BiP and Calreticulin Form an Abundant Complex That Is Independent of Endoplasmic Reticulum Stress

Abstract

BiP is found in association with calreticulin, both in the presence and absence of endoplasmic reticulum stress. Although the BiP-calreticulin complex can be disrupted by ATP, several properties suggest that the calreticulin associated with BiP is neither unfolded nor partially or improperly folded. (1) The complex is stable in vivo and does not dissociate during 8 hr of chase. (2) When present in the complex, calreticulin masks epitopes at the C terminus of BiP that are not masked when BiP is bound to an assembly-defective protein. And (3) overproduction of calreticulin does not lead to the recruitment of more BiP into complexes with calreticulin. The BiP-calreticulin complex can be disrupted by low pH but not by divalent cation chelators. When the endoplasmic reticulum retention signal of BiP is removed, complex formation with calreticulin still occurs, and this explains the poor secretion of the truncated molecule. Gel filtration experiments showed that BiP and calreticulin are present in distinct high molecular weight complexes in which both molecules interact with each other. The possible functions of this complex are discussed. INTRODUCTIONThe endoplasmic reticulum (ER) is the primary organelle of the endomembrane system and is responsible for the biosynthesis and maturation of proteins that are destined for secretion, for the plasma membrane, and for transport to various organelles of the endocytic and exocytic pathways (reviewed in Palade, 1975;Helenius et al., 1992; reviewed for plants in Vitale et al., 1993). In addition to nascent polypeptides and newly synthesized proteins, the ER lumen also contains a large and abundant family of soluble resident proteins known as the reticuloplasmins (Koch, 1987; reviewed for plants in Denecke, 1996). Several reticuloplasmins are now recognized as molecular chaperones (Gething and Sambrook, 1992;Hartl, 1996), which prevent the aggregation of unfolded and partially folded polypeptides, thus increasing the yield but not the rate of correct folding and assembly. Knowledge of the binding specificity of chaperones is rapidly expanding; however, mechanistic details of how the binding of the chaperone results in the unfolding of the protein followed by refolding and release from the chaperone remain to be established.Several reticuloplasmins have been identified in plants, including the binding protein BiP (Denecke et al., 1991;Fontes et al., 1991;Anderson et al., 1994), calreticulin (Chen et al., 1994;Dresselhaus et al., 1996), protein disulfide isomerase (PDI; Shorrosh and Dixon, 1991), and endoplasmin (Walther-Larsen et al., 1993;Denecke et al., 1993. Endoplasmin has homology with the cytosolic heat shock protein Hsp90 family, but no definitive function has been proposed. PDI catalyzes the formation of disulfide bonds, a process restricted to the ER lumen, hence the absence of cytosolic homologs of PDI.The best-characterized ER resident protein is BiP (Munro and Pelham, 1986). BiP belongs to the Hsp70 family of molecular chaperones. This family has members in every c...

Aberrant localization and underglycosylation of highly accumulating single-chain Fv-Fc antibodies in transgenicArabidopsisseeds

“…The presence of calreticulin and BiP in the PS, as well as cruciferin that normally accumulates in the PSV (22), suggests that the overproduction of the recombinant scFv-Fc also disturbs the retention and sorting mechanisms of endogenous proteins. Given their strong biological and physical association (23), the colocalization of calreticulin and BiP in the PS is not surprising. Under normal circumstances, these ER-resident proteins are not found outside the plasma membrane (24).…”

The Secreted Plant N-Glycoproteome and Associated Secretory Pathways

“…After removal of three Glc residues the N -glycoproteins enter the calnexin-calreticulin cycle (CNX/CRT; Hebert et al, 1995). The alternate action of glucosidase II and UDP-glucose:glycoprotein glucosyltransferase drives the glycoprotein through this cycle until it is correctly folded and exported from the ER to the GA. Misfolded proteins are directed from the ER to the cytosol by the ER-associated degradation (ERAD) machinery for proteasomal hydrolysis (Hebert et al, 1995; Crofts et al, 1998; Helenius and Aebi, 2004; Jin et al, 2007; Lederkremer, 2009). Glucosyl transferase and glucosidases implicated the in the sugar trimming in the ER and identified in plants so far, are enlisted in Table 1.…”

Over-expression of calcium-dependent protein kinase 13 and calreticulin interacting protein 1 confers cold tolerance on rice plants

“…Although such calcium signatures may partially explain the speciWcity of cellular responses triggered by a particular stimulus, the molecules that "sense" and "interpret" the calcium signals provide additional speciWcity to the coupling of calcium parameters to cellular responses (McAinsh et al 1992). Several families of calcium sensors have been identiWed in higher plants and are broadly divided into four major classes: calmodulin, calmodulin-like and other EF hand-containing calcium-binding proteins, calcium-regulated protein kinase and calciumbinding proteins without an EF-hand motif such as calreticulin (Zielinski 1998;Harmon et al 2000;Crofts and Denecke 1998;Llewelyn et al 1998). …”