A role for the exosome in the in vivo degradation of unstable mRNAs

Abstract

In mammals, the mRNAs encoding many proteins involved in inflammation bear destabilizing AU-rich elements (AREs) in the 3-untranslated region. The exosome, a complex of 3 → 5 exonucleases, is rate limiting in the destruction of such mRNAs in a mammalian in vitro system, but a role in vivo has not been demonstrated. The phenomenon of ARE-mediated degradation also occurs in the protist parasite Trypanosoma brucei. Messenger RNAs with 3-untranslated region U-rich elements, which strongly resemble AREs, are extremely unstable in the trypanosome form that parasitizes mammals. The first step in degradation of these mRNAs in vivo is rapid destruction of the 3-untranslated region; subsequently the mRNA is destroyed by exonucleases acting in both 5 → 3 and 3 → 5 directions. We here investigated the roles of three subunits of the trypanosome exosome complex, RRP45, RRP4, and CSL4, in this process, depleting the individual subunits in vivo by inducible RNA interference. RRP45 depletion, which probably disrupts exosome integrity, caused a delay in the onset of degradation of the very unstable RNAs, but did not affect degradation of more stable species. Depletion of RRP4 or CSL4 does not affect the stability of the residual exosome and did not change mRNA degradation kinetics. We conclude that the exosome is required for the initiation of rapid degradation of unstable mRNAs in trypanosomes.

Cancer Cell-derived Exosomes Induce Mitogen-activated Protein Kinase-dependent Monocyte Survival by Transport of Functional Receptor Tyrosine Kinases

“…Exosomes mediate cell communication through fusion between the exosome membrane and the plasma membrane of the recipient cell, a process that requires the integrity of the exosome membrane (24). We confirmed this in our studies by showing that exosomes that had been disrupted had no effect on monocyte survival.…”

“…Previous studies suggested that the efficiency of exosomes in transport of functional biomolecules relies on their vesicle integrity (24). To validate the necessity of exosome integrity for monocyte survival, we disrupted MCF-7-derived exosomes by repeated freezing and thawing or by permeabilization of the exosome membrane with 0.1% saponin (25).…”

The evolutionarily conserved subunits Rrp4 and Csl4 confer different substrate specificities to the archaeal exosome

“…1). It is noteworthy that Rrp4 and Csl4 are conserved in the exosomes of Archaea and Eukarya [3,[16][17][18], suggesting important differential roles for these proteins, most probably in substrate selection. The archaeal exosome is a suitable system for analysis of the functions of Rrp4 and Csl4: in contrast to the eukaryotic exosome, active protein complexes with homotrimeric Rrp4 or Csl4 caps bound to the hexameric core can be reconstituted [6,[9][10][11].…”

Rrp6, Rrp47 and Cofactors of the Nuclear Exosome

“…While the majority of Rrp6 resides in the nucleus in S. cerevisiae, evidence suggests its presence in the cytoplasm in humans, T. brucei and A. thaliana. [1][2][3] In S. cerevisiae, where Rrp6 has been studied most extensively, deletion of the sole copy of its gene (RRP6) causes a slow growth phenotype at 30 o C and extremely poor growth at 37 o C. 4 Nevertheless, the fact that deletion of any of the other exosome genes causes lethality has made the use of rrp6-∆ strains a valuable tool for the study of exosome defects in nuclear RNA processing.…”