Analysis of the XBP1 splicing mechanism using endoplasmic reticulum stress-indicators.
Under endoplasmic reticulum (ER) stress conditions, XBP1 mRNA is processed by unconventional splicing and translated into a functional transcription factor. ER stress-specific XBP1 splicing is also known to be activated by IRE1. However, many aspects of the molecular mechanism of XBP1 splicing remain to be elucidated. We previously developed an indicator system that enabled detection of XBP1 splicing using fluorescent proteins as the reporter signals. Here, we use a modification of this method that employs modified ER stress-indicators and mutant IRE1 in vivo and in vitro to analyze XBP1 splicing mechanisms. Our analyses suggest that the 506-579 nt region of the XBP1 mRNA is necessary and sufficient for XBP1 splicing, that XBP1 splicing can occur in the cytoplasm, and that cleavage and ligation of XBP1 mRNA during splicing may occur as a coupled reaction.
Iwawaki
Takao
T
Iwawaki Initiative Research Unit, Frontier Research System, RIKEN, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan. iwawaki@riken.jp
Akai
Ryoko
R
D002478:Cells, Cultured; D004274:DNA, Recombinant / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D007668:Kidney / Q000502:physiology*; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism*; D018384:Oxidative Stress / Q000502:physiology; D000071701:Regulatory Factor X Transcription Factors; D014157:Transcription Factors; D000071717:X-Box Binding Protein 1
D004274:DNA, Recombinant; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D000071701:Regulatory Factor X Transcription Factors; D014157:Transcription Factors; D000071717:X-Box Binding Protein 1; C000605543:XBP1 protein, human