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Crotonylated lysine residues within histones are linked to transcriptional activation in a process involving histone mark ‘reader’ proteins. Crystallographic analysis of the YEATS domain of the Taf14 protein reveals a mode of crotonylated histone mark recognition via a π-sandwich motif. The discovery of new histone modifications is unfolding at startling rates; however, the identification of effectors capable of interpreting these modifications has lagged behind. Here we report the YEATS domain as an effective reader of histone lysine crotonylation, an epigenetic signature associated with active transcription. We show that the Taf14 YEATS domain engages crotonyllysine via a unique π–π–π-stacking mechanism and that other YEATS domains have crotonyllysine-binding activity.

In Arabidopsis , RNA-directed DNA methylation is a poorly understood gene silencing pathway in which small interfering RNAs generated by RNA polymerase IV (Pol-IV) target a DNA methyltransferase to its sites of action; here structural and genomic analyses demonstrate that SHH binds chromatin via repressive histone modifications and recruits Pol-IV to enable siRNA production. Polymerase IV targeting in DNA methylation In Arabidopsis , RNA-directed DNA methylation (RdDM) is a poorly understood gene-silencing pathway in which small interfering RNAs (siRNAs) generated by an RNA polymerase, Pol-IV, target a DNA methyltransferase to its sites of action. Here, Steven Jacobsen and colleagues report that the homeodomain protein SHH1 binds chromatin via repressive histone modifications and recruits Pol-IV to promote siRNA production. SHH1 has a novel chromatin-binding module that functions as a dual lysine reader for both K4 and K9 of the histone H3 tail. This work identifies an early targeting step that may be a critical control point in epigenetic modification through DNA methylation. DNA methylation is an epigenetic modification that has critical roles in gene silencing, development and genome integrity. In Arabidopsis , DNA methylation is established by DOMAINS REARRANGED METHYLTRANSFERASE 2 (DRM2) and targeted by 24-nucleotide small interfering RNAs (siRNAs) through a pathway termed RNA-directed DNA methylation (RdDM) 1 . This pathway requires two plant-specific RNA polymerases: Pol-IV, which functions to initiate siRNA biogenesis, and Pol-V, which functions to generate scaffold transcripts that recruit downstream RdDM factors 1 , 2 . To understand the mechanisms controlling Pol-IV targeting we investigated the function of SAWADEE HOMEODOMAIN HOMOLOG 1 (SHH1) 3 , 4 , a Pol-IV-interacting protein 3 . Here we show that SHH1 acts upstream in the RdDM pathway to enable siRNA production from a large subset of the most active RdDM targets, and that SHH1 is required for Pol-IV occupancy at these same loci. We also show that the SHH1 SAWADEE domain is a novel chromatin-binding module that adopts a unique tandem Tudor-like fold and functions as a dual lysine reader, probing for both unmethylated K4 and methylated K9 modifications on the histone 3 (H3) tail. Finally, we show that key residues within both lysine-binding pockets of SHH1 are required in vivo to maintain siRNA and DNA methylation levels as well as Pol-IV occupancy at RdDM targets, demonstrating a central role for methylated H3K9 binding in SHH1 function and providing the first insights into the mechanism of Pol-IV targeting. Given the parallels between methylation systems in plants and mammals 1 , 5 , a further understanding of this early targeting step may aid our ability to control the expression of endogenous and newly introduced genes, which has broad implications for agriculture and gene therapy.

Histone methyltransferase MLL4 is centrally involved in transcriptional regulation and is often mutated in human diseases, including cancer and developmental disorders. MLL4 contains a catalytic SET domain that mono-methylates histone H3K4 and seven PHD fingers of unclear function. Here, we identify the PHD6 finger of MLL4 (MLL4-PHD6) as a selective reader of the epigenetic modification H4K16ac. The solution NMR structure of MLL4-PHD6 in complex with a H4K16ac peptide along with binding and mutational analyses reveal unique mechanistic features underlying recognition of H4K16ac. Genomic studies show that one third of MLL4 chromatin binding sites overlap with H4K16ac-enriched regions in vivo and that MLL4 occupancy in a set of genomic targets depends on the acetyltransferase activity of MOF, a H4K16ac-specific acetyltransferase. The recognition of H4K16ac is conserved in the PHD7 finger of paralogous MLL3. Together, our findings reveal a previously uncharacterized acetyllysine reader and suggest that selective targeting of H4K16ac by MLL4 provides a direct functional link between MLL4, MOF and H4K16 acetylation. Histone methyltransferase MLL4 is a transcriptional regulator. Here the authors identify the PHD6 finger of MLL4 as a selective reader of the epigenetic modification H4K16ac and show that a subset of MLL4 chromatin binding sites overlap with H4K16ac-enriched regions, which depends on MOF activity.

The epigenetic inheritance of DNA methylation requires UHRF1, a histone- and DNA-binding RING E3 ubiquitin ligase that recruits DNMT1 to sites of newly replicated DNA through ubiquitylation of histone H3. UHRF1 binds DNA with selectivity towards hemi-methylated CpGs (HeDNA); however, the contribution of HeDNA sensing to UHRF1 function remains elusive. Here, we reveal that the interaction of UHRF1 with HeDNA is required for DNA methylation but is dispensable for chromatin interaction, which is governed by reciprocal positive cooperativity between the UHRF1 histone- and DNA-binding domains. HeDNA recognition activates UHRF1 ubiquitylation towards multiple lysines on the H3 tail adjacent to the UHRF1 histone-binding site. Collectively, our studies are the first demonstrations of a DNA-protein interaction and an epigenetic modification directly regulating E3 ubiquitin ligase activity. They also define an orchestrated epigenetic control mechanism involving modifications both to histones and DNA that facilitate UHRF1 chromatin targeting, H3 ubiquitylation, and DNA methylation inheritance. Cells are able to regulate the activity of their genes in response to different cues. Genetic information is encoded in DNA and one way to regulate gene activity is to modify the DNA by attaching chemical “epigenetic” markers to it. When a cell divides, these epigenetic markers can be inherited by the daughter cells so that they share the same patterns of gene activity as the parent cell. When the DNA of the parent cell is copied prior to cell division, the epigenetic markers are also copied onto the new DNA. Mistakes in this process are linked to a wide range of diseases in humans, such as cancer and neurological disorders. One type of epigenetic marker is known as a methyl tag and it is added to DNA by certain enzymes in a process called DNA methylation. A protein called UHRF1 is required for human cells to inherit patterns of DNA methylation through cell division. This protein binds to newly copied DNA that lacks some methyl tags as well as to another protein associated with DNA called histone H3. UHRF1 modifies histone H3 by attaching a small protein molecule called ubiquitin to it. This helps to recruit a DNA methylation enzyme to place methyl tags on the newly copied DNA. However, it was not clear how the various properties of UHRF1 allow it to control how DNA methylation is inherited. Harrison et al. addressed this question by studying purified proteins and DNA fragments outside of living cells. The results show that UHRF1 binding to DNA and histone H3 work together to bring UHRF1 to the sites on DNA that require methylation. Further experiments revealed that the methylation pattern on newly copied DNA is able to activate the ability of UHRF1 to place ubiquitin on histone H3. The findings of Harrison et al. reveal a new mechanism by which dividing cells control how DNA methylation is inherited by their daughter cells. A future challenge will be to find out how attaching ubiquitin to histone H3 activates DNA methylation.

Acetylation of histone H3K23 has emerged as an essential posttranslational modification associated with cancer and learning and memory impairment, yet our understanding of this epigenetic mark remains insufficient. Here, we identify the native MORF complex as a histone H3K23-specific acetyltransferase and elucidate its mechanism of action. The acetyltransferase function of the catalytic MORF subunit is positively regulated by the DPF domain of MORF (MORF DPF ). The crystal structure of MORF DPF in complex with crotonylated H3K14 peptide provides mechanistic insight into selectivity of this epigenetic reader and its ability to recognize both histone and DNA. ChIP data reveal the role of MORF DPF in MORF-dependent H3K23 acetylation of target genes. Mass spectrometry, biochemical and genomic analyses show co-existence of the H3K23ac and H3K14ac modifications in vitro and co-occupancy of the MORF complex, H3K23ac, and H3K14ac at specific loci in vivo. Our findings suggest a model in which interaction of MORF DPF with acylated H3K14 promotes acetylation of H3K23 by the native MORF complex to activate transcription. Acetylation of histone H3K23 has emerged as an essential posttranslational modification, yet this epigenetic mark remains poorly understood. Here, the authors identify the native MORF complex as a histone H3K23-specific acetyltransferase and show that interaction of the MORF subunit with acylated H3K14 promotes acetylation of H3K23 by this complex to activate transcription.

The clinical use of 1,25-dihydroxycholecalciferol (1,25D3), the active form of vitamin D3, is limited by its calcemic side effects and rapid metabolic degradation. To overcome these limitations, we designed novel vitamin D analogs with extended, rigidified, and branched side chains. Among them, PRI-1938, featuring a 5,6-trans triene system and 22,24-all-trans side-chain geometry, demonstrated markedly enhanced resistance to enzymatic catabolism. In vitro assays revealed that metabolic conversion of PRI-1938 by the nonselective cytochrome P450 3A4 (CYP3A4) enzyme was ca. 4-fold lower than that of the previously obtained PRI-1906 and over 9-fold lower than 1,25D3. All new analogs, including PRI-1927 and PRI-1937, exhibited significantly higher stability toward mitochondrial cytochrome P450 24A1 (CYP24A1), the vitamin D-selective catabolic enzyme, than that of 1,25D3. Molecular modeling and quantum mechanical calculations indicated that PRI-1938 adopts a highly stable conformation in the CYP24A1 active site, stabilized by four hydrogen bonds and multiple hydrophobic interactions. The spatially optimized interaction network reduces access to the catalytic heme, resulting in the lowest observed metabolic conversion. These findings highlight the critical role of the side-chain geometry in modulating metabolic stability and support the further development of PRI-1938 as a promising anticancer vitamin D analog.

Preliminary studies are presented showing to what extent nutrients available in the growth environment of Kombucha microorganisms affect the physical and mechanical properties of synthesized cellulose. With an increase in the amount of sucrose in the growth medium and with the presence of additional nutrients, peptone and tea extract, the thickness and strength of the biopolymer increased, while elongation was reduced. The best physical and mechanical parameters were obtained for bacterial cellulose from cultures with the addition of 10% sucrose and 0.25% peptone content. The increase in elongation correlated with the decrease in the degree of polymerization, which means that in media rich in nutrients, the number of molecules building the polymer decreases. The presented data is important in order to select ingredients that will help synthesize bacterial cellulose with the desired physio-mechanical properties.

Background Plant homeodomain (PHD) fingers are central “readers” of histone post-translational modifications (PTMs) with > 100 PHD finger-containing proteins encoded by the human genome. Many of the PHDs studied to date bind to unmodified or methylated states of histone H3 lysine 4 (H3K4). Additionally, many of these domains, and the proteins they are contained in, have crucial roles in the regulation of gene expression and cancer development. Despite this, the majority of PHD fingers have gone uncharacterized; thus, our understanding of how these domains contribute to chromatin biology remains incomplete. Results We expressed and screened 123 of the annotated human PHD fingers for their histone binding preferences using reader domain microarrays. A subset (31) of these domains showed strong preference for the H3 N-terminal tail either unmodified or methylated at H3K4. These H3 readers were further characterized by histone peptide microarrays and/or AlphaScreen to comprehensively define their H3 preferences and PTM cross-talk. Conclusions The high-throughput approaches utilized in this study establish a compendium of binding information for the PHD reader family with regard to how they engage histone PTMs and uncover several novel reader domain–histone PTM interactions (i.e., PHRF1 and TRIM66). This study highlights the usefulness of high-throughput analyses of histone reader proteins as a means of understanding how chromatin engagement occurs biochemically.

The aim was to investigate the impact of an automatic in ovo injection of the raffinose family oligosaccharides (RFO) extracted from the seeds of Lupinus luteus L, on the chicken performance and resistance in a production environment. At day 12 of incubation, a total of 57,900 eggs (Ross 308) were divided into two groups: 1/ Control, injected with 0.9% NaCl and 2/ RFO group, injected with 1.9 mg/egg of the lupin seed extract, dissolved in 0.2 mL NaCl. The performance parameters, biochemical indices (lipid profile, hepatic parameters), gut histomorphology and duodenum structure, oxidative stability of the meat and microbiological counts of the major commensal microbiota species were analyzed. Mortality, body weight, and feed conversion ratio (FCR) were not affected. By day 42, several health indices were improved with RFO and were reflected in a beneficial lipid blood profile, increased villi surface and better combating opportunistic pathogens through reduction of Clostridia and decreased coccidia counts. The RFO increased meat oxidation, but only at the beginning of the storage. The RFO sourced from local legumes can be considered a promising prebiotic for broiler chickens. In ovo delivery of prebiotics and/or synbiotics should be further optimized as an important strategy for the earliest possible modulation of chicken resistance.

The results of enzymatic hydrolysis of birch and beech kraft cellulosic pulps indicate that they may be promising feedstocks for fermentation processes including biofuel manufacturing. The aim of this study was to investigate whether birch and beech wood require the same degree of delignification by kraft pulping as pine wood. The differences observed in the efficiency of hydrolysis for the raw materials tested suggest that the differences in the anatomical structure of the examined wood in relation to pine wood is essential for the efficiency of the enzymatic hydrolysis process. The yields of glucose and other reducing sugars obtained from the birch and beech cellulosic pulps were similar (up to around 75% and 98.3% dry weight, and 76% and 98.6% dry weight, respectively). The highest glucose yields from cellulose contained in the birch and beech pulp were around 81.2% (at a Kappa number of 28.3) and 83.1% (at a Kappa number of 30.4), respectively. The maximum glucose yields and total reducing sugars of birch wood on a dry weight basis (39.8% and 52.1%, respectively) were derived from the pulp at a Kappa number of 28.3, while the highest yields of glucose and total reducing sugars of beech wood on a dry weight basis (around 36.9% and 48.2%, respectively) were reached from the pulp at a Kappa number of 25.3. To obtain the highest glucose yields and total reducing sugars of a wood on a dry weight basis, total lignin elimination from the birch and beech pulps was not necessary. However more in-depth delignification of birch and beech wood is required than for pine wood.