Thapsigargin inhibits the sarcoplasmic or endoplasmic reticulum Ca-ATPase family of calcium pumps.

TLDR

ATP‑dependent calcium uptake into intracellular stores is essential for hormonally induced signaling, and thapsigargin is used to manipulate calcium levels by inhibiting the endoplasmic reticulum calcium pump. The study directly tested thapsigargin’s specificity against all known intracellular calcium pumps of the SERCA family. Full‑length cDNA clones of SERCA1, SERCA2a, SERCA2b, and SERCA3 were expressed in COS cells, and calcium uptake and ATPase activity were measured in isolated microsomes. Thapsigargin inhibited all SERCA isozymes with equal potency, rapidly and irreversibly abolishing calcium uptake and ATPase activity in sarcoplasmic reticulum from fast‑twitch and cardiac muscle while sparing plasma membrane Ca‑ATPase and Na,K‑ATPase, indicating a unique recognition site shared by all ER/SR calcium pumps.

Abstract

The role of ATP-dependent calcium uptake into intracellular storage compartments is an essential feature of hormonally induced calcium signaling. Thapsigargin, a non-phorboid tumor promoter, increasingly is being used to manipulate calcium stores because it induces a hormone-like elevation of cytosolic calcium. It has been suggested that thapsigargin acts through inhibition of the endoplasmic reticulum calcium pump. We have directly tested the specificity of thapsigargin on all of the known intracellular-type calcium pumps (referred to as the sarcoplasmic or endoplasmic reticulum Ca-ATPase family (SERCA]. Full-length cDNA clones encoding SERCA1, SERCA2a, SERCA2b, and SERCA3 enzymes were expressed in COS cells, and both calcium uptake and calcium-dependent ATPase activity were assayed in microsomes isolated from them. Thapsigargin inhibited all of the SERCA isozymes with equal potency. Furthermore, similar doses of thapsigargin abolished the calcium uptake and ATPase activity of sarcoplasmic reticulum isolated from fast twitch and cardiac muscle but had no influence on either the plasma membrane Ca-ATPase or Na,K-ATPase. The interaction of thapsigargin with the SERCA isoforms is rapid, stoichiometric, and essentially irreversible. These properties demonstrate that thapsigargin interacts with a recognition site found in, and only in, all members of the endoplasmic and sarcoplasmic reticulum calcium pump family.