Partitioning and translation of mRNAs encoding soluble proteins on membrane-bound ribosomes.
In eukaryotic cells, it is generally accepted that protein synthesis is compartmentalized; soluble proteins are synthesized on free ribosomes, whereas secretory and membrane proteins are synthesized on endoplasmic reticulum (ER)-bound ribosomes. The partitioning of mRNAs that accompanies such compartmentalization arises early in protein synthesis, when ribosomes engaged in the translation of mRNAs encoding signal-sequence-bearing proteins are targeted to the ER. In this report, we use multiple cell fractionation protocols, in combination with cDNA microarray, nuclease protection, and Northern blot analyses, to assess the distribution of mRNAs between free and ER-bound ribosomes. We find a broad representation of mRNAs encoding soluble proteins in the ER fraction, with a subset of such mRNAs displaying substantial ER partitioning. In addition, we present evidence that membrane-bound ribosomes engage in the translation of mRNAs encoding soluble proteins. Single-cell in situ hybridization analysis of the subcellular distribution of mRNAs encoding ER-localized and soluble proteins identify two overall patterns of mRNA distribution in the cell-endoplasmic reticular and cytosolic. However, both partitioning patterns include a distinct perinuclear component. These results identify previously unappreciated roles for membrane-bound ribosomes in the subcellular compartmentalization of protein synthesis and indicate possible functions for the perinuclear membrane domain in mRNA sorting in the cell.
Lerner
Rachel S
RS
Departments of Cell Biology and Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina 27710, USA.
Seiser
Robert M
RM
Zheng
Tianli
T
Lager
Patrick J
PJ
Reedy
Mary C
MC
Keene
Jack D
JD
Nicchitta
Christopher V
CV
D002458:Cell Fractionation; D018076:DNA, Complementary / Q000235:genetics; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006801:Humans; D019169:Jurkat Cells; D008854:Microscopy, Electron; D020411:Oligonucleotide Array Sequence Analysis; D014176:Protein Biosynthesis / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism*
D018076:DNA, Complementary; D012333:RNA, Messenger