Role of nuclear RNP assembly in cytoplasmic mRNA localization

Messenger RNA localization occurs in the cytoplasm and allows
temporal and spatial regulation of gene expression. In yeast, the
localization of ASH1 mRNA to the tip of budding cells allows the
asymmetric sorting of Ash1 protein, which has a key function in the
regulation of mating-type switching. After cell division,
asymmetric distribution of Ash1p restricts mating type switching to
only the mother cell. The cytoplasmic transport of ASH1 mRNA to the
bud tip depends on the myosin Myo4p, its adaptor She3p, and the
specific RNA binding protein She2p. Three additional trans-acting
factors Khd1p, Puf6p and Loc1p are involved in this process. All
known RNA-binding proteins of ASH1 mRNA have revealed a nuclear
connection, when following their cellular distribution by indirect
immunofluorescence. Thus, an early step in the localization pathway
might be the early recruitment of specific trans-acting factors to
the mRNA already in the nucleus. The aim of this thesis was to
investigate how nuclear key events such as early binding to
localized transcripts and the subsequent assembly into a nuclear
RNP can account for effective RNA localization. Following the route
of She2p, it was possible to show nucleo-cytoplasmic shuttling of
this RNA binding protein. Moreover, ASH1 mRNA and She2p were found
accumulated within the nucleolus upon arrest of mRNA export.
Interestingly, two additional trans-acting factors, Loc1 and Puf6p,
both involved in ASH1 mRNA localization are also nucleolar
proteins. Moreover, She2’s nuclear history seems to be important
for an effective sorting of Ash1p. When restricting ASH1-She2p
association to the cytoplasmic compartment artificially, the ASH1
mRNA was still localized but was prematurely translated during its
transport. This suggests that nuclear RNP assembly has an influence
on the later stages of cytoplasmic translational control. The
nucleolus might represent the appropriate cellular compartment to
provide the spatial framework for the assembly of localization
competent RNPs since many RNPs are assembled in this region.