A crucial RNA-binding lysine residue in the Nab3 RRM domain undergoes SET1 and SET3-responsive methylation.

The Nrd1-Nab3-Sen1 (NNS) complex integrates molecular cues to direct termination of noncoding transcription in budding yeast. NNS is positively regulated by histone methylation as well as through Nrd1 binding to the initiating form of RNA PolII. These cues collaborate with Nrd1 and Nab3 binding to target RNA sequences in nascent transcripts through their RRM RNA recognition motifs. In this study, we identify nine lysine residues distributed amongst Nrd1, Nab3 and Sen1 that are methylated, suggesting novel molecular inputs for NNS regulation. We identify mono-methylation of one these residues (Nab3-K363me1) as being partly dependent on the H3K4 methyltransferase, Set1, a known regulator of NNS function. Moreover, the accumulation of Nab3-K363me1 is essentially abolished in strains lacking SET3, a SET domain containing protein that is positively regulated by H3K4 methylation. Nab3-K363 resides within its RRM and physically contacts target RNA. Mutation of Nab3-K363 to arginine (Nab3-K363R) decreases RNA binding of the Nab3 RRM in vitro and causes transcription termination defects and slow growth. These findings identify SET3 as a potential contextual regulator of Nab3 function through its role in methylation of Nab3-K363. Consistent with this hypothesis, we report that SET3 exhibits genetic activation of NAB3 that is observed in a sensitized context.

Combinatorial Genetic Control of Rpd3S Through Histone H3K4 and H3K36 Methylation in Budding Yeast.

Much of euchromatin regulation occurs through reversible methylation of histone H3 lysine-4 and lysine-36 (H3K4me and H3K36me). Using the budding yeast Saccharomyces cerevisiae, we previously found that levels of H3K4me modulated temperature sensitive alleles of the transcriptional elongation complex Spt6-Spn1 through an unknown H3K4me effector pathway. Here we identify the Rpd3S histone deacetylase complex as the H3K4me effector underlying these Spt6-Spn1 genetic interactions. Exploiting these Spt6-Spn1 genetic interactions, we show that H3K4me and H3K36me collaboratively impact Rpd3S function in an opposing manner. H3K36me is deposited by the histone methyltransferase Set2 and is known to promote Rpd3S function at RNA PolII transcribed open reading frames. Using genetic epistasis experiments, we find that mutations perturbing the Set2-H3K36me-Rpd3S pathway suppress the growth defects caused by temperature sensitive alleles of SPT6 and SPN1, illuminating that this pathway antagonizes Spt6-Spn1 Using these sensitive genetic assays, we also identify a role for H3K4me in antagonizing Rpd3S that functions through the Rpd3S subunit Rco1, which is known to bind H3 N-terminal tails in a manner that is prevented by H3K4me. Further genetic experiments reveal that the H3K4 and H3K36 demethylases JHD2 and RPH1 mediate this combinatorial control of Rpd3S. Finally, our studies also show that the Rpd3L complex, which acts at promoter-proximal regions of PolII transcribed genes, counters Rpd3S for genetic modulation of Spt6-Spn1, and that these two Rpd3 complexes balance the activities of each other. Our findings present the first evidence that H3K4me and H3K36me act combinatorially to control Rpd3S.

H3K4 Methylation Dependent and Independent Chromatin Regulation by JHD2 and SET1 in Budding Yeast.

Set1 and Jhd2 regulate the methylation state of histone H3 lysine-4 (H3K4me) through their opposing methyltransferase and demethylase activities in the budding yeast Saccharomyces cerevisiae H3K4me associates with actively transcribed genes and, like both SET1 and JHD2 themselves, is known to regulate gene expression diversely. It remains unclear, however, if Set1 and Jhd2 act solely through H3K4me. Relevantly, Set1 methylates lysine residues in the kinetochore protein Dam1 while genetic studies of the S. pombe SET1 ortholog suggest the existence of non-H3K4 Set1 targets relevant to gene regulation. We interrogated genetic interactions of JHD2 and SET1 with essential genes involved in varied aspects of the transcription cycle. Our findings implicate JHD2 in genetic inhibition of the histone chaperone complexes Spt16-Pob3 (FACT) and Spt6-Spn1 This targeted screen also revealed that JHD2 inhibits the Nrd1-Nab3-Sen1 (NNS) transcription termination complex. We find that while Jhd2's impact on these transcription regulatory complexes likely acts via H3K4me, Set1 governs the roles of FACT and NNS through opposing H3K4-dependent and -independent functions. We also identify diametrically opposing consequences for mutation of H3K4 to alanine or arginine, illuminating that caution must be taken in interpreting histone mutation studies. Unlike FACT and NNS, detailed genetic studies suggest an H3K4me-centric mode of Spt6-Spn1 regulation by JHD2 and SET1 Chromatin immunoprecipitation and transcript quantification experiments show that Jhd2 opposes the positioning of a Spt6-deposited nucleosome near the transcription start site of SER3, a Spt6-Spn1 regulated gene, leading to hyper-induction of SER3 In addition to confirming and extending an emerging role for Jhd2 in the control of nucleosome occupancy near transcription start sites, our findings suggest some of the chromatin regulatory functions of Set1 are independent of H3K4 methylation.

Simultaneous nutrient removal, optimised CO2 mitigation and biofuel feedstock production by Chlorogonium sp. grown in secondary treated non-sterile saline sewage effluent.

The phycoremediation process has great potential for effectively addressing environmental pollution. To explore the capabilities of simultaneous algal nutrient removal, CO2 mitigation and biofuel feedstock production from spent water resources, a Chlorogonium sp. isolated from a tilapia pond in Hong Kong was grown in non-sterile saline sewage effluent for a bioremediation study. With high removal efficiencies of NH3-N (88.35±14.39%), NO3(-)-N (85.39±14.96%), TN (93.34±6.47%) and PO4(3-)-P (91.80±17.44%), Chlorogonium sp. achieved a CO2 consumption rate of 58.96 mg L(-1) d(-1), which was optimised by the response surface methodology. Under optimised conditions, the lipid content of the algal biomass reached 24.26±2.67%. Overall, the isolated Chlorogonium sp. showed promising potential in the simultaneous purification of saline sewage effluent in terms of tertiary treatment and CO2 sequestration while delivering feedstock for potential biofuel production in a waste-recycling manner.

Developmentally programmed nuclear destruction during yeast gametogenesis.

Autophagy controls cellular catabolism in diverse eukaryotes and modulates programmed cell death in plants and animals. While studies of the unicellular yeast Saccharomyces cerevisiae have provided fundamental insights into the mechanisms of autophagy, the roles of cell death pathways in yeast are less well understood. Here, we describe widespread developmentally programmed nuclear destruction (PND) events that occur during yeast gametogenesis. PND is executed through apoptotic-like DNA fragmentation in coordination with an unusual form of autophagy that is most similar to mammalian lysosomal membrane permeabilization and mega-autophagy, a form of plant autophagic cell death. Undomesticated strains execute gametogenic PND broadly in maturing colonies to the apparent benefit of sibling cells, confirming its prominence during the yeast life cycle. Our results reveal that diverse cell-death-related processes converge during gametogenesis in a microbe distantly related to plants or animals, highlighting gametogenesis as a process during which programmed cell death mechanisms may have evolved.