QUANTITATIVE MEASUREMENT OF SPLICED XBP1 mRNA AS AN INDICATOR OF ENDOPLASMIC RETICULUM STRESS

Abstract

The unfolded protein response (UPR) events triggered by the accumulation of unfolded protein in endoplasmic reticulum (ER) activate the three UPR signaling pathways mediated by IRE1, ATF6 and PERK. Spliced XBP1 mRNA induced by activated IRE1 is translated to the protein, a potent transcription factor that induces BiP expression. XBP1 is also induced by activated ATF6. It is thus thought to be an important marker reflecting both IRE1 and ATF6 signaling in response to ER stress. For quantitative measurement of XBP1 gene expression, it is important to distinguish between the spliced and non-spliced form of XBP1 mRNA. We have developed a new method to detect the spliced XBP1 mRNA by means of real-time PCR and we compared the result with measurements of the expression of the ER stress inducible gene BiP. A good correlation was found between spliced XBP1 expression and BiP expression. Thus, our method may be useful for simple and quantitative evaluation of ER stress.

Palmitate modulates intracellular signaling, induces endoplasmic reticulum stress, and causes apoptosis in mouse 3T3-L1 and rat primary preadipocytes

“…6B). Figure 6C shows that treatment with palmitate alone or mixed with oleate/linoleate induced partial but significant XBP-1 mRNA splicing, further evidence of increased ER stress (22). As a positive control, we showed that thapsigargin, a chemical inducer for ER stress, induced a greater increase of CHOP and GRP78 mRNA (Fig.…”

Activation of the Unfolded Protein Response Is Associated with Favorable Prognosis in Acute Myeloid Leukemia

“…6). Activation of IRE1 induces the cleavage of the XBP1 mRNA, generating a spliced mRNA, which encodes a potent novel transcriptional activator of downstream unfolded protein response target genes (7)(8)(9)(10)13). This spliced mRNA can rapidly be detected by semiquantitative PCR, allowing the identification of activated unfolded protein response.…”

Antitumor alkyl-lysophospholipid analog edelfosine induces apoptosis in pancreatic cancer by targeting endoplasmic reticulum

“…The presence in this 26-nucleotide fragment of a PstI restriction site further allowed us to distinguish between both XBP1 forms by restriction analysis of PCR-amplified complementary DNA, and thus to assay for ER stress response activation (Hirota et al, 2006). By reverse transcriptase-PCR we found the induction of sXBP1 mRNA following treatment of BxPC-3 cells with 20 mM edelfosine ( Figure 2f).…”