Riboproteomic Approaches to Understanding IRES Elements - Biophysical Approaches to Translational Control of Gene Expression - page 107

as already shown for the Hu antigen R (HuR) protein that represses the activity of the
HIV-1 IRES while it stimulates that of HCV (Rivas-Aravena et al. 2009 ) . HIV-1
IRES-mediated translation is also stimulated by hnRNP A1/2 or the RNA helicase
DDX3, hRIP, and Sam68 (Liu et al. 2011 ) .

6.4 ITAFs Interacting with Cellular IRES

Cellular IRES are typically present in mRNAs encoding stress response proteins;
hence, they have evolved mechanism to evade global repression of translation
(Holcik and Korneluk 2000 ; Spriggs et al. 2005 ). A characteristic of cellular IRES
elements is to depend on its interaction with ITAFs to facilitate the binding of the
mRNA to 40S subunits. In this way, changes in the abundance, posttranslational
modifications, or subcellular location of ITAFs contribute to control IRES-mediated
translation during stress conditions, including nutrient stress, temperature shock,
hypoxia, cell cycle arrest, or apoptosis (Conte et al. 2008 ; Komar and Hatzoglou
2011 ; Lewis and Holcik 2008 ; Spriggs et al. 2010 ) .
Conventional biochemical approaches carried out to identify ITAFs interacting
with cellular IRES, in conjunction with functional characterization, have provided a
long list of factors (Table 6.2 ), basically consisting in abundant RNA-binding pro-
teins that shuttle between the nucleus and the cytoplasm. Identification of ITAFs
relied basically on the characterization of ribonucleoprotein complexes assembled
with the IRES of interest (Fox and Stover 2009 ; Graber et al. 2010 ; Lewis et al.
2008 ; Majumder et al. 2009 ; Spriggs et al. 2009 ) .
The identification of factors controlling the expression of apoptotic proteins as
well as the myc family of transcription factors has been cumbersome in gathering
information of proteins mediating IRES-dependent translation (Bushell et al. 2006 ;
Cobbold et al. 2008, 2010 ; Henis-Korenblit et al. 2002 ; Mitchell et al. 2001 ) . Many
of these factors have been also identified associated to other cellular IRES (Table 6.2 ).
This is the case of PTB (hnRNP I) that stimulates translation driven by IRES of
mRNAs encoding proteins required for cell survival under apoptosis, hypoxia,
nutrient deprivation, or cell growth dysregulation (the apoptotic protease activating
factor 1 (apaf-1), BCL2-associated athanogene (BAG)-1, p53, hypoxia-inducible
factor (HIF1a), among others) (Dobbyn et al. 2008 ; Grover et al. 2008 ; Schepens
et al. 2005 ), although it represses translation initiation driven by the glucose-regu-
lated protein GRP78 immunoglobulin heavy chain-binding protein (BiP) IRES
(Kim et al. 2000b ) .
Recently, a proteomic approach devoted to isolate RNA-protein complexes from
living cells has been applied to obtain quantitative profiles of proteins interacting
with the lymphoid enhancer factor (LEF-1) IRES (Tsai et al. 2011 ) . This approach
is based on the use of biotin-tagged transfected RNAs expressing the bacteriophage
protein MS2, which allows affinity purification of UV-cross-linked ribonucleopro-
tein complexes assembled in living cells, combined with stable isotope labeling
with amino acids in cell culture (SILAC)-based quantitative mass spectrometry.