AZM activates UPR activity through facilitating the dissociation
of GRP78 from its interactors
The activation of ER stress occurred upon the dissociation of GRP78 from
the luminal part of the ER integral membrane proteins: PERK, IRE1α and
ATF6α (Yoo et al., 2013). Co-immunoprecipitation experiments showed that
incubation of RA FLSs lysates with AZM decreased the GRP78 level in the
PERK, IRE1α, and ATF6α immunoprecipitated fractions (Fig. 5A), thus
indicating that AZM induced a dissociation of the GRP78, PERK, IRE1α and
ATF6α complex and representing the initial event necessary to induce the
observed ER stress.
Furthermore, SREBP-SCAP complex is like other key integral membrane
proteins of the UPR associated with GRP78, and, the export of the
complex requires its dissociation from GRP78 (Jin et al., 2000). GRP78
dissociates from SREBP allows its cleavage and nuclear translocation for
its regulatory function (Katanasaka et al., 2010). Furthermore,
attenuation of ER stress by overexpression of GRP78 blocked SREBP
activation and decreased the expression of genes responsible for
cholesterol and fatty acid biosynthesis (Kammoun et al., 2009). Here, an
enhanced expression of m-SREBP-1c by AZM was seen in nuclear extract of
RA FLSs, but not in cytoplasmic protein (Fig. 5B). Furthermore, the
enhancement of m-SREBP-1c (mature SREBP-1c) nuclear translocation
resulting from AZM treatment could be attenuated by overexpression of
GRP78 in RA FLSs (Fig. 5B). These data supported that AZM inhibits the
activity of GRP78, thereby provoking ER stress and activating
SREBP-mediated cholesterol and lipid metabolism.