This Key Event Relationship is licensed under the Creative Commons BY-SA license. This license allows reusers to distribute, remix, adapt, and build upon the material in any medium or format, so long as attribution is given to the creator. The license allows for commercial use. If you remix, adapt, or build upon the material, you must license the modified material under identical terms.

Relationship: 3367

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

Increased, SREBP2 protein expression leads to Increased, cholesterol synthesis enzymes

Upstream event

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

Increased, SREBP2 protein expression

Downstream event

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

Increased, cholesterol synthesis enzymes

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
Activation, Pregnane-X receptor, NR1l2 leads to increased plasma low-density lipoprotein (LDL) cholesterol via increased cholesterol synthesis	adjacent	High		John Frisch (send email)	Under development: Not open for comment. Do not cite

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
mammals	mammals	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	Moderate

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

Sterol Regulatory Element Binding Transcription Protein 2 (SREBP2) is synthesized as an inactive precursor protein, with inactive form located in the endoplasmic reticulum membrane (Horton et al. 2002). SREBP2 has an important role in lipid synthesis regulation. At low cholesterol levels, free SREBP cleavage-activating protein (SCAP) binds to Coat Protein Complex II (COPII; Ouyang et al. 2020). The SCAP-COPII complex enables Sterol Regulatory Element Binding Proteins (SREBPs) to move through the endoplasmic reticulum to the Golgi, where membrane-bound transcription factor site-1-protease (S1P) and site-2-protease (S2P) enable proteolytic processing that allows SREBPs to enter the nucleus (Yabe et al. 2002; Yang et al. 2002). In the nucleus, SREBP2 increases gene expression for enzymes involved in the mevalonate pathway of cholesterol synthesis (Sakakura et al. 2001; Ouyang et al. 2020; Itkonen et al., 2023) including not only rate-limited enzymes including 3-hydroxy-3-methylglutarylcoenzyme (HMGCR) (Sakakura et al. 2001; Itkonen et al. 2023) but many enzymes in the pathway: mevalonate kinase, mevalonate pyrophosphate decarboxylase, isopentenyl phosphate isomerase, geranylgeranyl pyrophosphate synthase, farnesyl pyrophosphate synthase, squalene synthase, squalene epoxidase, lanosterol synthase, lanosterol demethylase, and 7-dehydro-cholesterol reductase (Sakakura et al. 2001; Horton et al. 2002). At high cholesterol levels, INSIG1 binds to SCAP, competitively inhibiting the ability of SCAP to bind to COPII (Ouyang et al. 2020). SREBPs are retained in the endoplasmic reticulum rather than being transferred to the Golgi, reducing levels of cholesterol synthesis (Ouyang et al. 2020; Itkonen et al., 2023).

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

This Key Event Relationship was developed as part of an Environmental Protection Agency effort to represent putative AOPs from peer-reviewed literature which were heretofore unrepresented in the AOP-Wiki. Itkonen et al. (2023) focused on identifying Adverse Outcome Pathways that linked PXR activation to increased level of plasma low-density lipoprotein (LDL) cholesterol through review of existing literature, and provided initial network analysis.

Cited empirical studies are focused on increased SREBP2 expression and resulting increased cholesterol synthesis enzymes in mammals, in support of development of AOP 545 for Itkonen et al. (2023) content.

Authors of KER 3367 did a further evaluation of published peer-reviewed literature to provide additional evidence in support of the key event relationship.

Evidence Supporting this KER

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

Biological Plausibility

Addresses the biological rationale for a connection between KEupstream and KEdownstream. This field can also incorporate additional mechanistic details that help inform the relationship between KEs, this is useful when it is not practical/pragmatic to represent these details as separate KEs due to the difficulty or relative infrequency with which it is likely to be measured. More help

Sterol Regulatory Element Binding Protein-2 (SREBP2) and cholesterol synthesis enzymes have been studied in a variety of gene-knockout, gene transfection, and diet studies designed to disrupt maintenance of lipid homeostasis in laboratory mammals. Evidence from gene expression and protein expression studies show a consistent response in increase of SREBP2 activity leading to an increase in cholesterol synthesis enzymes abundance and activity. In addition, study of cholesterol and lipid levels, as well as binding/activation of SREBP by SREBP cleavage-activating protein (SCAP), help to understand the mechanism for regulation of cholesterol synthesis in this key event relationship.

Empirical Evidence

Provides specific (citable) evidence that supports the idea of a change in the upstream KE (KEupstream) leading to, or being associated with, a subsequent change in the downstream KE (KEdownstream), assuming the perturbation of KEupstream is sufficient. More help

Species	Duration	Dose	Increased SREBP2 activity?	Increased cholesterol synthesis enzymes?	Summary	Citation
Mouse (Mus musculus)	12-16 weeks	Transgenic mice that overexpress SREBP2.	yes	yes	Transgenic mice that overexpress SREBP2 compared to wild-type mice led to statistically significant increased gene expression of 19 loci that code for enzymes involved in cholesterol synthesis in transgenic mice versus wild-type mice.	Horton et al. (2003)
Mouse (Mus musculus)	10 weeks	Transgenic mice that overexpress SREBP2.	yes	yes	Transgenic mice that overexpress SREBP2 compared to wild-type mice led to statistically increased gene expression loci that code for enzymes involved in cholesterol synthesis in transgenic mice versus wild-type mice including HMG-CoA reductase (HMGCR) and HMGCoA synthase (HMGCS).	Maxwell et al. (2003)
Human (Homo sapiens)	24 hours	Transgenic, gene knock-out in HeLa cells.	yes	yes	A combination of gene transfection for SREBP2 and gene knock-out studies in human (HeLa) cells showed statistically significant increase in SREBP2 gene expression led to statistically significant gene expression of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR), coding for a rate-limiting enzyme in cholesterol synthesis.	Howe et al. (2017)
Mouse (Mus musculus)	16 weeks	60% calories from fat (High fat diet) for 15 weeks, 50 mg/kg pregnenolone-16α-carbonitrile.	yes	yes	6 week old C57BL/6 N mice had statistically significant increase in SREBP2 gene and protein expression leading to statistically significant protein expression of hydroxymethylglutaryl-CoA reductase (HMGCR) and Liver 24-dehydrocholesterol reductase (DHCR24), enzymes involved in cholesterol synthesis, gene expression levels at these two loci and approximately 20 other loci for cholesterol synthesis enzymes increased but were not statistically analyzed.	Karpale et al. (2021)

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

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

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

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

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

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

Life Stage: All life stages.

Sex: Applies to both males and females.

Taxonomic: Primarily studied in humans and laboratory rodents.

References

List of the literature that was cited for this KER description. More help

Horton, J.D., Goldstein, J.L., and Brown, M.S. 2002. SREBPs: activators of the complete program of cholesterol and fatty acid synthesis in the liver. Journal of Clinical Investigation 109: 1125–1131.

Horton, J.D., Shah, N.A., Warrington, J.A., Anderson, N.N., Park, S.W., Brown, M.S., and Goldstein, J.L. 2003. Combined analysis of oligonucleotide microarray data from transgenic and knockout mice identifies direct SREBP target genes. Proceedings of the National Academy of Sciences 100(21): 12027–12032.

Howe, V., Sharpe, L.J., Prabhu, A.V., and Brown, A.J. 2017. New insights into cellular cholesterol acquisition: promoter analysis of human HMGCR and SQLE, two key control enzymes in cholesterol synthesis. Biochim Biophys Acta 1862: 647–657.

Itkonen, A., Hakkola, J., and Rysa, J. 2023. Adverse outcome pathway for pregnane X receptor‑induced hypercholesterolemia. Archives of Toxicology 97: 2861–2877.

Karpale, M. Karajamaki, A.J., Kummu, O., Gylling, H., Hyotylainen, T., Oresic, M., Tolonen, A., Hautajarvi, H., Savolainen, M.J., Ala-Korpela, M., Hukkanen, J., and Hakkola, J. 2021. Activation of pregnane X receptor induces atherogenic lipids and PCSK9 by a SREBP2-mediated mechanism. British Journal of Pharmacology 178: 2461–2481.

Maxwell, K.N., Soccio, R.E., Duncan, E.M., Sehayek, E., and Breslow, J.L. 2003. Novel putative SREBP and LXR target genes identified by microarray analysis in liver of cholesterol-fed mice. Journal of Lipid Research 44: 2109-2119.

Ouyang, S., Mo, Z., Sun, S., Yin, K., and Lv, Y. 2020. Emerging role of Insig-1 in lipid metabolism and lipid disorders. Clinica Chimica Acta 508: 206–212.

Sakakura, Y., Shimano, H., Sone, H., Takahashi, A., Inoue, K., Toyshima, H., Suzuki, S. and Yamada, N. 2001. Sterol regulatory element-binding proteins induce an entire pathway of cholesterol synthesis. Biochemical and Biophysical Research Communications 286: 176–183.

Yabe, D., Brown, M.S., and Goldstein, J.L. 2002. Insig-2, a second endoplasmic reticulum protein that binds SCAP and blocks export of sterol regulatory element-binding proteins. Proceedings of the National Academy of Sciences 99(20): 12753–12758.

Yang, T., Espenshade, P.J., Wright, M.E., Yabe, D., Gong, Y., Aebersold, R., Goldstein, J.L., and Brown, M.S. 2002. Crucial step in cholesterol homeostasis: sterols promote binding of SCAP to INSIG-1, a membrane protein that facilitates retention of SREBPs in ER. Cell 110: 489–500.

NOTE: Italics indicate edits from John Frisch October 2024.