Abstract:Cells are rejuvenated during gametogenesis. The induction of gametogenesis-related genes is the common response to dietary restriction across a variety of regimens and genotypes. Long-lived genetic DR-mimetics upregulate Ndt80. Constitutively upregulation of Ndt80 renders cells immortal. Ime1 and Ndt80 regulate ageing genes. Using an integrated approach the potential target genes of Ndt80 crucial for lifespan resetting were identified among them is the homolog of human BARKOR and telomerase TERT.
Introduction
DR Induces Gametogenesis Genes
Genetic DR-Mimetics Utilize Rejuvenation Processes
Ndt80 Activation Prevents Chronological Ageing
Ageing Genes Regulated by Meiotic Transcription Factors
NDT80 Target Genes Identification
Common Targets of Ime1 and Ndt80
Ndt80 Antagonist Sum1
Discussion
Ndt80 Target Gene Induction Can Slow-Down, Stop, and Reverse Ageing
Transcription Factor Target Genes Inference
Methods
Sequences
Significance
Tables & Figures
Table: Upregulated terms common to the DR response of wt, atg15 and erg6
Table: Sterol-related genes, ATG14 and EST2 scoring for regulation by Ndt80
Table: Top-scoring Ndt80 targets
Table: Top-scoring Ndt80 and Ime1 targets
Table: Motif number-corrected top-scoring Ndt80 target genes
Table: SUM1 regulation of OSH genes
In eukaryotes the genome average of 5'-UTR (upstream translated region) lengths is remarkably similar across diverse taxonomic classes, ranging from 100 to 200 bp [11897027; 18451266; 19169243; 20810668]. In contrast, the 5'-UTR length varies considerably among genes in a given genome, ranging from a few bp to several thousand [11591473; 11897027; 18451266]. Genes with a long 5'-UTR, such as those involved in development or meiosis, are generally highly and finely regulated [21965341]. The majority of transcription factor (TF) binding sites for coding genes lie within 500 bp upstream of the transcription start site (TSS) [21177963], while for non-coding genes, the binding sites are even closer to the TSS, as they are positioned between 300 bp upstream and 200 bp downstream of the TSS.
In Saccharomyces cerevisiae regulated transcription of most genes is mediated through short upstream regulatory sequences (437 bp), residing 100 - 200 bp upstream of the start codon [17418911]. Approximately 150 DNA-binding proteins that affect transcription have known consensus site(s), although 218 DNA-binding proteins are known.
For most yeast transcription factors, binding sites are positioned further upstream and vary over a wider range in TATA promoters than in TATA-less promoters. A group of 6 'proximal promoter motifs' (GAT1/GLN3/DAL80, FKH1/FKH2, PBF1/PBF2, RPN4, NDT80, and ROX1*) occur preferentially in TATA-less promoters and have strong preferences for binding close to the transcription start site in these promoters [21931670].
In budding yeast consensus sequences for known transcription factors can be obtained from the Saccharomyces Genome Database (SGD) [http://www.yeastgenome.org/], Saccharomyces Cerevisiae Promoter Database (SCPD) [http://rulai.cshl.edu/SCPD/], and YEASTRACT [http://www.yeastract.com/] as well as from ChIP-chip [15343339; 16522208] and from specific scientific articles [20739709].
The yeast transcription factor DNA-binding specificities for 112 DNA-binding proteins representing 19 distinct structural classes were systemically determined [19111667].
In this article, transcription factor target genes will be identified via a combination approach using different data sources. Sequence based identification (consensus sequence and/or weight matrix), genome-wide location data, gene expression profiles (knockout, overexpression, induction, and regulated process) and eventually phylogenetic conservation will be utilized:
Consensus sequence
Weight matrix for strength
Direct binding (ChIP)
Overexpression and deletion gene expression
Conservation
Dietary restriction (DR) is capable of extending lifespan in wide-variety of species; from small yeast up to big mammals, but its mechanisms is unknown. Transient expression of either one of two transcription factors (Ime1 and Ndt80) is sufficient to totally reset lifespan [21700873]. Ime1 is highly responsive to nutrients and a variety of stresses. It is upstream of Ndt80. Ndt80 target genes can be identified via the use of genome-wide expression studies on meiotic cells [9784122; 11101837; 16542486], especially the expression during middle meiosis (cluster 4, 7, and 13) [20716365], Ndt80 deletion and overexpression [9784122] and existence of Ndt80 binding sites [15684073]. NDT80 promoter was also replaced by an inducible promoter (MK-ER-NDT80) that can be turned on by administration of Estradiol [20716365].
Common to the transcriptional response to DR of wild-type (wt) as well as two DR-essential gene mutants (atg15 and erg6) was the upregulation of genes related to autophagy, sporulation, response to temperature stimulus and peroxisome and downregulation of genes majorly associated to ribosome [Table: Upregulated terms common to the DR response of wt, atg15 and erg6]. However, in comparison of these mutants to wild-type, there are concrete differences which indicate an impaired response of exactly these processes. erg6 majorly upregulates nucleolus-related genes (Benjamini p-value, p = 2.210^-110), while it downregulates genes associated with response to temperature stimulus (p = 1.410^-10), general substrate transporter (p = 6.810^-6), vacuolar protein catabolic process (p = 1.410^-10) and sporulation (p = 3.610^-5). Upon DR wild-type and atg15 upregulate sporulation (p = 3.610^-6). NDT80 is induced with increasing strength of glucose restriction. SRC1, a potential target gene of Ndt80 is also slightly higher upon DR and peaks at 0.5% glucose [34]. SRC1 is also 4-fold induced upon DR [22912585].
Numerous DR-essential gene mutants which prolong lifespan if inactivated (e.g. ras2, sch9, tor1, rpd3, hhf1, HAP4-Overexpression) induce expression of NDT80, while sgs1 a classical model of accelerated ageing downregulates NDT80. Ndt80 binding sites are in turn highly significant enriched in the promoter regions of the upregulated genes of DR-essential mutants.
Chronological ageing cells in absent of DR do not appear to age. Immobilized cells cease dividing, remain metabolic active and retain > 95% viability of periods of 17 days, while over the same time-course starved cells retained
Ime1 and Ndt80 regulate ageing genes in the gene-regulatory network more than expected by random (hypergeometric p-value = 0.001 and 0.01, respectively). Moreover, IME1 and NDT80 interact with far more ageing genes than expected by purely by chance (p
The consensus sequence of Ndt80 is 'gNCRCAAAY'. 800 bp upstream regions of all (7131) S. cerevisiae genes were obtained from Ensembl. Regular expression were designed and searched against all promoters. 331 gene promoters contained at least one perfect match. These genes were very highly enriched for Retrotransposons (p = 6.210^-31), RNA binding and ribosomes. There were also reproduction-related genes, but they did not reach significant enrichment. Among the 331 genes were four ageing genes, namely: mitochondrial glycerol-3-phosphate dehydrogenase (GUT2), Heat shock protein 104 (HSP104), cAMP-dependent protein kinase type 2 (TPK2) and Telomerase reverse transcriptase (EST2*). Transcription factors are capable of binding to variations of their consensus motif. For allowing one mismatch in the consensus motif, regular expression for each motif was generated. 4113 genes were found to harbour a Ndt80 consensus sequence motif with one base different from the perfect match. Searching only for the core motif (CRCAAA) yielded 2569 genes.
We finally identified Ndt80 target genes by integrating multiple datasets via an approach in which each gene gains score if it is found to be either significant bound by Ndt80, contains a consensus motif of Ndt80, upregulated upon Ndt80 overexpression or transient induction, and downregulated in NDT80 deletion mutants. About 19 genes obtained the highest possible score [Table: Top-scoring Ndt80 targets]. Most of these genes are associated to meiosis, as expected, but there are also a few others functions as well as some unknown genes.
As IME1 and NDT80 when transiently overexpressed are both capable to rejuvenate, the upregulated genes common to Ime1 and Ndt80 overexpression, which are also bound by either one of them, were determined. For instance, both induce HHO1, an H1-type linker histone represses recombination at the rDNA by a mechanism that is independent of Sir2. 11 highest scoring target genes of Ime1 and Ndt80 were identified [Table: Top-scoring Ndt80 and Ime1 targets]. Among those candidates is ATG14 which is an autophagy-specific subunit of the PI3K complex I involved in multiple forms of autophagy and the homolog of human BARKOR (Beclin 1-associated autophagy-related key regulator). BARKOR/ATG14 has two matches of Ndt80 consensus sequence, a significant p-value for NDT80 binding (0.01), a higher value of binding (1.07), is upregulated by Ndt80 overexpression (1.08), downregulated by NDT80 deletion (0.83), as well as upregulated by Ndt80 induction after 8h (1.16) and 14 h (1.3). Although the individual effects are relatively modest, together the all indicate a robust transcriptional induction by Ndt80.
As a promoter can harbour multiple binding sites we extended our approach by incorporating the number of motifs found in the promoter-proximal region. To our surprise, a few genes contain an exceptional high number of Nd80 consensus sequences. Most intriguing the highest scoring gene among the potential Ndt80 target genes encodes telomerase [Table: Motif number-corrected top-scoring Ndt80 target genes]. This gene encoding the catalytic component of the telomerase enzyme is also upregulated by DR by around two-fold [22912585]. It is the homolog of mammalian TERT. In human germ-line cells telomeres are long, whereas in cells of somatic tissues, telomerase activity is absent and the telomeres are short.
As ergosterol biosynthetic genes were proposed to be Ndt80 target genes [Sellam et al., 2009] and immortal cells have profound transcriptional changes in genes related to sterol biosyntheses, we checked the presence of genes with ERG or OSH in their gene symbols/alias. Several ergosterol-related genes appear to be directly or indirectly regulated by Ndt80 and some of these score as high as ATG14 [Table: Sterol-related genes, ATG14 and EST2 scoring for regulation by Ndt80]. For instance, ERG26 has one consensus sequence match, is significantly (p-value
Sum1 is the transcriptional antagonist of Ndt80 as it binds to similar sites and presumably competes with Ndt80. Sum1 directly binds to the promoter regions of OSH1 and OSH5 [Table: SUM1 regulation of OSH genes], both genes which when deleted extend the lifespan which is non-additive to DR (Xia et al., in preparation).
The approaches used in this study were devoted to identify the most promising target genes of rejuvenation transcription factors in order to unveil the mechanism of lifespan resetting. Surprisingly, we found that the process of rejuvenation (which originates from gametogenesis) is reused in a variety of lifespan-prolonging interventions, especially those manipulations presumed to work via diet responsive signalling (i.e. DR) and it appears to be the case of perhaps all DR-essential gene mutations that extend the lifespan.
Transient induction of Ndt80 resets replicative lifespan, but what is about chronological lifespan? We found that cells which activate NDT80 as well as the rejuvenation process do not age. They do not age even in the presence of excess nutrients (ad libitum, not DR). It appears due to the shut-down of nutrient sensing signalling (as it occurs under DR) but by cell cycle control because Rim15, Msn2/4 and meiotic transcription factors are highly activated. Therefore, it seems that the same process "rejuvenation" resets replicative lifespan and makes chronological cells immortal.
Another reason for the age-less phenotype of immobilized cells might be the environment itself. Usually cells expel stuff and sometimes detrimental components. For instance, acetic acid secreted by yeast cells shortens the lifespan [19305133]. In such it may be no wonder way immobilized cells appear to be so extraordinarily long-lived. Whether NDT80 upregulation in chronological cells might be causal for the age-less phenotype or not can be tested by moderate but continues overexpression of NDT80. An inducible promoter may allow tightly-regulated Ndt80 expression to an optimal level.
One of the most robust candidates is ATG14 as it was among the highly scored positively regulated target genes of Ndt80. ATG14 levels are elevated upon DR around 4-fold and non-ageing immobilized cells have also twice the amount of ATG14 transcript levels. Autophagy as well as sporulation is absent in ATG14 mutants, while ATG14 overexpression enhances autophagy. Further Osh1 accumulation is increased in ATG14 mutants.
Biogenesis of vacoules seems to be one target of Ndt80. While suppression of extrachromosomal rDNA circles, accumulation of Hsp104, and autophagy (Atg1) are not targets of Ndt80, chromatin remodeling might be one target. Many metals (cadmium, nickle, manganese) also cause chromatin remodelling, but they are toxic to cells. However, vacuole biogenesis may be one target of Ndt80 because of the following reasons:
During meiosis, the newly formed spores produce their own vacuoles. They do not inherit old vacuoles from the diploid cell [Shaw's result]. The transient expression of Ndt80 before spore formation should activate the vacuole biogenesis program (the exact mechanism of vacuole biogenesis is not known).
ime1-delta exhibits positive genetic interactions with ypt6-delta. Ypt6 helps the retrieval of Golgi components from late endosome (Multi-vesicular body). Vacuoles are probably formed from some form of late endosome. Assuming that Ime1-Ndt80 helps the formation of vacuoles, the positive genetic interaction between ime1-delta and ypt6-delta can be explained as that in the absence of Ime1-Ndt80, blocking the retromer from late endosome may help the formation of vacuoles.
the longevity effect of Perg-OSH6 (from 26 for wild-type to 37 for mutant) is similar to the expression of overexpression of Ndt80 in young cells (from 21 for control to 34 for Ndt80ox). Perg6-OSH6 probably works on late-endosome-vacuole transition.
The inference of transcription factor target genes as described here can be further improved by incorporating weighted matrices rather than sequence motifs, machine learning, and additionally considering the degree of sequence conservation.
Ndt80 binds to the consensus sequence gNCRCAAAY (N = any base, Y represents pyrimidine [C,T] and R indicates any purine [A,G]) of meiotic middle genes (MSE). Using position specific weight matrix instead of consensus sequence is superior.
Supervised learning approach for site identification using support vector machines by combining different data types is more sensitive when specificity and positive predictive value are the same. Processing of classifier outputs can provide high quality prediction and biological insights into functions of a defined transcription factors [19003435].
Five sensu stricto Saccharomcyes strain can be aligned. S. cerevisae closest relative is S. paradoxus, while S. bayanus is distant. Important cis-regulatory motifs in promoters regions are conserved throughout evolution, unlike other intergenic regions of DNA [11435399; 12775844]. However, the lack of conservation may not discriminate against a transcription factors regulatory role at a specific promoter. For instance, Sum1 and Sok2, regulate IME1, bind to non-perfect consensuses with non-conserved regions in sensu stricto Saccharomyces strains [20739709].
For each gene the average of all the probes belonging to the same condition were calculated, appropriate contrasts between conditions were made and signatures derived. Instead of averaging gene expression across replicates, the fold-change of each pair of comparison can be calculated. The ratios can then be averaged to get the mean fold-change of each probe-id [17971858]. For binding data only those genes which reached a significance of
800 bp upstream sequences of transcription start site were retrieved from Ensembl. These promoter proximal regions were searched with regular expressions for the presence of a motif. 2569 genes contained the Ndt80 core consensus sequence (CRCAAA), while 331 genes contained the consensus sequence (gNCRCAAAY) which is approximately the number of sites identified via direct binding measurement.
Functional enriched terms for a set of genes was evaluated with the Benjamini p-value. Intersection between sets was performed the with the hypergeometric test.
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Category Term Count % Benjamini GOTERM_BP_FAT response to temperature stimulus 49 8 3.90E-05 GOTERM_BP_FAT vacuolar protein catabolic process 32 5 8.60E-05 GOTERM_BP_FAT cellular response to heat 42 7 9.80E-05 GOTERM_BP_FAT response to heat 44 7 1.30E-04 GOTERM_BP_FAT response to abiotic stimulus 64 10 1.40E-04 GOTERM_BP_FAT sporulation 47 7 1.40E-04 GOTERM_BP_FAT sporulation resulting in formation of a cellular spore 47 7 1.40E-04 GOTERM_BP_FAT autophagy 34 5 4.60E-04 GOTERM_BP_FAT cellular lipid catabolic process 12 2 1.00E-03 GOTERM_CC_FAT peroxisomal part 14 2 1.80E-03 GOTERM_CC_FAT microbody part 14 2 1.80E-03 GOTERM_CC_FAT intrinsic to membrane 164 25 2.20E-03 GOTERM_CC_FAT peroxisome 19 3 2.30E-03 GOTERM_CC_FAT microbody 19 3 2.30E-03 GOTERM_CC_FAT integral to membrane 158 25 2.40E-03 GOTERM_CC_FAT peroxisomal matrix 8 1 2.40E-03 GOTERM_CC_FAT microbody lumen 8 1 2.40E-03 GOTERM_BP_FAT fatty acid catabolic process 8 1 3.10E-03 UP_SEQ_FEATURE short sequence motif:Microbody targeting signal 10 2 9.20E-03 GOTERM_BP_FAT fatty acid beta-oxidation 7 1 1.80E-02 KEGG_PATHWAY Citrate cycle (TCA cycle) 10 2 1.90E-02 GOTERM_BP_FAT carboxylic acid catabolic process 16 3 2.00E-02 GOTERM_BP_FAT organic acid catabolic process 16 3 2.00E-02 SP_PIR_KEYWORDS fatty acid metabolism 9 1 2.30E-02 SP_PIR_KEYWORDS peroxisome 17 3 2.40E-02 GOTERM_BP_FAT regulation of gluconeogenesis 8 1 2.70E-02 GOTERM_BP_FAT fatty acid oxidation 7 1 2.80E-02 GOTERM_BP_FAT lipid oxidation 7 1 2.80E-02 GOTERM_BP_FAT lipid catabolic process 13 2 2.80E-02 GOTERM_BP_FAT regulation of cellular ketone metabolic process 9 1 3.30E-02
All values in the table represent expression ratios, except those for Ndt80 B and Ime1 B which are p-values (B = Binding; OE = Over-Expression; I = Induction). Symbol Score Matches Ndt80 B Ndt80 B 9h NDT80 OE ndt80 D NDT80 I 8h NDT80 I 14h Ime1 B IME1 OE SWH1 6 0.31 1.64 1.04 0.94 0.81 0.98 0.3 1.01 OSH2 7 GCCAGAAAT GACACGAAT 0.34 0.96 1.03 0.81 0.83 1.55 0.21 0.99 OSH3 4 0.33 0.93 0.96 1 0.95 0.43 KES1 7 GAAACAAAT 0.4 0.93 1.07 1.27 1.05 1.16 0.55 1.02 HES1 5 GACGCGAAC 0.63 0.8 0.87 1.07 1.29 2.23 0.83 0.89 OSH6 7 GACGTAAAC GACACAGAT GGCACAACC 0.72 1.27 0.99 0.91 0.87 1.6 0.99 OSH7 8 CTCGCAAAT 0.81 2.26 1.02 0.89 1.13 1.99 0.18 1 ERG1 5 TCCACAAAC 0.74 0.81 0.97 1.24 2.95 1.14 0.21 0.93 ERG2 6 0.73 1.02 1.01 1.27 1.98 0.56 0.23 1.03 ERG3 6 1 0.9 1.05 1.55 5.51 1.15 0.3 1.06 ERG4 5 0.79 1.02 1.01 1.21 0.73 0.76 0.69 1.03 ERG5 4 0.9 0.89 0.97 1.34 5.63 1.01 0.71 0.98 ERG6 6 GGCCCAAAT 0.76 1.09 0.99 1.22 1.72 0.5 0.3 1.05 ERG7 6 0.21 0.86 1.03 0.98 1 0.93 0.39 1.04 ERG8 4 GAAACAAAC 0.47 0.91 0.99 1.17 1.6 0.51 0.6 0.97 ERG9 7 GTCACATAT 0.59 1.5 0.98 1.13 1.04 1.3 0.11 1.06 ERG10 4 GTCACAAAG 0.26 0.77 0.99 1.2 1.88 0.9 0.21 0.98 ERG11 5 GAGACAAAC 0.68 0.91 0.9 2.58 0.88 0.46 ERG12 6 GGCAAAAAT 0.64 0.74 1.04 1.19 1.29 0.75 0.24 1.05 ERG13 5 GTCGAAAAT 0.77 0.74 0.98 1.37 1.58 0.72 0.18 1.05 ERG20 8 GTGACAAAT 0.58 1.47 1.04 1.04 1.99 1.14 0.71 1.06 ERG24 7 TTCACAAAT 0.82 0.82 1.03 0.87 0.98 1.26 0.65 1.02 ERG25 6 GGCACAAAA 0.95 1.07 1.04 1.05 0.83 0.71 0.74 1.04 ERG26 5 TACGCAAAT 0.02 1.58 0.99 1 1.56 0.61 0.98 ERG27 6 ATCGCAAAT GCCAAAAAC GACACAATC 0.61 0.8 1 1.21 1.26 0.57 0.22 0.98 ERG28 5 GACGTAAAC 0.12 0.94 1.08 1 1.2 0.83 1.06 ATG14 8 GCCGAAAAT GACACTAAT 0.01 1.07 1.08 0.83 1.16 1.3 0.97 EST2 17 GGCGCAAAC GGCGCAAAC GGCGCAAAC GGCGCAAAC GGCGCAAAC GGCGCAAAC GGCGCAAAC GGCGCAAAC GGCGCAAAC 0.85 1.15 1.07 0.91 1.06 1.13 0.23 1.09
Genes that gained the highest score to be potential Ndt80 targets. Symbol Name Description YDL109C Putative lipase; involved in lipid metabolism; YDL109C is not an essential gene YDR249C Putative protein of unknown function YDR333C Putative protein of unknown function ATG14 AuTophaGy related Autophagy-specific subunit of phosphatidylinositol 3-kinase complex I (with Vps34/1tp5/30p); Atg14p targets complex I to the phagophore assembly site (PAS); required for localizing additional ATG proteins to the PAS; homolog of human Barkor DOC1 Destruction Of Cyclin B Processivity factor required for the ubiquitination activity of the anaphase promoting complex (APC), mediates the activity of the APC by contributing to substrate recognition; involved in cyclin proteolysis; contains a conserved DOC1 homology domain ALG13 Asparagine-Linked Glycosylation Catalytic component of UDP-GlcNAc transferase, required for the second step of dolichyl-linked oligosaccharide synthesis; anchored to the ER membrane via interaction with Alg14p; similar to bacterial and human glycosyltransferases PRP21 Pre-mRNA Processing Subunit of the SF3a splicing factor complex, required for spliceosome assembly TPP1 Three Prime Phosphatase DNA 3'-phosphatase that functions in repair of endogenous damage of double-stranded DNA, activity is specific for removal of 3' phosphates at strand breaks; has similarity to the l-2-haloacid dehalogenase superfamily YMR258C Repressor Of Ypt52 GTPase inhibitor with similarity to F-box proteins; inhibits Ypt52p GTPase activity by preventing Ypt52p from binding GTP; involved in regulating intracellular trafficking; physically interacts with Skp1p CNM67 Chaotic Nuclear Migration Component of the spindle pole body outer plaque; required for spindle orientation and mitotic nuclear migration SPO11 SPOrulation Meiosis-specific protein that initiates meiotic recombination by catalyzing the formation of double-strand breaks in DNA via a transesterification reaction; required for homologous chromosome pairing and synaptonemal complex formation YHR003C Protein of unknown function, localized to the mitochondrial outer membrane IPI1 Involved in Processing ITS2 Essential component of the Rix1 complex (with Rix1p and Ipi3p) that is required for processing of ITS2 sequences from 35S pre-rRNA; Rix1 complex associates with Mdn1p in pre-60S ribosomal particles YEL020C Hypothetical protein with low sequence identity to Pdc1p NUP157 NUclear Pore Abundant subunit of the nuclear pore complex (NPC), present on both sides of the NPC; has similarity to Nup170p; essential role, with Nup170p, in NPC assembly BUD5 BUD site selection GTP/GDP exchange factor for Rsr1p (Bud1p) required for both axial and bipolar budding patterns; mutants exhibit random budding in all cell types YLL032C Protein of unknown function that may interact with ribosomes, based on co-purification experiments; green fluorescent protein (GFP)-fusion protein localizes to the cytoplasm; YLL032C is not an essential gene CLB4 CycLin B B-type cyclin involved in cell cycle progression; activates Cdc28p to promote the G2/M transition; may be involved in DNA replication and spindle assembly; accumulates during S phase and G2, then targeted for ubiquitin-mediated degradation YGL242C Putative protein of unknown function; deletion mutant is viable
Genes that gained the highest score to be targeted by both Ndt80 and Ime1. Symbol Name Description YDR249C Putative protein of unknown function YDR333C Putative protein of unknown function ATG14 AuTophaGy related Autophagy-specific subunit of phosphatidylinositol 3-kinase complex I (with Vps34/15/30p); Atg14p targets complex I to the phagophore assembly site (PAS); required for localizing additional ATG proteins to the PAS; homolog of human Barkor DOC1 Destruction Of Cyclin B Processivity factor required for the ubiquitination activity of the anaphase promoting complex (APC), mediates the activity of the APC by contributing to substrate recognition; involved in cyclin proteolysis; contains a conserved DOC1 homology domain ALG13 Asparagine-Linked Glycosylation Catalytic component of UDP-GlcNAc transferase, required for the second step of dolichyl-linked oligosaccharide synthesis; anchored to the ER membrane via interaction with Alg14p; similar to bacterial and human glycosyltransferases IPI1 Involved in Processing ITS2 Essential component of the Rix1 complex (with Rix1p and Ipi3p) that is required for processing of ITS2 sequences from 35S pre-rRNA; Rix1 complex associates with Mdn1p in pre-60S ribosomal particles YEL020C Hypothetical protein with low sequence identity to Pdc1p NUP157 NUclear Pore Abundant subunit of the nuclear pore complex (NPC), present on both sides of the NPC; has similarity to Nup170p; essential role, with Nup170p, in NPC assembly BUD5 BUD site selection GTP/GDP exchange factor for Rsr1p (Bud1p) required for both axial and bipolar budding patterns; mutants exhibit random budding in all cell types YLL032C Protein of unknown function that may interact with ribosomes, based on co-purification experiments; green fluorescent protein (GFP)-fusion protein localizes to the cytoplasm; YLL032C is not an essential gene CLB4 CycLin B B-type cyclin involved in cell cycle progression; activates Cdc28p to promote the G2/M transition; may be involved in DNA replication and spindle assembly; accumulates during S phase and G2, then targeted for ubiquitin-mediated degradation
Genes that gained the highest score after a correction which takes into account the number of motifs. Symbol Name Description EST2 Ever Shorter Telomeres Reverse transcriptase subunit of the telomerase holoenzyme, essential for telomerase core catalytic activity, involved in other aspects of telomerase assembly and function; mutations in human homolog are associated with aplastic anemia KEI1 Kex2-cleavable protein Essential for Inositol phosphorylceramide synthesis Component of inositol phosphorylceramide (IPC) synthase; forms a complex with Aur1p and regulates its activity; required for IPC synthase complex localization to the Golgi; post-translationally processed by Kex2p; KEI1 is an essential gene STP1 Species-specific tRNA Processing Transcription factor, undergoes proteolytic processing by SPS (Ssy1p-Ptr3p-Ssy5p)-sensor component Ssy5p in response to extracellular amino acids; activates transcription of amino acid permease genes and may have a role in tRNA processing YNL018C Putative protein of unknown function NRM1 Negative Regulator of MBF targets Transcriptional co-repressor of MBF (MCB binding factor)-regulated gene expression; Nrm1p associates stably with promoters via MBF to repress transcription upon exit from G1 phase YLL032C Protein of unknown function that may interact with ribosomes, based on co-purification experiments; green fluorescent protein (GFP)-fusion protein localizes to the cytoplasm; YLL032C is not an essential gene VPH2 Vacuolar pH Integral membrane protein required for vacuolar H+-ATPase (V-ATPase) function, although not an actual component of the V-ATPase complex; functions in the assembly of the V-ATPase; localized to the endoplasmic reticulum (ER) KGD2 alpha-KetoGlutarate Dehydrogenase B Dihydrolipoyl transsuccinylase, component of the mitochondrial alpha-ketoglutarate dehydrogenase complex, which catalyzes the oxidative decarboxylation of alpha-ketoglutarate to succinyl-CoA in the TCA cycle; phosphorylated
Genes that are potentially regulated by Sum1, the antagonist of Ndt80. Symbol Score Matches genotype SUM1 Deletion 17h profile SUM1 ChIP-chip genotype SUM1 Deletion 10h profile Ndt80 & Sum1 genotype SUM1 Deletion 21h profile SWH1 4 GCGACTGTAAT 0.04 0.01 1.36 present 1.09 OSH2 2 GCGACACGAAT 0.03 0.73 1.32 1.05 OSH3 2 0.0 0.67 1.15 0.81 KES1 1 0.05 0.67 1.11 1.3 HES1 3 0.02 0.04 0.96 1.24 OSH6 1 0.01 0.45 1.28 1.24 OSH7 2 0.05 0.59 1.5 0.98
