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The Ccr4-Not complex has been implicated in the control of multiple steps of mRNA metabolism; however, its functions in transcription remain ambiguous. The discovery that Ccr4/Pop2 is the major cytoplasmic mRNA deadenylase and the detection of Not proteins within mRNA processing bodies have raised questions about the roles of the Ccr4-Not complex in transcription. Here we firmly establish Ccr4-Not as a positive elongation factor for RNA polymerase II (RNAPII). The Ccr4-Not complex is targeted to the coding region of genes in a transcription-dependent manner similar to RNAPII and promotes elongation in vivo. Furthermore, Ccr4-Not interacts directly with elongating RNAPII complexes and stimulates transcription elongation of arrested polymerase in vitro. Ccr4-Not can reactivate backtracked RNAPII using a mechanism different from that of the well-characterized elongation factor TFIIS. While not essential for its interaction with elongation complexes, Ccr4-Not interacts with the emerging transcript and promotes elongation in a manner dependent on transcript length, although this interaction is not required for it to bind RNAPII. Our comprehensive analysis shows that Ccr4-Not directly regulates transcription, and suggests it does so by promoting the resumption of elongation of arrested RNAPII when it encounters transcriptional blocks in vivo.

BackgroundBurnout among nurses not only threatens their own health, but also that of their patients. Exploring risk factors of nurse’ burnout is important to improve nurses’ health and to increase the quality of health care services. This study aims to explore the relationship between work-family conflict and burnout among Chinese female nurses and the mediating role of psychological capital in this relationship.MethodsThis cross-sectional study was performed during the period of September and October 2010. A questionnaire that consisted of the Maslach Burnout Inventory-General Survey (MBI-GS), the work-family conflict scale and the psychological capital questionnaire (PCQ-24) scale, as well as demographic and working factors, was distributed to nurses in Liaoning province, China. A total of 1,332 individuals (effective response rate: 78.35%) became our subjects. Hierarchical linear regression analyses were performed to explore the mediating role of psychological capital.ResultsBoth work interfering family conflict and family interfering work conflict were positively related with emotional exhaustion and cynicism. However, work interfering family conflict was positively related with professional efficacy whereas family interfering work conflict was negatively related with it. Psychological capital partially mediated the relationship of work interfering family conflict with emotional exhaustion and cynicism; and partially mediated the relationship of family interfering work conflict with emotional exhaustion, cynicism and professional efficacy.ConclusionWork-family conflict had effects on burnout and psychological capital was a mediator in this relationship among Chinese nurses. Psychological capital was a positive resource for fighting against nurses’ burnout.

BackgroundAlthough occupational stress is an identified predictor of depressive symptoms, the mechanism behind the association is not well understood. The purpose of this study was to examine how psychological capital (PsyCap), a positive psychological state, mediates the association between occupational stress and depressive symptoms among Chinese physicians.MethodsA cross-sectional survey was conducted in Liaoning Province, China, during September–October 2010. Self-administered questionnaires including items on depressive symptoms assessed by the Center for Epidemiologic Studies Depression Scale, occupational stress assessed by the effort–reward imbalance scale and PsyCap estimated by a 24-item Psychological Capital Questionnaire, together with age, gender, marital status and education were distributed to 1300 physicians employed in large general hospitals. The final sample consisted of 998 participants. Asymptotic and resampling strategies were used to examine how PsyCap mediates the association between occupational stress and depressive symptoms.ResultsBoth the effort/reward ratio (ERR) and overcommitment were significantly associated with depressive symptoms among male and female physicians. There was a gender difference in the mediating role of PsyCap on the occupational stress–depressive symptoms association. For male physicians, PsyCap did not mediate the association between occupational stress and depressive symptoms. For female physicians, ERR and overcommitment were negatively associated with PsyCap, and PsyCap was negatively associated with depressive symptoms. As a result, PsyCap significantly mediated the associations of ERR and overcommitment with depressive symptoms. The proportion of PsyCap mediation was 19.07% for ERR, and 24.29% for overcommitment.ConclusionsPsyCap could be a positive resource for combating depressive symptoms in Chinese physicians. In addition to reducing occupational stress, PsyCap development should be included in depression prevention and treatment strategies, especially for female physicians.

In all domains of life, elongating RNA polymerases require the assistance of accessory factors to maintain their processivity and regulate their rate. Among these elongation factors, the Spt5/NusG factors stand out. Members of this protein family appear to be the only transcription accessory proteins that are universally conserved across all domains of life. In archaea and eukaryotes, Spt5 associates with a second protein, Spt4. In addition to regulating elongation, the eukaryotic Spt4-Spt5 complex appears to couple chromatin modification states and RNA processing to transcription elongation. This review discusses the experimental bases for our current understanding of Spt4-Spt5 function and recent studies that are beginning to elucidate the structure of Spt4-Spt5/RNA polymerase complexes and mechanism of Spt4-Spt5 action.

Pcf11 is one of numerous proteins involved in pre-mRNA 3-end processing and transcription termination. Using elongation complexes (ECs) formed from purified yeast RNA polymerase II (Pol II), we show that a 140-amino acid polypeptide from yeast Pcf11 is capable of dismantling the EC in vitro. This action depends on the C-terminal domain (CTD) of the largest subunit of Pol II and the CTD-interaction domain (CID) of Pcf11. Our experiments reveal a novel termination mechanism whereby Pcf11 bridges the CTD to the nascent transcript and causes dissociation of both Pol II and the nascent transcript from the DNA in the absence of nucleotide hydrolysis. We posit that conformational changes in the CTD are transduced through Pcf11 to the nascent transcript to cause termination. Transcription termination in eukaryotes is essential for recycling polymerase II (Pol II) and for preventing Pol II from perturbing promoters of genes located downstream from a transcription unit (Proudfoot et al. 2002;Proudfoot 2004). The mechanism of Pol II termination is poorly understood. Termination depends on the polyadenylation site in the nascent transcript. Numerous proteins involved in cleavage and polyadenylation also appear to participate in termination since mutations in several of these proteins affect both RNA processing and termination (Birse et al. 1998;Sadowski et al. 2003). Termination also depends on the C-terminal domain (CTD) of the largest subunit of Pol II (McCracken et al. 1997;Park et al. 2004). The CTD is composed of repeating heptapeptide motifs with the consensus YSPTSPS, and undergoes cycles of phosphorylation and dephosphorylation during the transcription process (Buratowski 2003). It functions as both a binding site and an allosteric regulator of the RNA processing machinery and serves to couple RNA processing and transcription elongation (Bentley 2002).In addition to the termination that occurs at the end of a transcription unit, premature termination within the body of genes can affect the processivity of Pol II in ways that regulate gene expression. One of the clearest examples of this is HIV. In the absence of the virally encoded protein Tat, transcription from the LTR results in accumulation of short transcripts in the cytoplasm due to premature termination (Feinberg et al. 1991;Kessler and Mathews 1992). HIV Tat modifies the elongation complex (EC) by recruiting the kinase, P-TEFb (Price 2000). P-TEFb phosphorylates the CTD of Pol II, and phosphorylation of the CTD increases the processivity of Pol II at least in part by counteracting the inhibitory action of two elongation factors, NELF and DSIF Yamaguchi et al. 1999Yamaguchi et al. , 2002Renner et al. 2001). Several studies have provided evidence that cellular activators stimulate gene expression by increasing the processivity of Pol II, although it is not clear if this occurs by preventing premature termination or by reducing the frequency of pauses and arrests (Krumm et al. 1993;Yankulov et al. 1994;Bentley 1995;Blau et al. 1996;Lis 1998;Lis et al. 2000;Barb...

RNA polymerase II (RNAPII) undergoes structural changes during the transitions from initiation, elongation, and termination, which are aided by a collection of proteins called elongation factors. NusG/Spt5 is the only elongation factor conserved in all domains of life. Although much information exists about the interactions between NusG/Spt5 and RNA polymerase in prokaryotes, little is known about how the binding of eukaryotic Spt4/5 affects the biochemical activities of RNAPII. We characterized the activities of Spt4/5 and interrogated the structural features of Spt5 required for it to interact with elongation complexes, bind nucleic acids, and promote transcription elongation. The eukaryotic specific regions of Spt5 containing the Kyrpides, Ouzounis, Woese domains are involved in stabilizing the association with the RNAPII elongation complex, which also requires the presence of the nascent transcript. Interestingly, we identify a region within the conserved NusG N-terminal (NGN) domain of Spt5 that contacts the non-template strand of DNA both upstream of RNAPII and in the transcription bubble. Mutating charged residues in this region of Spt5 did not prevent Spt4/5 binding to elongation complexes, but abrogated the cross-linking of Spt5 to DNA and the anti-arrest properties of Spt4/5, thus suggesting that contact between Spt5 (NGN) and DNA is required for Spt4/5 to promote elongation. We propose that the mechanism of how Spt5/NGN promotes elongation is fundamentally conserved; however, the eukaryotic specific regions of the protein evolved so that it can serve as a platform for other elongation factors and maintain its association with RNAPII as it navigates genomes packaged into chromatin.The conversion of DNA to RNA is a fundamental aspect of all life, and this process is carried out by RNA polymerases (RNAPs).2 These enzymatic powerhouses must maintain both high levels of fidelity and processivity over long distances to ensure that RNAs are accurately produced on a time scale amenable to life. Families of proteins called elongation factors have evolved to assist RNA polymerases during transcription elongation. The oldest and most conserved of these factors is the NusG/suppressor of Ty element (Spt) 5 family (1, 2). NusG is the eubacterial version of Spt5 and functions as a single polypeptide; however, archaea and eukaryotic Spt5 associate with an additional small protein, Spt4. In yeast, SPT5 is essential, but SPT4 is not. Deleting the gene encoding Spt4 impairs elongation, transcription-coupled repair, and mRNA processing (2-5). Some of the functions of Spt4 may be partially dependent on its ability to prevent degradation of Spt5 in cells (4). The NusG/Spt5 family of proteins has been shown to enhance RNA polymerase transcription elongation in all domains of life (6 -10). NusG regulates RNAP activity by stabilizing the post-translocated state thereby inhibiting backtracking and reducing long lifetime pauses (6, 11). The NusG homolog RfaH has also been implicated in regulating movement of the RNAP bridge hel...

Appropriate treatment of X-ray diffraction from an unoriented 18-heavy atom cluster derivative of a yeast RNA polymerase II crystal gave significant phase information to 5 A resolution. The validity of the phases was shown by close similarity of a 6 A electron density map to a 16 A molecular envelope of the polymerase from electron crystallography. Comparison of the 6 A X-ray map with results of electron crystallography of a paused transcription elongation complex suggests functional roles for two mobile protein domains: the tip of a flexible arm forms a downstream DNA clamp; and a hinged domain may serve as an RNA clamp, enclosing the transcript from about 8-18 residues upstream of the 3'-end in a tunnel.