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Molecular and Cellular Biology, April 2009, p. 2296-2307, Vol. 29, No. 8
0270-7306/09/$08.00+0     doi:10.1128/MCB.01514-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

The Essential N Terminus of the Pta1 Scaffold Protein Is Required for snoRNA Transcription Termination and Ssu72 Function but Is Dispensable for Pre-mRNA 3'-End Processing{triangledown}

Mohamed A. Ghazy,1 Xiaoyuan He,1 Badri Nath Singh,2 Michael Hampsey,2 and Claire Moore1*

Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, Massachusetts 02111,1 Department of Biochemistry, Robert Wood Johnson Medical School, Piscataway, New Jersey 088542

Received 29 September 2008/ Returned for modification 5 November 2008/ Accepted 12 January 2009

Saccharomyces cerevisiae Pta1 is a component of the cleavage/polyadenylation factor (CPF) 3'-end processing complex and functions in pre-mRNA cleavage, poly(A) addition, and transcription termination. In this study, we investigated the role of the N-terminal region of Pta1 in transcription and processing. We report that a deletion of the first 75 amino acids (pta1-{Delta}75) causes thermosensitive growth, while the deletion of an additional 25 amino acids is lethal. The pta1-{Delta}75 mutant is defective for snoRNA termination, RNA polymerase II C-terminal domain Ser5-P dephosphorylation, and gene looping but is fully functional for mRNA 3'-end processing. Furthermore, different regions of Pta1 interact with the CPF subunits Ssu72, Pti1, and Ysh1, supporting the idea that Pta1 acts as a scaffold to organize CPF. The first 300 amino acids of Pta1 are sufficient for interactions with Ssu72, which is needed for pre-mRNA cleavage. By the degron-mediated depletion of Pta1, we show that the removal of this essential region leads to a loss of Ssu72, yet surprisingly, in vitro cleavage and polyadenylation remain efficient. In addition, a fragment containing amino acids 1 to 300 suppresses 3'-end processing in wild-type extracts. These findings suggest that the amino terminus of Pta1 has an inhibitory effect and that this effect can be neutralized through the interaction with Ssu72.

* Corresponding author. Mailing address: Tufts University School of Medicine and Sackler School of Graduate Biomedical Sciences, 136 Harrison Ave., Boston, MA 02111. Phone: (617) 636-6935. Fax: (617) 636-0337. E-mail: claire.moore{at}tufts.edu

{triangledown} Published ahead of print on 2 February 2009.

Molecular and Cellular Biology, April 2009, p. 2296-2307, Vol. 29, No. 8
0270-7306/09/$08.00+0     doi:10.1128/MCB.01514-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Richard, P., Manley, J. L. (2009). Transcription termination by nuclear RNA polymerases. Genes Dev. 23: 1247-1269 [Abstract] [Full Text]  


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