SLIP1, a Factor Required for Activation of Histone mRNA Translation by the Stem-Loop Binding Protein

Abstract

Replication-dependent histone mRNAs are the only eukaryotic cellular mRNAs that are not polyadenylated, ending instead in a conserved stem-loop. The 3 end of histone mRNA is required for histone mRNA translation, as is the stem-loop binding protein (SLBP), which binds the 3 end of histone mRNA. We have identified five conserved residues in a 15-amino-acid region in the amino-terminal portion of SLBP, each of which is required for translation. Using a yeast two-hybrid screen, we identified a novel protein, SLBP-interacting protein 1 (SLIP1), that specifically interacts with this region. Mutations in any of the residues required for translation reduces SLIP1 binding to SLBP. The expression of SLIP1 in Xenopus oocytes together with human SLBP stimulates translation of a reporter mRNA ending in the stem-loop but not a reporter with a poly(A) tail. The expression of SLIP1 in HeLa cells also stimulates the expression of a green fluorescent protein reporter mRNA ending in a stem-loop. RNA interference-mediated downregulation of endogenous SLIP1 reduces the rate of translation of endogenous histone mRNA and also reduces cell viability. SLIP1 may function by bridging the 3 end of the histone mRNA with the 5 end of the mRNA, similar to the mechanism of translation of polyadenylated mRNAs.

INT6 interacts with MIF4GD/SLIP1 and is necessary for efficient histone mRNA translation

“…It has been reported previously that MIF4GD interacts with SLBP and is required for efficient translation of histone mRNAs (Cakmakci et al 2008). Because INT6 is associated with the eIF3 translation initiation complex and binds MIF4GD, we next examined whether INT6 is also important for histone mRNA translation.…”

“…Besides its role in formation of the 39 end, SLBP is also important for export of the histone mRNA (Ghule et al 2008;Sullivan et al 2009a), as well as for their productive translation, and this translational activity relies on interactions with eIF4G and eIF3 translation initiation factors (Ling et al 2002;Gorgoni et al 2005). More recently, it has also been reported that this effect involves an interaction with a protein identified in a two-hybrid screen performed with SLBP as bait, which was accordingly named SLBP interacting protein 1 (SLIP1) (Cakmakci et al 2008). This small 25-kDa protein corresponds almost entirely to a middle domain of eukaryotic initiation factor 4G (MIF4G) and has been designated accordingly as MIF4G domain-containing protein (MIF4GD) by the HUGO Gene Nomenclature Committee, which name is further used in this article.…”

“…The bands corresponding to histones H3, H2B, and H4 are indicated. (C) The experiment was repeated three times, and the amount of radioactivity in the H3/H2B bands as well as in the H4 band was quantified using the MultiGauge software with normalization with respect to bands of the upper part of the gels as previously described (Cakmakci et al 2008). The mean of the ratios with respect to cells transfected with control siRNAs is represented with an error bar corresponding to the standard deviation.…”

“…lane 1 with lanes 2-5). This experiment was repeated three times, and the radioactivity in the histone signals was normalized with bands of the upper part of the gel (Cakmakci et al 2008). Because they were poorly separated, the bands corresponding to histones H2B and H3 were grouped for quantification.…”

The C-terminal extension of Lsm4 interacts directly with the 3′ end of the histone mRNP and is required for efficient histone mRNA degradation

“…As Lsm4 showed the most robust binding, we focused only on it for this study. We also tested the ability of the seven Lsm proteins to interact with GST-SLIP1, a component of the cytoplasmic histone mRNP that is involved in translation (Cakmakci et al 2008). None of the subunits of the Lsm1-7 complex interacted directly with SLIP1 (Supplemental Data).…”