Coordination of Endoplasmic Reticulum and mRNA Localization to the Yeast Bud

Abstract

Localization of messenger RNAs and local protein synthesis contribute to asymmetric protein distribution not only of cytoplasmic but also of membrane or secreted proteins. Since synthesis of the latter protein classes occurs at the rough endoplasmic reticulum (ER), mRNA localization and distribution of ER should be coordinated. However, this coordination is not yet understood. In yeast, mRNA localization to the growing bud depends on the myosin Myo4p, its adaptor She3p, and the specific RNA binding protein She2p. These proteins mediate the localization of 23 mRNAs including ASH1 mRNA and mRNAs encoding membrane proteins. In addition, Myo4p and She3p are required for segregation of cortical ER to the bud. Here we show, with ASH1 mRNA as a model mRNA, that localizing messenger ribonucleoprotein (mRNP) particles comigrate with tubular ER structures to the bud, which requires the RNA binding protein She2p. Coordinated movement of the ASH1 mRNP with ER tubules but not their association with each other depends on Myo4p and She3p. Subcellular fractionation experiments demonstrate a cosegregation of ER and She2p, which is independent of Myo4p, She3p, or polysomes. Our findings suggest a novel model for mRNA localization that involves association of She2p and mRNPs with ER tubules and myosin-dependent cotransport of tubules and localized mRNPs.

Structure of a myosin•adaptor complex and pairing by cargo

“…6B, two Myo4p•She3p heterotrimers are proposed to be cross-linked by putative tER ligands into a transportcompetent motor complex. Coupling two Myo4p•She3p heterotrimers by either She2p•zipcode or tER ligand(s) may occur separately or in concert, in agreement with reported observations of discrete transport or cotransport of these two different cargos (29,32). Again, as for ASH1 mRNA, multiple ligands arranged consecutively along the surface of tubular ER could recruit multiple motor complexes, each consisting of two cross-linked Myo4p•She3p heterotrimers, yielding a more processive mode of transport in a millipede-like fashion.…”

“…4D). We propose this highly conserved She3p coiled coil as a candidate binding site that represents the genetically mapped binding site on She3p for a hitherto unidentified ligand on the surface of tubular ER that can undergo transport from the mother to the daughter cell (28,32,39). As indicated in the model in Fig.…”

“…The modular arrangement of two distinct cargo-binding sites, one for mRNP and the other one for tER, would permit simultaneous transport of these two cargos. Indeed, coupled transport of zipcoded mRNPs and tER has been reported although cotransport appears not to be obligatory (29,32).…”

Hooking She3p onto She2p for myosin-mediated cytoplasmic mRNA transport

“…Instead, cargo binds via an "adaptor" protein, swi5p-dependent HO expression 3 protein (She3p) (7). Notably, the 425-residue-long She3p contains at least four binding modules that appear to be sequentially arranged along its primary structure (8)(9)(10)(11). Our recent composite crystal structure of a region of She3p (comprising residues showed that two She3p molecules form a pseudocoiled coil (6).…”

Monomeric myosin V uses two binding regions for the assembly of stable translocation complexes

“…In this translocation complex, the myosin motor, termed Myo4p, interacts with the N-terminal half of its adapter protein She3p to translocate at least 26 different types of mRNA as well as endoplasmic reticulum (ER) (15)(16)(17)(18)(19)(20)(21)(22)(23)(24). For mRNA transport, the adapter protein She3p binds with its C-terminal half to the RNA-binding protein She2p (16,19,22,23).…”