Background: Eukaryotic cells localize selected mRNAs to a region of the cel
l as a means to sequester proteins. Signals within the 3' untranslated regi
on (3'UTR) facilitate mRNA localization by both actin and microtubule cytos
keletal systems. Recently, an mRNA in the yeast Saccharomyces cerevisiae, A
SH1, was shown to coalesce into a discrete particle that is maintained at t
he bud tip. Mutations in five genes, SHE1 -SHE5, cause defects in particle
formation and/or localization of the ASH1 transcript. Factors at the destin
ation of the mRNA transport remain to be identified.
Results: We have developed a system to label mRNA in living yeast with gree
n fluorescent protein (GFP) and follow the dynamics of mRNA movement and lo
calization. Constitutively expressing an ASH1 mRNA containing the bacteriop
hage MS2 coat-protein binding site adjacent to the ASH1 3'UTR allowed us to
visualize ASH1 mRNA with an MS2-coat-protein-GFP fusion protein (together
denoted gRNA(ASH1)). The gRNA(ASH1) was restricted to the bud tip in small
to large budded cells, migrated to the bud neck prior to cell separation an
d then rapidly relocalized to the incipient site of bud growth. It also loc
alized to regions of polarized growth during mating. In cells lacking Bud6p
/Aip3p or Bni1p/She5p, which are involved in polarity establishment and act
in organization, gRNA(ASH1) migrated to the bud but failed to remain at the
bud tip. These studies reveal discrete transport and anchoring steps in mR
NA localization,
Conclusions: The ASH1 mRNA was maintained at sites of polarized growth thro
ughout the vegetative and mating cell cycles, Bud6p/Aip3p and Bni1p/She5p a
re required to maintain the transcript at the cortical bud cap.