A 69-kDa RNA-binding protein from Xenopus oocytes recognizes a common motif in two vegetally localized maternal mRNAs.

Abstract

Vgl mRNA, a maternal message encding a member of the transforming growth factor (3 superfamily, undergoes localization to the vegetal cortex of Xenopus laevis oocytes during a narrow period of oogenesis. A 340-nudweotde sequence has been identified in Vgl RNA that directs its vegetal localization [Mowry, K. L. & Melton, D. A. (1992) Science 255, [991][992][993][994]. To understand how cis-and trans-acting factors are involved in Vgl mRNA localization, we have looked for specific interactions in vitro between oocyte proteins and Vgl mRNA. S100 extracts of late-stage oocytes contain a proteinbinding activity that protects specific regions of labeled Vgl mRNA from degradation by RNase T1. The use of different regions of Vgl RNA in competition reactions reveals two binding sites, both in the first half of the 3' untranslated region of Vgl message. UV crosslinking predominantly labels a 69-kDa protein; saturation analysis and competitor studies indicate that this protein binds with a high affinity to the downstream site, which corresponds to the 340-nucleotide vegetal localization sequence. Binding to this region is inhibited by another vegetally localized message, transforming growth factor (35 but is not inhibited by an animally localized RNA, An2. These data indicate that vegetally localized mRNAs share a binding motif that helps them achieve their intracellular distribution through specific RNA-protein interactions.The polarity of Xenopus oocytes is evident from very early stages and is perhaps presaged by an asymmetry detectable in primordial germ cells (1, 2). As oogenesis proceeds, this polarity is elaborated by a series of morphological events that occur along the animal-vegetal axis, such as asymmetric distribution of yolk platelets, migration of pigment granules to the animal hemisphere, accumulation of polar granules at the vegetal pole, and the displacement ofthe germinal vesicle to the animal hemisphere (3,4). On a molecular level, this organization is reflected in a small class of RNA molecules that are localized to particular regions of the oocyte cytoplasm (5, 6). The heterogeneity of the single-cell egg is then translated into cellular differences in the developing embryo by early cleavage planes, helping to create the cell-cell interactions that contribute to producing the body axis. Despite the importance of polarity in oocytes, little is understood about its generation, molecular nature, or interpretation.Vgl mRNA, a member of the transforming growth factor ,8 (TGFB) superfamily, is localized to the vegetal cortex of late-stage oocytes and remains in the vegetal hemisphere of early embryos until after gastrulation (5, 7). Synthesized early during oogenesis and distributed homogeneously throughout stage I and II oocytes, Vgl message accumulates along the vegetal cortex of oocytes during a short period in the middle of oogenesis, where it remains until maturation (8-10). Localization of Vgl mRNA appears to occur in at least two steps, the first being translocation ofthe message to the vegeta...

An3 mRNA encodes an RNA helicase that colocalizes with nucleoli in Xenopus oocytes in a stage-specific manner.

“…Maternal mRNAs localized to the vegetal hemisphere include two transforming growth factor f3 (TGFIB)-like proteins [Vgl (3) and TGF(35 (4)] and a protein with homology to Drosophila nanos (5). Although the function of these localized mRNAs is still not certain, the Vgl protein may serve as the maternal inducer of embryonic mesoderm formation.…”

Possible involvement of insulin-like growth factor 2 mRNA-binding protein 3 in zebrafish oocyte maturation as a novel cyclin B1 mRNA-binding protein that represses the translation in immature oocytes

“…In immature Xenopus oocytes, IMP3 is bound to Vg1 mRNA that localizes to the vegetal cortex [14], and its phosphorylation by MAP kinase during oocyte maturation induces cortical detachment and solubilization of Vg1 mRNA [15]. While these findings suggest that IMP3 plays a role in the localization of Vg1 mRNA, the involvement of IMP3 in the translational repression or activation of Vg1 mRNA in…”

“…IMP3 was described as a protein bound to the vegetal localization element (VLE) in the 3'UTR of Vg1 mRNA that localizes to the vegetal cortex of Xenopus oocytes (thereby also called Vg1 RNA-binding protein) [14]. Since the 3' UTR of zebrafish cyclin B1 mRNA harbors an E2 motif (UUCAC), a cis-acting element in the VLE [16], it is plausible that IMP3 binds to cyclin B1 mRNA via this motif in zebrafish oocytes.…”

“…The molecular mass of Xenopus IMP3 is 69 kDa [14] and that estimated from the cDNA sequence of 9 zebrafish IMP3 is 64 kDa. The anti-IMP3-positive 69-kDa protein was enriched in fractions 7 and 8, similar to cyclin B1 mRNA, Pumilio1 and CPEB (Fig.…”

GRSF-1: a poly(A)+mRNA binding protein which interacts with a conserved G-rich element

“…In Xenopus, the interaction between a 340-nucleotide sequence in the 3' UTR of vg 1 mRNA and a 69 kDa protein directs the vegetal localization of the transcript (34,35). Cooperation between staufen (a double-stranded RNA binding protein) and the 3' UTR of bicoid mRNA is required for anterior and posterior pole localization of the mRNA in Drosophila oocytes (36,37).…”