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Abstract Background The largest sub-unit of RNA polymerase II, Rpb1p, has long been known to be subject to post-translational modifications that influence various aspects of pre-mRNA processing. However, the portion of the Rpb1p molecule subject to these modifications – the carboxy-terminal domain or CTD – remains the subject of much fascination. Intriguingly, the CTD possesses a unique repetitive structure consisting of multiple repeats of the heptapeptide sequence, Y1S2P3T4S5P6S7. While these repeats are critical for viability, they are not required for basal transcriptional activity in vitro. This suggests that – even though the CTD is not catalytically essential – it must perform other critical functions in eukaryotes. Presentation of the Hypothesis By formally applying the long-standing mathematical principles of information theory, I explore the hypothesis that complex post-translational modifications of the CTD represent a means for the dynamic “programming” of Rpb1p and thus for the discrete modulation of the expression of specific gene subsets in eukaryotes. Testing the Hypothesis Empirical means for testing the informational capacity and regulatory potential of the CTD – based on simple genetic analysis in yeast model systems – are put forward and discussed. Implications of the Hypothesis These ideas imply that the controlled manipulation of CTD effectors could be used to “program” the CTD and thus to manipulate biological processes in eukaryotes in a definable manner.

Abstract Background In Schizosaccharomyces pombe the SET domain protein, Set3p - together with its interacting partners, Snt1p, and Hif2p - form a complex that aids in preventing cell division failure upon mild cytokinetic stress. Intriguingly, the human orthologs of these proteins (MLL5, NCOR2, and TBL1X) are also important for the faithful completion of cytokinesis in tissue culture cells. Since MLL5, NCOR2, and TBL1X form a complex with the histone deacetylase, HDAC3, we sought to determine if an orthologous counterpart played a regulatory role in fission yeast cytokinesis. Results In this report we identify the hos2 gene as the fission yeast HDAC3 ortholog. We show that Hos2p physically interacts with Set3p, Snt1p, and Hif2p, and that hos2∆ mutants are indeed compromised in their ability to reliably complete cell division in the presence of mild cytokinetic stresses. Furthermore, we demonstrate that over-expression of hos2 causes severe morphological and cytokinetic defects. Lastly, through recombinase mediated cassette exchange, we show that expression of human HDAC3 complements the cytokinetic defects exhibited by hos2∆ cells. Conclusions These data support a model in which Hos2p functions as an essential component of the Set3p-Snt1p-Hif2p complex with respect to the regulation of cytokinesis. The ability of human HDAC3 to complement the cytokinesis defects associated with the deletion of the hos2 gene suggests that further analysis of this system could provide insight into the role of HDAC3 in both the regulation of cell division, as well as other biological processes influenced by HDAC3 deacetylation.

Histone deacetylase 2 (HD2) is a unique family of histone deacetylases (HDACs) in plants. Despite evidence that certain HD2 family HDACs play an important role in plant growth and stress response, the coordination of HD2s in these processes remains largely unknown. We found that HD2-type, HD2A and HD2C coordinate to play a role in drought stress response in Arabidopsis. We showed that the hd2a . hd2c double mutant (Mac16) exhibit decreased drought survival and increased water loss as compared to the single mutants, hd2a and hd2c . Gene expression analysis showed that the ABI1 and ABI2 genes were upregulated and SLAC1 was downregulated which led to the modified stomatal functioning in the Mac16 as compared to the single mutants. Overexpression of HD2A and HD2C showed enhanced drought survival and decreased water loss. We also showed that the GA2ox1 and GA2ox2 genes, which are involved in the catabolism of bioactive gibberellic acids, were upregulated in the Mac16 as compared to the single mutants, which led to a decreased root growth in the Mac16. Furthermore, we showed that HD2A and HD2C can physically interact and increased genome-wide H3K9 acetylation was observed in the Mac16, compared to the single mutants. Overall, our investigation revealed that HD2A and HD2C coordinate to play a cumulative role in drought stress response and root growth in Arabidopsis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Tahir, Karagiannis and Tian.)

Extreme environmental conditions, such as drought, are expected to increase in frequency and severity due to climate change, leading to substantial deficiencies in crop yield and quality. Medicago sativa (alfalfa) is an important crop that is relied upon as a staple source of forage in ruminant feed. Despite its economic importance, alfalfa production is constrained by abiotic stress, including drought. In this report, we investigate the role of Squamosa Promoter Binding Protein-Like 9 ( SPL9 ), a target of miR156, in drought tolerance. Transgenic alfalfa plants with RNAi-silenced MsSPL9 ( SPL9 -RNAi) were compared to wild-type (WT) alfalfa for phenotypic changes and drought tolerance indicators. In SPL9 -RNAi plants, both stem thickness and plant height were reduced in two- and six-month-old alfalfa, respectively; however, yield was unaffected. SPL9 -RNAi plants showed less leaf senescence and had augmented relative water content under drought conditions, indicating that SPL9 -RNAi plants had greater drought tolerance potential than WT plants. Interestingly, SPL9 -RNAi plants accumulated more stress-alleviating anthocyanin compared to WT under both drought and well-watered control conditions, suggesting that MsSPL9 may contribute to drought tolerance in alfalfa, at least in part, by regulating anthocyanin biosynthesis. The results suggest that targeting MsSPL9 is a suitable means for improving alfalfa resilience towards drought conditions.

Post-translational glycosylation of proteins with O-linked β-N-acetylglucosamine (O-GlcNAcylation) and changes of galectin expression profiles are essential in many cellular stress responses. We examine this regulation in the liver tissue of hibernating thirteen-lined ground squirrels (Ictidomys tridecemlineatus) representing a biological model of hypometabolism and physiological stress resistance. The tissue levels of O-GlcNAcylated proteins as well as galectin-1 and galectin-3 proteins detected by immunodot blot assay were significantly lower by 4.6-5.4-, 2.2-2.3- and 2.5-2.9-fold, respectively, in the non-hibernating summer squirrels compared with those in winter, whether hibernating or aroused. However, there were no differences in the expression of genes encoding enzymes involved in O-GlcNAc cycle (O-GlcNAc transferase and O-GlcNAcase) and such galectins as LGALS1, LGALS2, LGALS3, LGALS4 and LGALS9. Only the expression of LGALS8 gene in the liver tissue was significantly decreased by 37.6 ± 0.1% in hibernating ground squirrels relative to summer animals. Considering that the expression of a proven genetic biomarker ELOVL6 encoding ELOVL fatty acid elongase 6 was readily upregulated in non-hibernating animals by 11.3-32.9-fold, marginal differential changes in the expression of galectin genes cannot be classified as biomarkers of hibernation. Thus, this study provides evidence that hibernation in Ictidomys tridecemlineatus is associated with increasing O-GlcNAcylation of liver proteins and suggests that the contribution of galectins deserves further studies at the protein level.

Background/aim: The effects of O-linked β-N-acetyl-D-glucosamine (O-GlcNAc) transferase (OGT) and O-GlcNAcase (OGA) inhibitors on galectin gene expression profiles were examined in MCF7, HT-29, and HL-60 cancer cell lines., Materials and Methods: Cell cultures were treated for 24 h with OGA inhibitor thiamet G or OGT inhibitor 2-acetamido-1,3,4,6-tetra-O-acetyl-2-deoxy-5-thio-α-D-glucopyranose, and global O-GlcNAc levels and expression of galectin genes were determined using an immunodot blot assay and real-time quantitative polymerase chain reaction., Results: Two galectin genes, LGALS3 in MCF7 cells and LGALS12 in HL-60 cells, were up-regulated by O-GlcNAc, whereas other cell-specific galectins were unresponsive to changes in O-GlcNAc level. Of interest, basal levels of O-GlcNAc in resting HL-60 and HT-29 cells were significantly higher than those in cells differentiated into neutrophilic or enterocytic lineages, respectively., Conclusion: O-GlcNAc-mediated signaling pathways may be involved in regulating the expression of only a limited number of galectin genes. Additional O-GlcNAc-dependent mechanisms may work at the protein level (galectin secretion and intracellular localization) and warrant further investigation., (Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Tra1 is an essential component of the SAGA/SLIK and NuA4 complexes in S. cerevisiae , recruiting these co-activator complexes to specific promoters. As a PIKK family member, Tra1 is characterized by a C-terminal phosphoinositide 3-kinase (PI3K) domain. Unlike other PIKK family members ( e.g. , Tor1, Tor2, Mec1, Tel1), Tra1 has no demonstrable kinase activity. We identified three conserved arginine residues in Tra1 that reside proximal or within the cleft between the N- and C-terminal subdomains of the PI3K domain. To establish a function for Tra1's PI3K domain and specifically the cleft region, we characterized a tra1 allele where these three arginine residues are mutated to glutamine. The half-life of the Tra1[Formula: see text] protein is reduced but its steady state level is maintained at near wild-type levels by a transcriptional feedback mechanism. The tra1 [Formula: see text] allele results in slow growth under stress and alters the expression of genes also regulated by other components of the SAGA complex. Tra1[Formula: see text] is less efficiently transported to the nucleus than the wild-type protein. Likely related to this, Tra1[Formula: see text] associates poorly with SAGA/SLIK and NuA4. The ratio of Spt7 SLIK to Spt7 SAGA increases in the tra1 [Formula: see text] strain and truncated forms of Spt20 become apparent upon isolation of SAGA/SLIK. Intragenic suppressor mutations of tra1 [Formula: see text] map to the cleft region further emphasizing its importance. We propose that the PI3K domain of Tra1 is directly or indirectly important for incorporating Tra1 into SAGA and NuA4 and thus the biosynthesis and/or stability of the intact complexes., (Copyright © 2018 Berg et al.)

Tuberous sclerosis complex is an autosomal dominant disorder characterized by benign tumors arising from the abnormal activation of mTOR signaling in cells lacking TSC1 (hamartin) or TSC2 (tuberin) activity. To expand the genetic framework surrounding this group of growth regulators, we utilized the model eukaryote Schizosaccharomyces pombe to uncover and characterize genes that buffer the phenotypic effects of mutations in the orthologous tsc1 or tsc2 loci. Our study identified two genes: fft3 (encoding a DNA helicase) and ypa1 (encoding a peptidyle-prolyl cis/trans isomerase). While the deletion of fft3 or ypa1 has little effect in wild-type fission yeast cells, their loss in tsc1Δ or tsc2Δ backgrounds results in severe growth inhibition. These data suggest that the inhibition of Ypa1p or Fft3p might represent an 'Achilles' heel' of cells defective in hamartin/tuberin function. Furthermore, we demonstrate that the interaction between tsc1 / tsc2 and ypa1 can be rescued through treatment with the mTOR inhibitor, torin-1, and that ypa1Δ cells are resistant to the glycolytic inhibitor, 2-deoxyglucose. This identifies ypa1 as a novel upstream regulator of mTOR and suggests that the effects of ypa1 loss, together with mTOR activation, combine to result in a cellular maladaptation in energy metabolism that is profoundly inhibitory to growth., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2018. Published by The Company of Biologists Ltd.)

As part of an earlier study aimed at uncovering gene products with roles in defending against latrunculin A (LatA)-induced cytoskeletal perturbations, we identified three members of the oxidative stress response pathway: the Pap1p AP-1-like transcription factor, the Imp1p α-importin, and the Caf5p efflux pump. In this report, we characterize the pathway further and show that Pap1p translocates from the cytoplasm to the nucleus in an Imp1p-dependent manner upon LatA treatment. Moreover, preventing this translocation, through the addition of a nuclear export signal (NES), confers the same characteristic LatA-sensitive phenotype exhibited by pap1 Δ cells. Lastly, we show that the caf5 gene is induced upon exposure to LatA and that Pap1p is required for this transcriptional upregulation. Importantly, the expression of trr1 , a Pap1p target specifically induced in response to oxidative stress, is not significantly altered by LatA treatment. Taken together, these results suggest a model in which LatA-mediated cytoskeletal perturbations are sensed, triggering the Imp1p-dependent translocation of Pap1p to the nucleus and the induction of the caf5 gene (independently of oxidative stress)., (Copyright © 2017 Asadi et al.)

The heparin-binding protein midkine is a potent growth factor with emerging roles in numerous inflammatory diseases. Beyond its characterization in embryogenesis and organ development, ample insights into its function have been collected from experimental disease models using knockout animals or knockdown intervention strategies. Here a comprehensive overview on midkine and its functions in atherogenesis and kidney diseases is provided. Molecular clues to key signalling pathways (Akt, ERK, HIF1α) and key events in atherosclerotic vessels link midkine expression with vascular smooth muscle proliferation and (neo)angiogenesis. In acute and chronic kidney diseases, midkine expression is upregulated in tubular as well as endothelial cells. Experimental disease models that mimic diabetic nephropathy and/or immunologic glomerular damage indicate dichotomous midkine activities, with cytoprotective as well as injurious effects. This review also pinpoints the commonalities of the disease models. An understanding of the underlying molecular events will be required in order to design a targeted intervention into cardiovascular or renal diseases as well as inflammatory processes., (© The Author 2016. Published by Oxford University Press on behalf of ERA-EDTA. All rights reserved.)

Fission yeast cells treated with low doses of the actin depolymerizing drug, latrunculin A (LatA), delay entry into mitosis via a mechanism that is dependent on both the Clp1p and Rad24p proteins. During this delay, cells remain in a cytokinesis-competent state that is characterized by continuous repair and/or reestablishment of the actomyosin ring. In this manner, cells ensure the faithful completion of the preceding cytokinesis in response to perturbation of the cell division machinery. To uncover other genes with a role in this response, or simply genes with roles in adapting to LatA-induced stress, we carried out a genome-wide screen and identified a group of 38 gene deletion mutants that are hyper-sensitive to the drug. As expected, we found genes affecting cytokinesis and/or the actin cytoskeleton within this set (ain1, acp2, imp2). We also identified genes with roles in histone modification (tra1, ngg1), intracellular transport (apl5, aps3), and glucose-mediated signaling (git3, git5, git11, pka1, cgs2). Importantly, while the identified gene deletion mutants are prone to cytokinesis failure in the presence of LatA, they are nevertheless fully capable of cell division in the absence of the drug. These results indicate that fission yeast cells make use of a diverse set of regulatory modules to counter abnormal cytoskeletal perturbations, and furthermore, that these modules act redundantly to ensure cell survival and proliferation., (Copyright © 2016 Asadi et al.)

The TTT complex is composed of the three essential proteins Tel2, Tti1, and Tti2 The complex is required to maintain steady state levels of phosphatidylinositol 3-kinase-related kinase (PIKK) proteins, including mTOR, ATM/Tel1, ATR/Mec1, and TRRAP/Tra1, all of which serve as regulators of critical cell signaling pathways. Due to their association with heat shock proteins, and with newly synthesized PIKK peptides, components of the TTT complex may act as cochaperones. Here, we analyze the consequences of depleting the cellular level of Tti2 in Saccharomyces cerevisiae We show that yeast expressing low levels of Tti2 are viable under optimal growth conditions, but the cells are sensitive to a number of stress conditions that involve PIKK pathways. In agreement with this, depleting Tti2 levels decreased expression of Tra1, Mec1, and Tor1, affected their localization and inhibited the stress responses in which these molecules are involved. Tti2 expression was not increased during heat shock, implying that it does not play a general role in the heat shock response. However, steady state levels of Hsp42 increase when Tti2 is depleted, and tti2L187P has a synthetic interaction with exon 1 of the human Huntingtin gene containing a 103 residue polyQ sequence, suggesting a general role in protein quality control. We also find that overexpressing Hsp90 or its cochaperones is synthetic lethal when Tti2 is depleted, an effect possibly due to imbalanced stoichiometry of a complex required for PIKK assembly. These results indicate that Tti2 does not act as a general chaperone, but may have a specialized function in PIKK folding and/or complex assembly., (Copyright © 2016 Hoffman et al.)