Gcn4 Binding in Coding Regions Can Activate Internal and Canonical 5' Promoters in Yeast.

Gcn4 is a yeast transcriptional activator induced by amino acid starvation. ChIP-seq analysis revealed 546 genomic sites occupied by Gcn4 in starved cells, representing ∼30% of Gcn4-binding motifs. Surprisingly, only ∼40% of the bound sites are in promoters, of which only ∼60% activate transcription, indicating extensive negative control over Gcn4 function. Most of the remaining ∼300 Gcn4-bound sites are within coding sequences (CDSs), with ∼75 representing the only bound sites near Gcn4-induced genes. Many such unconventional sites map between divergent antisense and sub-genic sense transcripts induced within CDSs adjacent to induced TBP peaks, consistent with Gcn4 activation of cryptic bidirectional internal promoters. Mutational analysis confirms that Gcn4 sites within CDSs can activate sub-genic and full-length transcripts from the same or adjacent genes, showing that functional Gcn4 binding is not confined to promoters. Our results show that internal promoters can be regulated by an activator that functions at conventional 5'-positioned promoters.

SWI/SNF and RSC cooperate to reposition and evict promoter nucleosomes at highly expressed genes in yeast.

The nucleosome remodeling complex RSC functions throughout the yeast genome to set the positions of -1 and +1 nucleosomes and thereby determines the widths of nucleosome-depleted regions (NDRs). The related complex SWI/SNF participates in nucleosome remodeling/eviction and promoter activation at certain yeast genes, including those activated by transcription factor Gcn4, but did not appear to function broadly in establishing NDRs. By analyzing the large cohort of Gcn4-induced genes in mutants lacking the catalytic subunits of SWI/SNF or RSC, we uncovered cooperation between these remodelers in evicting nucleosomes from different locations in the promoter and repositioning the +1 nucleosome downstream to produce wider NDRs-highly depleted of nucleosomes-during transcriptional activation. SWI/SNF also functions on a par with RSC at the most highly transcribed constitutively expressed genes, suggesting general cooperation by these remodelers for maximal transcription. SWI/SNF and RSC occupancies are greatest at the most highly expressed genes, consistent with their cooperative functions in nucleosome remodeling and transcriptional activation. Thus, SWI/SNF acts comparably with RSC in forming wide nucleosome-free NDRs to achieve high-level transcription but only at the most highly expressed genes exhibiting the greatest SWI/SNF occupancies.

Differential requirements for Gcn5 and NuA4 HAT activities in the starvation-induced versus basal transcriptomes.

The histone acetyltransferase (HAT) subunit of coactivator complex SAGA, Gcn5, stimulates eviction of promoter nucleosomes at certain highly expressed yeast genes, including those activated by transcription factor Gcn4 in amino acid-deprived cells; however, the importance of other HAT complexes in this process was poorly understood. Analyzing mutations that disrupt the integrity or activity of HAT complexes NuA4 or NuA3, or HAT Rtt109, revealed that only NuA4 acts on par with Gcn5, and functions additively, in evicting and repositioning promoter nucleosomes and stimulating transcription of starvation-induced genes. NuA4 is generally more important than Gcn5, however, in promoter nucleosome eviction, TBP recruitment, and transcription at most other genes expressed constitutively. NuA4 also predominates over Gcn5 in stimulating TBP recruitment and transcription of genes categorized as principally dependent on the cofactor TFIID versus SAGA, except for the most highly expressed subset including ribosomal protein genes, where Gcn5 contributes strongly to PIC assembly and transcription. Both SAGA and NuA4 are recruited to promoter regions of starvation-induced genes in a manner that might be feedback controlled by their HAT activities. Our findings reveal an intricate interplay between these two HATs in nucleosome eviction, PIC assembly, and transcription that differs between the starvation-induced and basal transcriptomes.

Distinct functions of three chromatin remodelers in activator binding and preinitiation complex assembly.

The nucleosome remodeling complexes (CRs) SWI/SNF, RSC, and Ino80C cooperate in evicting or repositioning nucleosomes to produce nucleosome depleted regions (NDRs) at the promoters of many yeast genes induced by amino acid starvation. We analyzed mutants depleted of the catalytic subunits of these CRs for binding of transcriptional activator Gcn4 and recruitment of TATA-binding protein (TBP) during preinitiation complex (PIC) assembly. RSC and Ino80 were found to enhance Gcn4 binding to both UAS elements in NDRs upstream of promoters and to unconventional binding sites within nucleosome-occupied coding sequences; and SWI/SNF contributes to UAS binding when RSC is depleted. All three CRs are actively recruited by Gcn4 to most UAS elements and appear to enhance Gcn4 binding by reducing nucleosome occupancies at the binding motifs, indicating a positive regulatory loop. SWI/SNF acts unexpectedly in WT cells to prevent excessive Gcn4 binding at many UAS elements, indicating a dual mode of action that is modulated by the presence of RSC. RSC and SWI/SNF collaborate to enhance TBP recruitment at Gcn4 target genes, together with Ino80C, in a manner associated with nucleosome eviction at the TBP binding sites. Cooperation among the CRs in TBP recruitment is also evident at the highly transcribed ribosomal protein genes, while RSC and Ino80C act more broadly than SWI/SNF at the majority of other constitutively expressed genes to stimulate this step in PIC assembly. Our findings indicate a complex interplay among the CRs in evicting promoter nucleosomes to regulate activator binding and stimulate PIC assembly.

Rational construction of dicarbonous CB/CNT@PI composites toward low filler loading and low-frequency electromagnetic wave absorption.

With the application of low frequency radar and the demand for stealth of high temperature resistant components, it is increasingly urgent to develop absorbing materials with both low frequency and high temperature resistant properties. Here, we successfully prepared various carbon/polyimide composites as low-frequency electromagnetic wave (EMW) absorbing materials by simple blending method. The well-designed mesh lap structure introduces a large amount of free space, further optimizing the impedance matching of the material. At the same time, the multiple loss mechanism formed by the combination of carbon black dominated polarization and carbon nanotube dominated conductive loss enhances the loss of incident EMW. The results showed that only 10 wt% filler loading of the CB/CNT@PI is achieved in the low frequency range (1-4 GHz) with a minimum reflection loss strength of -18.3 dB, which has obvious advantages compared with other works in recent years. This study provides a way for the design and preparation of resin-based absorbing materials.

Chromatin remodeler Ino80C acts independently of H2A.Z to evict promoter nucleosomes and stimulate transcription of highly expressed genes in yeast.

The chromatin remodelers SWI/SNF and RSC function in evicting promoter nucleosomes at highly expressed yeast genes, particularly those activated by transcription factor Gcn4. Ino80 remodeling complex (Ino80C) can establish nucleosome-depleted regions (NDRs) in reconstituted chromatin, and was implicated in removing histone variant H2A.Z from the -1 and +1 nucleosomes flanking NDRs; however, Ino80C's function in transcriptional activation in vivo is not well understood. Analyzing the cohort of Gcn4-induced genes in ino80Δ mutants has uncovered a role for Ino80C on par with SWI/SNF in evicting promoter nucleosomes and transcriptional activation. Compared to SWI/SNF, Ino80C generally functions over a wider region, spanning the -1 and +1 nucleosomes, NDR and proximal genic nucleosomes, at genes highly dependent on its function. Defects in nucleosome eviction in ino80Δ cells are frequently accompanied by reduced promoter occupancies of TBP, and diminished transcription; and Ino80 is enriched at genes requiring its remodeler activity. Importantly, nuclear depletion of Ino80 impairs promoter nucleosome eviction even in a mutant lacking H2A.Z. Thus, Ino80C acts widely in the yeast genome together with RSC and SWI/SNF in evicting promoter nucleosomes and enhancing transcription, all in a manner at least partly independent of H2A.Z editing.

Forecasting the incidence of mumps in Chongqing based on a SARIMA model.

BACKGROUND: Mumps is classified as a class C infection disease in China, and the Chongqing area has one of the highest incidence rates in the country. We aimed to establish a prediction model for mumps in Chongqing and analyze its seasonality, which is important for risk analysis and allocation of resources in the health sector. METHODS: Data on incidence of mumps from January 2004 to December 2018 were obtained from Chongqing Municipal Bureau of Disease Control and Prevention. The incidence of mumps from 2004 to 2017 was fitted using a seasonal autoregressive comprehensive moving average (SARIMA) model. The root mean square error (RMSE) and mean absolute percentage error (MAPE) were used to compare the goodness of fit of the models. The 2018 incidence data were used for validation. RESULTS: From 2004 to 2018, a total of 159,181 cases (93,655 males and 65,526 females) of mumps were reported in Chongqing, with significantly more men than women. The age group of 0-19 years old accounted for 92.41% of all reported cases, and students made up the largest proportion (62.83%), followed by scattered children and children in kindergarten. The SARIMA(2, 1, 1) × (0, 1, 1)12 was the best fit model, RMSE and MAPE were 0.9950 and 39.8396%, respectively. CONCLUSION: Based on the study findings, the incidence of mumps in Chongqing has an obvious seasonal trend, and SARIMA(2, 1, 1) × (0, 1, 1)12 model can also predict the incidence of mumps well. The SARIMA model of time series analysis is a feasible and simple method for predicting mumps in Chongqing.

Forecasting the incidence of acute haemorrhagic conjunctivitis in Chongqing: a time series analysis.

Acute haemorrhagic conjunctivitis is a highly contagious eye disease, the prediction of acute haemorrhagic conjunctivitis is very important to prevent and grasp its development trend. We use the exponential smoothing model and the seasonal autoregressive integrated moving average (SARIMA) model to analyse and predict. The monthly incidence data from 2004 to 2017 were used to fit two models, the actual incidence of acute haemorrhagic conjunctivitis in 2018 was used to validate the model. Finally, the prediction effect of exponential smoothing is best, the mean square error and the mean absolute percentage error were 0.0152 and 0.1871, respectively. In addition, the incidence of acute haemorrhagic conjunctivitis in Chongqing had a seasonal trend characteristic, with the peak period from June to September each year.

Genome-wide cooperation by HAT Gcn5, remodeler SWI/SNF, and chaperone Ydj1 in promoter nucleosome eviction and transcriptional activation.

Chaperones, nucleosome remodeling complexes, and histone acetyltransferases have been implicated in nucleosome disassembly at promoters of particular yeast genes, but whether these cofactors function ubiquitously, as well as the impact of nucleosome eviction on transcription genome-wide, is poorly understood. We used chromatin immunoprecipitation of histone H3 and RNA polymerase II (Pol II) in mutants lacking single or multiple cofactors to address these issues for about 200 genes belonging to the Gcn4 transcriptome, of which about 70 exhibit marked reductions in H3 promoter occupancy on induction by amino acid starvation. Examining four target genes in a panel of mutants indicated that SWI/SNF, Gcn5, the Hsp70 cochaperone Ydj1, and chromatin-associated factor Yta7 are required downstream from Gcn4 binding, whereas Asf1/Rtt109, Nap1, RSC, and H2AZ are dispensable for robust H3 eviction in otherwise wild-type cells. Using ChIP-seq to interrogate all 70 exemplar genes in single, double, and triple mutants implicated Gcn5, Snf2, and Ydj1 in H3 eviction at most, but not all, Gcn4 target promoters, with Gcn5 generally playing the greatest role and Ydj1 the least. Remarkably, these three cofactors cooperate similarly in H3 eviction at virtually all yeast promoters. Defective H3 eviction in cofactor mutants was coupled with reduced Pol II occupancies for the Gcn4 transcriptome and the most highly expressed uninduced genes, but the relative Pol II levels at most genes were unaffected or even elevated. These findings indicate that nucleosome eviction is crucial for robust transcription of highly expressed genes but that other steps in gene activation are more rate-limiting for most other yeast genes.

Phosphorylated Pol II CTD recruits multiple HDACs, including Rpd3C(S), for methylation-dependent deacetylation of ORF nucleosomes.

Methylation of histone H3 by Set1 and Set2 is required for deacetylation of nucleosomes in coding regions by histone deacetylase complexes (HDACs) Set3C and Rpd3C(S), respectively. We report that Set3C and Rpd3C(S) are cotranscriptionally recruited in the absence of Set1 and Set2, but in a manner stimulated by Pol II CTD kinase Cdk7/Kin28. Consistently, Rpd3C(S) and Set3C interact with Ser5-phosphorylated Pol II and histones in extracts, but only the histone interactions require H3 methylation. Moreover, reconstituted Rpd3C(S) binds specifically to Ser5-phosphorylated CTD peptides in vitro. Hence, whereas interaction with methylated H3 residues is required for Rpd3C(S) and Set3C deacetylation activities, their cotranscriptional recruitment is stimulated by the phosphorylated CTD. We further demonstrate that Rpd3, Hos2, and Hda1 have overlapping functions in deacetylating histones and suppressing cotranscriptional histone eviction. A strong correlation between increased acetylation and lower histone occupancy in HDA mutants implies that histone acetylation is important for nucleosome eviction.

Phosphorylation of the Pol II CTD by KIN28 enhances BUR1/BUR2 recruitment and Ser2 CTD phosphorylation near promoters.

Cyclin-dependent kinase BUR1/BUR2 appears to be the yeast ortholog of P-TEFb, which phosphorylates Ser2 of the RNA Pol II CTD, but the importance of BUR1/BUR2 in CTD phosphorylation is unclear. We show that BUR1/BUR2 is cotranscriptionally recruited to the 5' end of ARG1 in a manner stimulated by interaction of the BUR1 C terminus with CTD repeats phosphorylated on Ser5 by KIN28. Impairing BUR1/BUR2 function, or removing the CTD-interaction domain in BUR1, reduces Ser2 phosphorylation in bulk Pol II and eliminates the residual Ser2P in cells lacking the major Ser2 CTD kinase, CTK1. Impairing BUR1/BUR2 or CTK1 evokes a similar reduction of Ser2P in Pol II phosphorylated on Ser5 and in elongating Pol II near the ARG1 promoter. By contrast, CTK1 is responsible for the bulk of Ser2P in total Pol II and at promoter-distal sites. In addition to phosphorylating Ser2 near promoters, BUR1/BUR2 also stimulates Ser2P formation by CTK1 during transcription elongation.

Accumulation of a threonine biosynthetic intermediate attenuates general amino acid control by accelerating degradation of Gcn4 via Pho85 and Cdk8.

Gcn4 is a master transcriptional regulator of amino acid and vitamin biosynthetic enzymes subject to the general amino acid control (GAAC), whose expression is upregulated in response to amino acid starvation in Saccharomyces cerevisiae. We found that accumulation of the threonine pathway intermediate ß-aspartate semialdehyde (ASA), substrate of homoserine dehydrogenase (Hom6), attenuates the GAAC transcriptional response by accelerating degradation of Gcn4, already an exceedingly unstable protein, in cells starved for isoleucine and valine. The reduction in Gcn4 abundance on ASA accumulation requires Cdk8/Srb10 and Pho85, cyclin-dependent kinases (CDKs) known to mediate rapid turnover of Gcn4 by the proteasome via phosphorylation of the Gcn4 activation domain under nonstarvation conditions. Interestingly, rescue of Gcn4 abundance in hom6 cells by elimination of SRB10 is not accompanied by recovery of transcriptional activation, while equivalent rescue of UAS-bound Gcn4 in hom6 pho85 cells restores greater than wild-type activation of Gcn4 target genes. These and other findings suggest that the two CDKs target different populations of Gcn4 on ASA accumulation, with Srb10 clearing mostly inactive Gcn4 molecules at the promoter that are enriched for sumoylation of the activation domain, and Pho85 clearing molecules unbound to the UAS that include both fully functional and inactive Gcn4 species.

Pol II CTD kinases Bur1 and Kin28 promote Spt5 CTR-independent recruitment of Paf1 complex.

Paf1 complex (Paf1C) is a transcription elongation factor whose recruitment is stimulated by Spt5 and the CDKs Kin28 and Bur1, which phosphorylate the Pol II C-terminal domain (CTD) on Serines 2, 5, and 7. Bur1 promotes Paf1C recruitment by phosphorylating C-terminal repeats (CTRs) in Spt5, and we show that Kin28 enhances Spt5 phosphorylation by promoting Bur1 recruitment. It was unclear, however, whether CTD phosphorylation by Kin28 or Bur1 also stimulates Paf1C recruitment. We find that Paf1C and its Cdc73 subunit bind diphosphorylated CTD repeats (pCTD) and phosphorylated Spt5 CTRs (pCTRs) in vitro, and that cdc73 mutations eliminating both activities reduce Paf1C recruitment in vivo. Phosphomimetic (acidic) substitutions in the Spt5 CTR sustain high-level Paf1C recruitment in otherwise wild-type cells, but not following inactivation of Bur1 or Kin28. Furthermore, inactivating the pCTD/pCTR-interaction domain (PCID) in Cdc73 decreases Paf1C-dependent histone methylation in cells containing non-phosphorylatable Spt5 CTRs. These results identify an Spt5 pCTR-independent pathway of Paf1C recruitment requiring Kin28, Bur1, and the Cdc73 PCID. We propose that pCTD repeats and Spt5 pCTRs provide separate interaction surfaces that cooperate to ensure high-level Paf1C recruitment.

Decapping factor Dcp2 controls mRNA abundance and translation to adjust metabolism and filamentation to nutrient availability.

Degradation of most yeast mRNAs involves decapping by Dcp1/Dcp2. DEAD-box protein Dhh1 has been implicated as an activator of decapping, in coupling codon non-optimality to enhanced degradation, and as a translational repressor, but its functions in cells are incompletely understood. RNA-Seq analyses coupled with CAGE sequencing of all capped mRNAs revealed increased abundance of hundreds of mRNAs in dcp2Δ cells that appears to result directly from impaired decapping rather than elevated transcription. Interestingly, only a subset of mRNAs requires Dhh1 for targeting by Dcp2, and also generally requires the other decapping activators Pat1, Edc3, or Scd6; whereas most of the remaining transcripts utilize nonsense-mediated mRNA decay factors for Dcp2-mediated turnover. Neither inefficient translation initiation nor stalled elongation appears to be a major driver of Dhh1-enhanced mRNA degradation. Surprisingly, ribosome profiling revealed that dcp2Δ confers widespread changes in relative translational efficiencies (TEs) that generally favor well-translated mRNAs. Because ribosome biogenesis is reduced while capped mRNA abundance is increased by dcp2Δ, we propose that an increased ratio of mRNA to ribosomes increases competition among mRNAs for limiting ribosomes to favor efficiently translated mRNAs in dcp2Δ cells. Interestingly, genes involved in respiration or utilization of alternative carbon or nitrogen sources are upregulated, and both mitochondrial function and cell filamentation are elevated in dcp2Δ cells, suggesting that decapping sculpts gene expression post-transcriptionally to fine-tune metabolic pathways and morphological transitions according to nutrient availability.

Decapping factor Dcp2 controls mRNA abundance and translation to adjust metabolism and filamentation to nutrient availability.

Degradation of most yeast mRNAs involves decapping by Dcp1/Dcp2. DEAD-box protein Dhh1 has been implicated as an activator of decapping, in coupling codon non-optimality to enhanced degradation, and as a translational repressor, but its functions in cells are incompletely understood. RNA-Seq analyses coupled with CAGE sequencing of all capped mRNAs revealed increased abundance of hundreds of mRNAs in dcp2 Δ cells that appears to result directly from impaired decapping rather than elevated transcription, which was confirmed by ChIP-Seq analysis of RNA Polymerase II occupancies genome-wide. Interestingly, only a subset of mRNAs requires Dhh1 for targeting by Dcp2, and also generally requires the other decapping activators Pat1, Lsm2, Edc3 or Scd6; whereas most of the remaining transcripts utilize NMD factors for Dcp2-mediated turnover. Neither inefficient translation initiation nor stalled elongation appears to be a major driver of Dhh1-enhanced mRNA degradation. Surprisingly, ribosome profiling revealed that dcp2 Δ confers widespread changes in relative TEs that generally favor well-translated mRNAs. Because ribosome biogenesis is reduced while capped mRNA abundance is increased by dcp2 Δ, we propose that an increased ratio of mRNA to ribosomes increases competition among mRNAs for limiting ribosomes to favor efficiently translated mRNAs in dcp2 Δ cells. Interestingly, genes involved in respiration or utilization of alternative carbon or nitrogen sources are derepressed, and both mitochondrial function and cell filamentation (a strategy for nutrient foraging) are elevated by dcp2 Δ, suggesting that mRNA decapping sculpts gene expression post-transcriptionally to fine-tune metabolic pathways and morphological transitions according to nutrient availability.

Yeast arginine methyltransferase Hmt1p regulates transcription elongation and termination by methylating Npl3p.

The heterogeneous nuclear ribonucleoprotein Npl3p of budding yeast is a substrate of arginine methyltransferase Hmt1p, but the role of Hmt1p in regulating Npl3p's functions in transcription antitermination and elongation were unknown. We found that mutants lacking Hmt1p methyltransferase activity exhibit reduced recruitment of Npl3p, but elevated recruitment of a component of mRNA cleavage/termination factor CFI, to the activated GAL10-GAL7 locus. Consistent with this, hmt1 mutants displayed increased termination at the defective gal10-Delta56 terminator. Remarkably, hmt1Delta cells also exhibit diminished recruitment of elongation factor Tho2p and a reduced rate of transcription elongation in vivo. Importantly, the defects in Npl3p and Tho2p recruitment, antitermination and elongation in hmt1Delta cells all were mitigated by substitutions in Npl3p RGG repeats that functionally mimic arginine methylation by Hmt1p. Thus, Hmt1p promotes elongation and suppresses termination at cryptic terminators by methylating RGG repeats in Npl3p. As Hmt1p stimulates dissociation of Tho2p from an Npl3p-mRNP complex, it could act to recycle these elongation and antitermination factors back to sites of ongoing transcription.

Time series analysis of rubella incidence in Chongqing, China using SARIMA and BPNN mathematical models.

INTRODUCTION: Chongqing is among the areas with the highest rubella incidence rates in China. This study aimed to analyze the temporal distribution characteristics of rubella and establish a forecasting model in Chongqing, which could provide a tool for decision-making in the early warning system for the health sector. METHODOLOGY: The rubella monthly incidence data from 2004 to 2019 were obtained from the Chongqing Center of Disease and Control. The incidence from 2004 to June 2019 was fitted using the seasonal autoregressive integrated moving average (SARIMA) model and the back-propagation neural network (BPNN) model, and the data from July to December 2019 was used for validation. RESULTS: A total of 30,083 rubella cases were reported in this study, with a significantly higher average annual incidence before the nationwide introduction of rubella-containing vaccine (RCV). The peak of rubella notification was from April to June annually. Both SARIMA and BPNN models were capable of predicting the expected incidence of rubella. However, the linear SARIMA model fits and predicts better than the nonlinear BPNN model. CONCLUSIONS: Based on the results, rubella incidence in Chongqing has an obvious seasonal trend, and SARIMA (2,1,1) × (1,1,1) 12 model can predict the incidence of rubella well. The SARIMA model is a feasible tool for producing reliable rubella forecasts in Chongqing.

Activator Gcn4 employs multiple segments of Med15/Gal11, including the KIX domain, to recruit mediator to target genes in vivo.

Mediator is a multisubunit coactivator required for initiation by RNA polymerase II. The Mediator tail subdomain, containing Med15/Gal11, is a target of the activator Gcn4 in vivo, critical for recruitment of native Mediator or the Mediator tail subdomain present in sin4Delta cells. Although several Gal11 segments were previously shown to bind Gcn4 in vitro, the importance of these interactions for recruitment of Mediator and transcriptional activation by Gcn4 in cells was unknown. We show that interaction of Gcn4 with the Mediator tail in vitro and recruitment of this subcomplex and intact Mediator to the ARG1 promoter in vivo involve additive contributions from three different segments in the N terminus of Gal11. These include the KIX domain, which is a critical target of other activators, and a region that shares a conserved motif (B-box) with mammalian coactivator SRC-1, and we establish that B-box is a critical determinant of Mediator recruitment by Gcn4. We further demonstrate that Gcn4 binds to the Gal11 KIX domain directly and, by NMR chemical shift analysis combined with mutational studies, we identify the likely binding site for Gcn4 on the KIX surface. Gcn4 is distinctive in relying on comparable contributions from multiple segments of Gal11 for efficient recruitment of Mediator in vivo.

Yeast cap binding complex impedes recruitment of cleavage factor IA to weak termination sites.

Nuclear cap binding complex (CBC) is recruited cotranscriptionally and stimulates spliceosome assembly on nascent mRNAs; however, its possible functions in regulating transcription elongation or termination were not well understood. We show that, while CBC appears to be dispensable for normal rates and processivity of elongation by RNA polymerase II (Pol II), it plays a direct role in preventing polyadenylation at weak termination sites. Similarly to Npl3p, with which it interacts, CBC suppresses the weak terminator of the gal10-Delta56 mutant allele by impeding recruitment of termination factors Pcf11p and Rna15p (subunits of cleavage factor IA [CF IA]) and does so without influencing Npl3p occupancy at the termination site. Importantly, deletion of CBC subunits or NPL3 also increases termination at a naturally occurring weak poly(A) site in the RNA14 coding sequences. We also show that CBC is most likely recruited directly to the cap of nascent transcripts rather than interacting first with transcriptional activators or the phosphorylated C-terminal domain of Pol II. Thus, our findings illuminate the mechanism of CBC recruitment and extend its function in Saccharomyces cerevisiae beyond mRNA splicing and degradation of aberrant nuclear mRNAs to include regulation of CF IA recruitment at poly(A) selection sites.