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UBC Theses and Dissertations

Implicating calcium signaling at intercellular junctions and structures associated with junction turnover in rat Sertoli cells Lyon, Kevin


The endoplasmic reticulum (ER) is a prominent organelle in Sertoli cells. It is an integral component of unique adhesion junctions (ectoplasmic specializations - ESs) in this cell type, and is closely associated with structures termed tubulobulbar complexes (TBCs) that internalize intercellular junctions during sperm release and during the translocation of spermatocytes through the blood-testis barrier. A role for the ER in Ca²⁺ regulation at ESs and TBCs has been suspected, but evidence for this function has proved elusive. The focus of this thesis is identification of molecular machinery involved in Ca²⁺ signaling and obtaining functional evidence of Ca²⁺ regulation of the actin networks at TBCs. Functional experiments using EGTA and thapsigargin to lower and raise Ca²⁺ levels did not provide evidence that TBC actin networks are regulated by Ca²⁺. Using immunofluorescence, I demonstrated that Ca²⁺ regulatory machinery is present at the ESs attached to spermatid heads, and at ER-PM contacts. SERCA2 is present at ESs, IP3R is present at ER-PM contacts associated with TBC bulbs, and STIM1, ORAI1 and SERCA2 are present at the ER-PM contacts around the margins of Sertoli cell apical processes. The results support the conclusion that the molecular machinery necessary for ER generated Ca²⁺ fluxes is present in regions and structures directly related to junction remodeling in Sertoli cells, a process necessary for sperm release.

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