Deciphering the role of STAP1 in cholesterol metabolism

knockout mouse functional genetics (medical)genetics

Prof. dr. T.P.W. Kamphuisen
dr. J.A. Kuivenhoven
J. Balder

Nature of the research:
Animal research, fundamental research

Fields of study:
biomaterials genetics vascular medicine

Background / introduction
Familial hypercholesterolemia (FH) is a common monogenic disease which presents with increased levels of plasma cholesterol1,2. Increased levels of cholesterol lead to accelerated development of atherosclerosis. In 60% of the individuals with FH, mutations are found in low-density lipoprotein receptor (LDLR), apolipoprotein B (APOB) and proprotein convertase subtilisin/kexin type 9 (PCSK9). Recently a fourth candidate gene for FH was found in families, namely signal transducing adaptor family member 1 (STAP1)3,4. So far, it is not known how STAP1 interferes with the cholesterol metabolism.
Last year we used a new technology to insert a mutation in STAP1 in mice, creating a knock out (KO) mouse model. The expression of STAP1 in the mouse model is extremely decreased compared to the mice without this mutation (wild type, WT). We compared 32 mice with a mutation in STAP1 to 32 mice without this mutation. At the age of 12 weeks, there was no difference in cholesterol between the mice. It is known that mice have many compensatory mechanisms for the loss of proteins, in such cases the phenotype can be triggered by putting pressure on the cholesterol homeostasis. Therefore, we fed the mice a high fat + high cholesterol diet for four weeks. After the diet intervention no differences in cholesterol were found. However, the weight of the KO mice is lower compared to WT mice. So we know that something is going on.
To further provoke a phenotype, we are planning to inject the mice with a viral vector that will drive the expression of a gain of function mutation in PCSK9 which will increase cholesterol levels.
The student can help us in designing the experiment, and perform practical work.
Research question / problem definition
1. Is STAP1 an important regulator of cholesterol in mice?
1. Inject mice 16 wild-type and 16 control mice with AAV-based vector
2. Feed high sucrose diet and monitor food intake and body weight
3. Measure cholesterol levels in blood obtained by orbital puncture after 0, 2, 4 and 6 weeks
4. Harvest, store and analyze tissues (liver, spleen, kidney, thymus, and lymph nodes)
a. Transcriptome analyses, quantitative PCR and proteomics of genes/proteins involved in cholesterol metabolism
1. Benn M, Watts GF, Tybjaerg-Hansen A, Nordestgaard BG. Familial hypercholesterolemia in the danish general population: Prevalence, coronary artery disease, and cholesterol-lowering medication. J Clin Endocrinol Metab. 2012;97(11):3956-3964.
2. Nordestgaard BG, Chapman MJ, Humphries SE, et al. Familial hypercholesterolaemia is underdiagnosed and undertreated in the general population: Guidance for clinicians to prevent coronary heart disease: Consensus statement of the european atherosclerosis society. Eur Heart J. 2013;34(45):3478-90a.
3. Fouchier SW, Dallinga-Thie GM, Meijers JC, et al. Mutations in STAP1 are associated with autosomal dominant hypercholesterolemia. Circ Res. 2014;115(6):552-555.
4. Braenne I, Kleinecke M, Reiz B, et al. Systematic analysis of variants related to familial hypercholesterolemia in families with premature myocardial infarction. Eur J Hum Genet. 2016;24(2):191-197.
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