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Abstract
hDIS3 is a mainly nuclear, catalytic subunit of the human exosome complex, containing exonucleolytic (RNB) and endonucleolytic (PIN) active domains. Mutations in hDIS3 have been found in ∼10% of patients with multiple myeloma (MM). Here, we show that these mutations interfere with hDIS3 exonucleolytic activity. Yeast harboring corresponding mutations in DIS3 show growth inhibition and changes in nuclear RNA metabolism typical for exosome dysfunction. Construction of a conditional DIS3 knockout in the chicken DT40 cell line revealed that DIS3 is essential for cell survival, indicating that its function cannot be replaced by other exosome-associated nucleases: hDIS3L and hRRP6. Moreover, HEK293-derived cells, in which depletion of endogenous wild-type hDIS3 was complemented with exogenously expressed MM hDIS3 mutants, proliferate at a slower rate and exhibit aberrant RNA metabolism. Importantly, MM mutations are synthetically lethal with the hDIS3 PIN domain catalytic mutation both in yeast and human cells. Since mutations in PIN domain alone have little effect on cell physiology, our results predict the hDIS3 PIN domain as a potential drug target for MM patients with hDIS3 mutations. It is an interesting example of intramolecular synthetic lethality with putative therapeutic potential in humans.
Highlights
Multiple myeloma (MM) is a lethal neoplastic disease accounting for 10–15% of hematologic malignances and 20% of deaths related to cancer of the blood and bone marrow [1]
To analyze the impact of hDIS3 mutations identified in MM patients by Chapman et al [4] on the exonucleolytic activity of the protein, recombinant versions of hDIS3 [wild-type (WT) or bearing different mutations detected in MM] were purified from E. coli
Using our model cell lines, we examined whether expression of mutant hDIS3 proteins led to aberrations in metabolism of RNA molecules representing various classes
Summary
Multiple myeloma (MM) is a lethal neoplastic disease accounting for 10–15% of hematologic malignances and 20% of deaths related to cancer of the blood and bone marrow [1]. The human genome encodes three Dis homologues, of which only hDIS3 and hDIS3L were found to associate with the exosome [26,27] Both of them are processive 30–50 hydrolytic exonucleases, whereas only hDIS3 has retained endonuclease activity in its PIN domain. Apart from its well-documented role in RNA metabolism, a recent study has implicated the exosome in the activation-induced immunoglobulin heavy-chain class switch recombination and immunoglobulin variable region somatic hypermutation in human B lymphocytes [48]. We explored the consequences of hDIS3 gene mutations found in MM patients using biochemical, genetic and functional analyses, and examined possible synthetic genetic interactions between hDIS3 exonucleolytic activity and other catalytic activities of the nuclear exosome
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