increase or decrease of the sensitivity of the downstream effect (by a factor of...) 4.) Provision of supporting scientific evidence for an effect of this MF on this KER. Give a list of references. More help

At present, no evidence is found.

Quantitative Understanding of the Linkage

Captures information that helps to define how much change in the upstream KE, and/or for how long, is needed to elicit a detectable and defined change in the downstream KE. More help

Response-response Relationship

Provides sources of data that define the response-response relationships between the KEs. More help

The relationship of the interference of nuclear localization of NFAT leading to reduced NFAT/AP-1 complex formation bound at the promoter sites of cytokine genes in the presence of CNIs is well known as mentioned above.

KE1:

Dose-dependent interference with nuclear translocation of NFAT1 was observed with increasing FK506 concentrations from 0.01nM (Jarkat T cells) up to 1 μM (1000 nM). Higher concentrations induced cellular toxicity and resulted in cell death. Dose-dependent interference of nuclear NFAT1 translocation per CN inhibition was also observed in CD4+ T cells from healthy donors, again from 10nM to maximal concentrations of 1 μM (Maguire et al. 2013). Both parameters were measured after 2 hour culture of T cells with FK506.

KE2:

Reduction in generation of NFAT/AP-1 complexes can be detected using a gel shift assay (Rao et al. 1997, Jain et al. 1992, Jain et al. 1993).

Decreased NFAT translocated to the nucleus, induced by FK506 at 100ng/mL (124nM) or CsA at 500ng/mL (416nM) after 2 hours treatment, hinders the formation of the functional NFAT/AP-1 complexes necessary to binding at the site of IL-2 promoters (Flanagan et al. 1991). As mentioned above, gel mobility shift assays also showed that NFAT/AP-1 complexes were formed only in the nucleus after T cell activation with unchanged preexisting NFAT in the cytoplasm and that treatment of T cells with 1µM FK506 led to decease the levels of NFAT/AP-1 complex (Jain et al. 1992).

These findings suggest that nuclear translocation of NFAT after T cell stimulation is strongly related to the complex formation with AP-1 in the nucleus, and FK506 was shown to inhibit NFAT/AP-1 complex formation in the nucleus at the concentrations within the concentration range of FK506 for suppressing nuclear translocation of NFAT (Maguire et al. 2013).

Time-scale

Information regarding the approximate time-scale of the changes in KEdownstream relative to changes in KEupstream (i.e., do effects on KEdownstream lag those on KEupstream by seconds, minutes, hours, or days?). More help

Nuclear translocation of NFAT was shown to be inhibited in vitro using imaging flowcytometry after 2 hours culture of T cells with FK506 (Maguire et al. 2013), and gel mobility shift assays revealed the inhibition of nuclear translocation of NFAT and following complex formation with AP-1 within the nucleus after 2 hours culture of T cells with FK506 (Jain et al. 1992, Flanagan et al. 1991).

Known Feedforward/Feedback loops influencing this KER

Define whether there are known positive or negative feedback mechanisms involved and what is understood about their time-course and homeostatic limits. More help

At present, no evidence is found.

Domain of Applicability

A free-text section of the KER description that the developers can use to explain their rationale for the taxonomic, life stage, or sex applicability structured terms. More help

NFAT expresses in B cells, mast cells, neutrophils, granulocytes, dendritic cells, macrophages, and natural killer cells as well as T cells from humans, rodents, and other mammalian species (Rao et al. 1997).

CN-NFAT system functionality is common among mammalian species, including humans and rodents. It is also possible that FK506-induced interference with NFAT/AP-1 complex formation at the promoter site of the IL-2 gene is common among mammalian T cells, including those of humans and rodents (Flanagan et al. 1991).