Conformational changes in inhibitory PAS domain protein associated with binding of HIF-1α and Bcl-xL in living cells.

Abstract:

:Inhibitory PAS domain protein (IPAS) is a dual function protein acting as a transcriptional repressor and as a pro-apoptotic protein. Simultaneous dual-color single-molecule imaging of EGFP-IPAS coexpressed with Mit-TagRFP-T in living HeLa cells revealed that fraction of EGFP-IPAS was arrested in the nucleus and on mitochondria. Transiently expressed Cerulean-IPAS in HEK293T cells was present in nuclear speckles when coexpressed with Citrine-HIF-1α or Citrine-HLF. Fluorescence lifetime imaging microscopy (FLIM) analysis of Citrine-IPAS-Cerulean in living CHO-K1 cells clarified the presence of intramolecular FRET. Reduced lifetimes of the donor were partially restored by coexpression of HIF-1α or Bcl-xL, binding proteins of IPAS in the nucleus and mitochondria, respectively. This alteration in lifetimes demonstrates that conformational changes occurred in IPAS by their binding.

journal_name

J Biochem

journal_title

Journal of biochemistry

authors

Kasai S,Kajimoto S,Ito Y,Saito T,Yasumoto KI,Tokunaga M,Sakata-Sogawa K,Fukumura H,Sogawa K

doi

10.1093/jb/mvw068

subject

Has Abstract

pub_date

2017-03-01 00:00:00

pages

291-296

issue

3

eissn

0021-924X

issn

1756-2651

pii

mvw068

journal_volume

161

pub_type

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