Colocalization of sterol isomerase and sigma(1) receptor at endoplasmic reticulum and nuclear envelope level.
SR31747A is a sigma ligand previously described as having original immunosuppressive properties. Two SR31747A targets were recently identified and termed sigma(1) or SR-BP-1 (SR31747A-binding protein-1) and hSI (human sterol isomerase). In order to characterize these proteins further, we examined their expression and localization at the subcellular level. Based on the amino acid sequence deduced from the cloned hSI, anti-hSI polyclonal antibody was raised against the N-terminal fragment of the protein. Using this antibody, we performed Western-blot experiments to demonstrate the presence of hSI in various B and T cell lines, and hSI expression was quantified in these cell lines by flow cytometry and estimated at 15 000-30 000 sites per cell. Subcellular localization studies by both confocal and electron microscopy, performed on THP1 cells with anti-hSI antibody and with the previously described anti-(SR-BP-1) monoclonal antibody, demonstrated that: (a) hSI was colocalized with SR-BP-1; (b) hSI and SR-BP-1 were associated with the endoplasmic reticulum and with the outer and inner membranes of the nuclear envelope; (c) both proteins were delocalized during the cell cycle at the mitosis step when the nuclear membranes disappeared. Taken together our results suggest that both SR31747A-binding proteins not only play a role in sterol metabolism but indirectly affect lipoprotein functions.
Sanofi Recherche, Montpellier, France Sanofi Recherche, Labège, France.
D
D
Dussossoy
P
P
Carayon
S
S
Belugou
D
D
Feraut
A
A
Bord
C
C
Goubet
C
C
Roque
H
H
Vidal
T
T
Combes
G
G
Loison
P
P
Casellas
D000911:Antibodies, Monoclonal; D002352:Carrier Proteins / Q000276:immunology* / Q000378:metabolism*; D002460:Cell Line; D004306:Dose-Response Relationship, Immunologic; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D005434:Flow Cytometry; D018922:HL-60 Cells; D006801:Humans; D007150:Immunohistochemistry; D019169:Jurkat Cells; D018613:Microscopy, Confocal; D008954:Models, Biological; D009685:Nuclear Envelope / Q000378:metabolism*; D011957:Receptors, Opioid; D017480:Receptors, sigma; D013251:Steroid Isomerases; D020298:U937 Cells; D000097605:Sigma-1 Receptor
D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D011957:Receptors, Opioid; D017480:Receptors, sigma; D013251:Steroid Isomerases; D000097605:Sigma-1 Receptor; C090445:EBP protein, human