Nucleic Acids Research Advance Access originally published online on November 22, 2007
Nucleic Acids Research 2008 36(2):353-363; doi:10.1093/nar/gkm990
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Nucleic Acids Research, 2008, Vol. 36, No. 2 353-363
© 2007 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Nucleic Acid Enzymes |
Cordycepin-hypersensitive growth links elevated polyphosphate levels to inhibition of poly(A) polymerase in Saccharomyces cerevisiae
1Institute of Molecular Biology, University of Zürich, Zürich, Switzerland, 2PhD Program in Molecular Life Sciences, University of Zürich and ETH Zürich, Zürich, Switzerland and 3The Institute of Plant Sciences, ETH Zürich, Zürich, Switzerland
*To whom correspondence should be addressed. Tel: + 41 44 635 3160; Fax: + 41 44 635 6811; Email: Bernhard.Dichtl{at}molbio.uzh.ch
Received July 12, 2007. Revised October 21, 2007. Accepted October 22, 2007.
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. 
pho80 and pho85 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 kD in the low nanomolar range. Poly P also bound mammalian poly(A) polymerase, however, with a 10-fold higher kD compared to yeast Pap1p. Genetic tests with double mutants of pho80 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 pho80 and pho85 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 pho80. Based on these observations, we suggest that binding of poly P to poly(A) polymerase negatively regulates its activity.
The authors wish it to be known that, in their opinion, the first two authors should be regarded as joint First Authors.