Oncogene (2007) 26, 7499–7507 & 2007 Nature Publishing Group All rights reserved 0950-9232/07 $30.00 www.nature.com/onc REVIEW Human RNA polymerase II-associated factor complex: dysregulation in cancer K Chaudhary1,3, S Deb1,3, N Moniaux1, MP Ponnusamy1 and SK Batra1,2 1Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, NE, USA and 2Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, Omaha, NE, USA Genetic instabilities are believed to be one of the major Keywords: PAF complex; transcription; tumor suppressor causes of developing a cancer phenotype in humans. genes; Wnt signaling During the progression of cancer, aberrant expression of proteins, either owing to genetic (amplification, mutation and deletion) or epigenetic modifications (DNA methyla- tion and histone deacetylation), contributes in different ways to the development of cancer. By differential Introduction screening analysis, an amplification of the 19q13 locus containing a novel pancreatic differentiation 2 (PD2) gene The dysregulation of critical genes is the crux of cancer was identified. PD2 is the human homolog of the yeast development and progression. This results in activation RNA polymerase II-associated factor 1 (yPaf1) and is of proto-oncogenes, inactivation of antioncogenes and part of the human RNA polymerase II-associated factor chromosomal instability (Batra et al., 1994; Vogelstein (hPAF) complex. hPAF is comprised of five subunits that and Kinzler, 2004). Genetic and epigenetic alterations include PD2/hPaf1, parafibromin, hLeo1, hCtr9 and lead to losses of or abnormal function of genes affecting hSki8. This multifaceted complex was first identified in processes that maintain or regulate orderly normal cell yeast (yPAF) and subsequently in Drosophila and human. function resulting in the phenotypic manifestation of Recent advances in the study on PAF have revealed specific types of the cancer (Thiagalingam, 2006). The various functions of the complex in human, which are aberrant expression of molecules that influence or are similar to yPAF, including efficient transcription elonga- involved in transcription and cell cycle, like cyclin D1 tion, mRNA quality control and cell-cycle regulation. (Knudsen et al., 2006) and Myc oncoprotein (Grandori Although the precise function of this complex in cancer is et al., 2004), is primarily responsible for oncogenic not clearly known, some of its subunits have been linked to signals to cells. These proteins are found to be a malignant phenotype. Its core subunit, PD2/hPaf1, is associated with cancer progression either by overexpres- amplified and overexpressed in many cancers. Further, an sion or by downregulation during the course of the overexpression of PD2/hPaf1 results in the induction of a disease. We have reported a new candidate oncogene, transformed phenotype, suggesting its possible involve- pancreatic differentiation 2 (PD2)/hPaf1, a member of ment in tumorigenesis. The parafibromin subunit of the the hPAF complex, which is dysregulated in cancer hPAF complex is a product of the HRPT-2 (hereditary (Moniaux et al., 2006). Another component of the hyperparathyroidism type 2) tumor suppressor gene, hPAF complex with a known role in cancer is the which is mutated in the germ line of hyperparathyroidism- hCdc73 subunit, also called parafibromin (Zhang et al., jaw tumor patients. This review focuses on the functions of 2006). The RNA polymerase II-associated factor (PAF) the PAF complex and its individual subunits, the complex was initially reported in yeast (yPAF) (Supple- interaction of the subunits with each other and/or with mentary text – section A) and comprises five subunits, other molecules, and dysregulation of the complex, Paf1, Ctr9, Leo1, Rtf1 and Cdc73 (Krogan et al., 2002). providing an insight into its potential involvement in the yPAF is associated with both the promoter and coding development of cancer. regions of transcriptionally active genes and plays a role Oncogene (2007) 26, 7499–7507; doi:10.1038/sj.onc.1210582; in transcription elongation. This complex also plays a published online 18 June 2007 role in mRNA processing and maturation, being responsible for maintaining proper poly(A) tail lengths (Mueller et al., 2004). Although both the yeast and human PAF (hPAF) Correspondence: Dr SK Batra, Department of Biochemistry and complexes have five subunits, the hPAF complex differs Molecular Biology, University of Nebraska Medical Center, Omaha, from yPAF in having an additional eukaryotic specific NE, 68198-5870, USA. hSki8 subunit and lacking the human homolog of yRtf1 E-mail: [email protected] 3These authors have contributed equally to this work. subunit (Figure 1). Rtf1 does not appear to be a stable Received 29 January 2007; revised 4April 2007; accepted 1 May 2007; part of the human and Drosophila PAF complex. published online 18 June 2007 However, Paf1 and Rtf1 associate during active hPAF complex in cancer K Chaudhary et al 7500 a hPAF1 531 aa b hCdc73 531 aa c hLeo1 666 aa d hCtr9 1173 aa e hSki8 305 aa Myc type helix- Leucine zipper RRM loop-helix Ser/Asp rich domain Glu domain Nuclear localization signal (NLS) Lys domain Tetratrico peptide repeat RGD WD40 repeat Figure 1 A schematic diagram of the structural domains of various components of human RNA polymerase II-associated factor (hPAF) complex. (a) Pancreatic differentiation 2 (PD2)/hPaf1 contains three Myc type helix–loop–helix, a leucine zipper, an RNA recognition motif (RRM), Ser/Asp-rich region with possible catalytic function, a Glu-rich domain and one nuclear localization signal (NLS). (b) Parafibromin/hCdc73 has five NLSs, but only two bipartite NLS shown are functional. (c) hLeo1 is a Ser/Asp-rich protein containing RGD (Arg-Gly-Asp) sequence that can function for cell adhesion, and an NLS domain. (d) hCtr9 is the largest component of the complex containing a tetratrico peptide repeat for protein–protein interactions. (e) hSki8 is a WD40-repeat-rich protein that is involved in post-transcriptional processing of RNA. WD40 repeats are found in a number of eukaryotic proteins that cover a wide variety of functions including adaptor/regulatory modules in signal transduction, pre-mRNA processing, cytoskeleton assembly and cell-cycle control. transcription in Drosophila (Supplementary text – Functions of the human PAF complex section B). Like the yPAF complex, hPAF regulates transcription-associated histone monoubiquitination of The hPAF complex was first identified during char- H2B, wherein recruitment of histone chaperone FACT acterization of proteins associated with parafibromin (facilitates chromatin transcription), hPAF and the H2B (Rozenblatt-Rosen et al., 2005). It is well documented monoubiquitination machinery is a necessary step for that hPAF shares four subunits with yPAF (Ctr9, Paf1, elongation by RNAP II (Pavri et al., 2006). The hCdc73 Leo1 and Cdc73) and also contains a novel higher subunit of hPAF induces cell-cycle arrest in the G1 eukaryotic-specific subunit, hSki8 (Zhu et al., 2005). phase by blocking expression of cyclin D1 (Zhang et al., Rozenblatt-Rosen et al. (2005) identified the hPAF 2006). Thus, like its yeast counterpart, the hPAF complex proteins by immunoprecipitation experiments complex may also have an important role in cell-cycle in HEK293T cells using polyclonal anti-parafibromin regulation. The additional subunit, hSki8, is a core antibody. Subsequently, Zhu et al. (2005) determined protein of the hSKI complex, and the association of the hPAF complex components via conventional chro- hSKI with actively transcribed genes is dependent on the matography, following elution with PD2/hPaf1 anti- presence of the hPAF complex. This review discusses the body, and identified a novel interaction partner, hSki8. information available on hPAF and its individual The authors suggest that this ambiguity of the result subunits, describing its role in transcription, cell cycle may be due to the difference in purification approaches and other cellular functions. It also examines the used by the two groups. Rozenblatt-Rosen et al. (2005) amplifications and losses of the genes encoding the used antibodies against hCdc73 followed by elution with complex proteins in various cancers and the ability of acidic glycine, which causes elution of large amounts of hPAF to function individually or as a complex in the IgG. The presence of IgG might have thus masked the initiation and progression of cancer. low-molecular-weight proteins such as hSki8. Oncogene hPAF complex in cancer K Chaudhary et al 7501 The hPAF complex interacts with the non-phosphory- subunit of the hPAF complex regulates cell cycle by lated and Ser2- and Ser5-phosphorylated forms of the blocking expression of cyclin D1 and causing G1-phase RNAP II large subunit (Rozenblatt-Rosen et al., 2005), arrest (Zhang et al., 2006). In yeast, the SKI complex, indicating its involvement in both transcription initiation which contains the Ski8 subunit, is localized in the cyto- and elongation. Removal of the nucleosomal barrier, plasm and, along with the exosome, is responsible for which involves the monoubiquitination of H2B, is a 30–50 mRNA degradation. The localization of the human prerequisite for efficient elongation on chromatin by SKI complex, unlike yPAF, is extended to the nucleus and RNAP II. Pavri et al. (2006) showed that the establish- the cytoplasm. Interestingly, hPAF and hSKI complexes ment of H2B monoubiquitination is dependent on hPAF, interact, and the association of hSKI with transcription- the histone chaperone FACT and transcription (Fig- ally active genes is dependent on the presence of hPAF, ure 2). Like its yeast counterpart, hPAF has also been suggesting a role of this complex in mRNA quality implicated in other cellular functions. The parafibromin control (Zhu et al., 2005). RNAPII a Nucleosome Stopped RNAPII b FACT RNAPII c FACT PAF d D1 D2 D3 PAF PAF PAF Set1 Set2 K4M RNAPII K36M RNAPII Rad6/Bre1 Rpt4 FACT FACT SAGA Ub Ub Ubp8 Ub Sug1 e RNAPII FACT PAF Figure 2 A schematic representation of polymerase II-associated factor (PAF) complex-mediated H2B monoubiquitination for efficient transcription through chromatin.