Proline-Rich Transcript in Brain Protein Induces Stress Granule Formation

Abstract

The repression of translation in environmentally stressed eukaryotic cells causes the sequestration of translation initiation factors and the 40S ribosomal subunit into discrete cytoplasmic foci called stress granules (SGs). Most components of the preinitiation complex, such as eIF3, eIF4A, eIF4E, eIF4G, and poly(A)-binding protein, congregate into SGs under stress conditions. However, the molecular basis of translation factor sequestration into SGs has not been clearly elucidated. Here, we report that proline-rich transcript in brain (PRTB) protein interacts with eIF4G and participates in SG formation. PRTB was recruited to SG under sodium arsenite and heat stress conditions. When overexpressed, PRTB inhibited global translation and formed SGs containing TIA-1, eIF4G, and eIF3. Knockdown of PRTB reduced the SG formation induced by sodium arsenite. These results suggest that PRTB not only is a component of SG formed by cellular stresses but also plays an important role in SG formation via an interaction with the scaffold protein eIF4G, which is associated with many translation factors and mRNAs.The translation rate of eukaryotic mRNAs is modulated by environmental conditions, with that of many mRNAs being inhibited when cells are stressed by heat, high osmolarity, oxidative chemicals, etc. (15). The translation inhibition causes the sequestration of translation initiation factors and the 40S ribosomal subunit into discrete cytoplasmic foci, called stress granules (SGs), that are formed in environmentally stressed eukaryotic cells (1, 2, 14-17, 19, 26, 30). The SGs contain most of the components of the 48S preinitiation complex (i.e., small [but not large] ribosomal subunits, eukaryotic initiation factor 4G [eIF4G], eIF3, eIF4E, eIF2, and eIF2B), other RNA-binding proteins, such as T-cell-restricted intracellular antigen-1 (TIA-1) and T-cell-restricted intracellular antigen-related protein (TIAR), and untranslated mRNAs (1, 2, 15-17). As a consequence, mRNA translation generally is inhibited under stress conditions (6, 31).eIF4G plays a pivotal role in the initiation of translation, since it recruits many translation factors [poly(A)-binding protein (PABP) (12), eIF4E (20, 23), eIF4A (13), and eIF3 (13)] and the translation modulator Mnk1 (a Ser/Thr kinase) to the 40S ribosomal subunit via protein-protein interactions (28, 37). Moreover, the signaling molecule TRAF2 has been shown to bind to eIF4GI, one of the two functional homologues of eIF4G, and to block proinflammatory signaling via the sequestration of TRAF2 at the SGs under stress conditions (18,25). This indicates that eIF4G plays important roles in the regulation of cellular activities such as translation and signal transduction.The proline-rich transcript of the brain (PRTB) protein, which is a 17-kDa protein, originally was isolated in a gene trap screen as a transcript expressed in the developing mouse inner ear (38). PRTB also is known as DAZAP2 [deleted-inazoospermia (DAZ)-associated protein 2], which was identified as a protein interacting with the...

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“…DAZAP2 was originally identified as a highly conserved, small, proline-rich protein which interacts with the deleted in azoospermia (DAZ) and the DAZ-like (DAZL) proteins through the DAZ-like repeats (36). Subsequently, this protein was also found to interact with the transforming growth factor-beta signaling molecule SARA (Smad anchor for receptor activation), eukaryotic initiation factor 4G, and an E3 ubiquitinase that regulates its stability in splicing factor containing nuclear speckles (14,15,42). Thus, DAZAP2 may function in various biological and pathological processes, including spermatogenesis, cell signaling and transcription regulation, formation of stress granules during translation arrest, RNA splicing, and the pathogenesis of multiple myeloma (22).…”

DAZL is essential for stress granule formation implicated in germ cell survival upon heat stress

“…2A). Another SG component, DAZAP2, is known to interact with eIF4G and to induce SG formation (Kim et al, 2008). In normal testis, DAZAP2 was detected in the nucleus and cytoplasm of male germ cells of all developmental stages except for elongated spermatids ( Fig.…”

“…The coding region of the human DAZAP2 cDNAs (Kim et al, 2008) was subcloned into pGEX-4T vector (GE Healthcare) for the generation of GST-DAZAP2 fusion protein. A rabbit polyclonal anti-DAZAP2 antibody was raised against bacterially expressed GST-DAZAP2 and affinity-purified using a method described elsewhere (Kim et al, 2009).…”

“…It is also responsible for the specific interaction with PABP, a major SG component, suggesting that DAZL is recruited to SGs through specific interaction with PABP (Kedersha et al, 1999;Collier et al, 2005). It is also noteworthy that DAZL interacts with DAZAP2, an SG component and an interacting partner of the translation initiation factor eIF4G (Tsui et al, 2000;Kim et al, 2008). At the same time, DAZL might function as a nucleator of SG assembly, as ectopic expression of DAZL induces SG formation in HeLa cells (Lee et al, 2006).…”

RhoB induces apoptosis via direct interaction with TNFAIP1 in HeLa cells

“…19,20 RhoB bait cloned into EcoR1/BamHI sites of the pGBKT vector and cDNA libraries of human brain and HeLa cells were used for screening purposes. The yeast PBN204 strain that contains 3 reporters (Ura3, LacZ and Ade2) under the control of different GAL promoters was used for transformation.…”