Binding Protein Calreticulin Research Articles

Calcium uptake during mitogenic stimulation of human lymphocytes: characterization of intracellular calcium compartments and demonstration of the presence of immunoreactive calrecticulin.

Phytohemagglutinin (PHA) stimulation of human peripheral blood lymphocytes (HPBL) rapidly increases 45Ca2+ uptake into intracellular pools. Detectable increase in 45Ca2+ uptake occurred only on exposure to mitogenic lectins but not with non-mitogenic lectins. However, intracellular free Ca2+ concentration [(Ca2+)i] increased comparably on exposure to either mitogenic or non-mitogenic lectins. Permeabilization of 45Ca2+ loaded cells revealed distinct pools of Ca2+ uptake. The highly digitonin sensitive pool #I (permeabilized by 0.02% digitonin) exchanged slowly and included a part that represented endoplasmic reticulum. Pool II was defined by lower digitonin sensitivity, had a much faster initial uptake. Pool III was digitonin-resistant and predominantly non-vesicular. During the first 120 min of PHA stimulation, significant increase in 45Ca2+ uptake occurred only into pool II. Progressive increase in uptake into pool I then occurred so that by 24 hours, this pool constituted the major fraction of PHA induced increment in total 45Ca2+ uptake. Using specific antibody to the calcium binding protein calreticulin, an analogous immunoreactive protein was detectable in resting HPBL. PHA stimulation led to a striking increase in abundance of immunoreactive calreticulin so that 24 hrs after PHA stimulation, there was a 28 and 3.4 fold increase in the amount of immunoreactive calreticulin present in the non-particulate fraction and the total particulate membrane fraction, respectively. A major part (72%) of the total cellular immunoreactive calreticulin in PHA stimulated cells at 24 hrs was released into the medium after permeabilization of lymphocytes with 0.02% digitonin, corresponding to the location of calcium uptake pool I.