Exploring the Role of Endoplasmic Reticulum Calcium Dynamics in Maintaining Cholesterol Homeostasis

• Author / Creator

  Wang, Wen-An Jennifer

• Calreticulin is an endoplasmic reticulum (ER) protein chaperone and calcium (Ca2+) binding protein and is therefore important for maintaining ER homeostasis. Calreticulin deficiency is embryonic lethal in mice due to inadequate inositol trisphosphate receptor (InsP3R)-mediated Ca2+ signaling and Ca2+/calcineurin-dependent activation of transcription factors involved in cardiogenesis. Cardiac-specific expression of constitutively active calcineurin allows for normal cardiac development of calreticulin-deficient mice and rescues embryonic lethality in mice. However, the rescued Calr-/- mice have disrupted energy metabolism, with a high concentration of triacylglycerols and cholesterol in the blood plasma indicating the existence of a link between lipid metabolism and calreticulin.
  The objective of the research within this thesis was to explore the consequence of disrupted ER homeostasis in maintaining cholesterol homeostasis. Specifically, we investigated the consequence of the absence of calreticulin on cholesterol metabolism controlled through the sterol regulatory binding protein (SREBP) pathway. We discovered that in the absence of calreticulin, there was increased accumulation of lipids in Calr-/- cells and Caenorhabditis elegans and this increase was attributed to the increased activation of SREBP and de novo lipid biosynthesis. This increase in SREBP activity and lipid biosynthesis in the absence of calreticulin occur in the absence of cholesterol depletion and intracellular levels of unesterified cholesterol were abundant.
  To unravel this conundrum, we investigated the effect of the chaperone activity of calreticulin on components of the SREBP processing pathway and found that the SREBP processing pathway was fully functional in the absence of calreticulin. Next, we investigated the Ca2+ binding
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  function of calreticulin and identified that in the absence of calreticulin Ca2+ binding, the reduction of ER Ca2+ stores caused a re-distribution of the intracellular unesterified cholesterol away from the sensing mechanism of the SREBP processing pathway. Therefore we were able to establish a link between ER Ca2+ and cholesterol homeostasis.
  Having established the connection between of ER Ca2+ and cholesterol homeostasis, we next investigated the mechanism behind the role of ER Ca2+ in the distribution of intracellular cholesterol. ER Ca2+ dynamics is a tightly regulated event that involves the level of ER Ca2+ stores, ER Ca2+ release and ER Ca2+ refilling regulated by a process known as store-operated Ca2+ entry (SOCE). Therefore, we investigated the role of SOCE as a mechanism behind the observed redistribution of unesterified cholesterol and increased SREBP activity in the absence of calreticulin and reduced ER Ca2+ conditions. We discovered that the stromal interaction molecule 1 (STIM1), an integral ER membrane protein that senses ER Ca2+ levels and an important component of SOCE, and its recently discovered cholesterol binding domain plays an important role in intracellular unesterified cholesterol distribution and SREBP activity. We hypothesize that STIM1 may function in binding ER membrane unesterified cholesterol and play a role in the movement of cholesterol between the ER and plasma membrane. Furthermore, this movement and distribution of unesterified cholesterol may be affected by ER Ca2+ status and SOCE activity.

• Subjects / Keywords

  • Endoplasmic Reticulum
  • cholesterol
  • calreticulin
  • calcium
  • sterol regulatory binding protein
  • stromal interaction molecule 1

• Graduation date

  Fall 2018

• Type of Item

  Thesis

• Degree

  Doctor of Philosophy

• DOI

  https://doi.org/10.7939/R3JM23X9G

• License

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