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Cordycepin-hypersensitive growth links elevated polyphosphate levels to inhibition of poly(A) polymerase in Saccharomyces cerevisiae


Holbein, S; Freimoser, F M; Werner, T P; Wengi, A; Dichtl, B (2008). Cordycepin-hypersensitive growth links elevated polyphosphate levels to inhibition of poly(A) polymerase in Saccharomyces cerevisiae. Nucleic Acids Research, 36(2):353-363.

Abstract

To identify genes involved in poly(A) metabolism, we screened the yeast gene deletion collection for growth defects in the presence of cordycepin (3'-deoxyadenosine), a precursor to the RNA chain terminating ATP analog cordycepin triphosphate. Deltapho80 and Deltapho85 strains, which have a constitutively active phosphate-response pathway, were identified as cordycepin hypersensitive. We show that inorganic polyphosphate (poly P) accumulated in these strains and that poly P is a potent inhibitor of poly(A) polymerase activity in vitro. Binding analyses of poly P and yeast Pap1p revealed an interaction with a k(D) in the low nanomolar range. Poly P also bound mammalian poly(A) polymerase, however, with a 10-fold higher k(D) compared to yeast Pap1p. Genetic tests with double mutants of Deltapho80 and other genes involved in phosphate homeostasis and poly P accumulation suggest that poly P contributed to cordycepin hypersensitivity. Synergistic inhibition of mRNA synthesis through poly P-mediated inhibition of Pap1p and through cordycepin-mediated RNA chain termination may thus account for hypersensitive growth of Deltapho80 and Deltapho85 strains in the presence of the chain terminator. Consistent with this, a mutation in the 3'-end formation component rna14 was synthetic lethal in combination with Deltapho80. Based on these observations, we suggest that binding of poly P to poly(A) polymerase negatively regulates its activity.

To identify genes involved in poly(A) metabolism, we screened the yeast gene deletion collection for growth defects in the presence of cordycepin (3'-deoxyadenosine), a precursor to the RNA chain terminating ATP analog cordycepin triphosphate. Deltapho80 and Deltapho85 strains, which have a constitutively active phosphate-response pathway, were identified as cordycepin hypersensitive. We show that inorganic polyphosphate (poly P) accumulated in these strains and that poly P is a potent inhibitor of poly(A) polymerase activity in vitro. Binding analyses of poly P and yeast Pap1p revealed an interaction with a k(D) in the low nanomolar range. Poly P also bound mammalian poly(A) polymerase, however, with a 10-fold higher k(D) compared to yeast Pap1p. Genetic tests with double mutants of Deltapho80 and other genes involved in phosphate homeostasis and poly P accumulation suggest that poly P contributed to cordycepin hypersensitivity. Synergistic inhibition of mRNA synthesis through poly P-mediated inhibition of Pap1p and through cordycepin-mediated RNA chain termination may thus account for hypersensitive growth of Deltapho80 and Deltapho85 strains in the presence of the chain terminator. Consistent with this, a mutation in the 3'-end formation component rna14 was synthetic lethal in combination with Deltapho80. Based on these observations, we suggest that binding of poly P to poly(A) polymerase negatively regulates its activity.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:07 Faculty of Science > Institute of Molecular Life Sciences
Dewey Decimal Classification:570 Life sciences; biology
Language:English
Date:February 2008
Deposited On:07 Mar 2009 19:47
Last Modified:05 Apr 2016 13:09
Publisher:Oxford University Press
ISSN:0305-1048
Additional Information:Full final text Oxford Journal
Publisher DOI:https://doi.org/10.1093/nar/gkm990
PubMed ID:18033801
Permanent URL: https://doi.org/10.5167/uzh-17354

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