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Medium-chain-length polyprenol (C45–C55) formation in chloroplasts of Arabidopsis is brassinosteroid-dependent
Get PDFBrassinosteroids are important plant hormones influencing, among other processes, chloroplast development, the electron transport chain during light reactions of photosynthesis, and the Calvin-Benson cycle. Medium-chainlength polyprenols built of 9–11 isoprenoid units (C45–C55 carbons) are a class of isoprenoid compounds present in abundance in thylakoid membranes. They are synthetized in chloroplast by CPT7 gene from Calvin cycle derived precursors on MEP methylerythritol 4-phosphate) isoprenoid biosynthesis pathway. C45–C55 polyprenols affect thylakoid membrane ultra-structure and hence influence photosynthetic apparatus performance in plants such as Arabidopsis and tomato. So far nothing is known about the hormonal or environmental regulation of CPT7 gene expression. The aim of our study was to find out if medium-chain-length polyprenol biosynthesis in plants may be regulated by hormonal cues.We found that the CPT7 gene in Arabidopsis has a BZR1 binding element (brassinosteroid dependent) in its promoter. Brassinosteroid signaling mutants in Arabidopsis accumulate a lower amount of medium-chain-length C45–C55 polyprenols than control plants. At the same time carotenoid and chlorophyll content is increased, and the amount of PsbD1A protein coming from photosystem II
does not undergo a significant change. On contrary, treatment of WT plants with epi-brassinolide increases
C45–C55 polyprenols content. We also report decreased transcription of MEP enzymes (besides C45–C55 polyprenols,
precursors of numerous isoprenoids, e.g. phytol, carotenoids are derived from this pathway) and genes encoding biosynthesis of medium-chain-length polyprenol enzymes in brassinosteroid perception mutant bri1-116. Taken together, we document that brassinosteroids affect biosynthetic pathway of C45–C55 polyprenols
SnRK2.10 kinase differentially modulates expression of hub WRKY transcription factors genes under salinity and oxidative stress in Arabidopsis thaliana
Get PDFIn nature, all living organisms must continuously sense their surroundings and react to the occurring changes. In the cell, the information about these changes is transmitted to all cellular compartments, including the nucleus, by multiple phosphorylation cascades. Sucrose Non-Fermenting 1 Related Protein Kinases (SnRK2s) are plant-specific enzymes widely distributed across the plant kingdom and key players controlling abscisic acid (ABA)-dependent and ABA independent signaling pathways in the plant response to osmotic stress and salinity. The main deleterious effects of salinity comprise water deficiency stress, disturbances in
ion balance, and the accompanying appearance of oxidative stress. The reactive oxygen species (ROS) generated at the early stages of salt stress are involved in triggering intracellular signaling required for the fast stress response and modulation of gene expression. Here we established in Arabidopsis thaliana that salt stress or induction of ROS accumulation by treatment of plants with
H2 O 2 or methyl viologen (MV) induces the expression of several genes encoding transcription factors (TFs) from the WRKY DNA-Binding Protein (WRKY) family. Their induction by salinity was dependent on SnRK2.10, an ABA non-activated
kinase, as it was strongly reduced in snrk2.10 mutants. The effect of ROS was clearly dependent on their source. Following the H2 O 2 treatment, SnRK2.10 was activated in wild-type (wt) plants and the induction of the WRKY TFs expression was only moderate and was enhanced in snrk2.10 lines. In contrast, MV did not activate SnRK2.10 and the WRKY induction was very strong and was similar in wt
and snrk2.10 plants. A bioinformatic analysis indicated that the WRKY33, WRKY40, WRKY46, and WRKY75 transcription factors have a similar target range comprising numerous stress-responsive protein kinases. Our resultsindicate that the stress-related functioning of SnRK2.10 is fine-tuned by the
source and intracellular distribution of ROS and the co-occurrence of other stress factors
Toward an understanding of the conformational plasticity of S100A8 and S100A9 Ca2+-binding proteins
Get PDFS100A8 and S100A9 are small, human, Ca2+-binding proteins with multiple intracellular and extracellular functions in signaling, regulation, and defense. The two proteins are not detected as monomers but form various noncovalent homo- or hetero-oligomers related to specific activities in human physiology. Because of their significant roles in numerous medical conditions, there has been intense research on the conformational properties of various S100A8 and S100A9 proteoforms as essential targets of drug discovery. NMR or crystal structures are currently available only for mutated or truncated protein complexes, mainly with bound metal ions, that may well reflect the proteins' properties outside cells but not in other biological contexts in which they perform. Here, we used structural mass spectrometry methods combined with molecular dynamics simulations to compare the conformations of wildtype full-length S100A8 and S100A9 subunits in biologically relevant homo- and heterodimers and in higher oligomers formed in the presence of calcium or zinc ions. We provide, first, rationales for their functional response to changing environmental conditions, by elucidating differences between proteoforms in flexible protein regions that may provide the plasticity of the binding sites for the multiple targets, and second, the key factors contributing to the variable stability of the oligomers. The described methods and a systematic view of the conformational properties of S100A8 and S100A9 complexes provide a basis for further research to characterize and modulate their functions for basic science and therapies
New molecular targets in Hodgkin and Reed-Sternberg cells
Get PDFRecent discoveries shed light on molecular mechanisms responsible for classical Hodgkin lymphoma (HL) development and progression, along with features of Hodgkin – Reed and Sternberg cells (HRS). Here, we summarize current knowledge on characteristic molecular alterations in HL, as well as existing targeted therapies and potential novel treatments for this disease. We discuss the importance of cluster of differentiation molecule 30 (CD30) and the programmed cell death-1 protein (PD-1) and ligands (PD-L1/2), and other molecules involved in immune modulation in HL. We highlight emerging evidence indicating that the altered function of SWI/SNF-type chromatin remodeling complexes, PRC2, and other epigenetic modifiers, contribute to variations in chromatin status, which are typical for HL. We postulate that despite of the existence of plentiful molecular data, the understanding of HL development remains incomplete. We therefore propose research directions involving analysis of reverse signaling in the PD-1/PD-L1 mechanism, chromatin remodeling, and epigenetics-related alterations, in order to identify HL features at the molecular level. Such attempts may lead to the identification of new molecular targets, and thus will likely substantially contribute to the future development of more effective targeted therapies
Promoter‐pervasive transcription causes RNA polymerase II pausing to boost DOG1 expression in response to salt
Get PDFEukaryotic genomes are pervasively transcribed by RNA polymerase II. Yet, the molecular and biological implications of such a phenomenon are still largely puzzling. Here, we describe noncoding RNA transcription upstream of the Arabidopsis thaliana DOG1 gene, which governs salt stress responses and is a key regulator of seed dormancy. We find that expression of the DOG1 gene is induced by salt stress, thereby causing a delay in seed germination. We uncover extensive transcriptional activity on the promoter of the DOG1 gene, which produces a variety of lncRNAs. These lncRNAs, named PUPPIES, are co-directionally transcribed and extend into the DOG1 coding region. We show that PUPPIES RNAs respond to salt stress and boost DOG1 expression, resulting in delayed germination. This positive role of pervasive PUPPIES transcription on DOG1 gene expression is associated with augmented pausing of RNA polymerase II, slower transcription and higher transcriptional burst size. These findings highlight the positive role of upstream co-directional transcription in controlling transcriptional dynamics of downstream genes
A non-canonical function of Arabidopsis ERECTA proteins and a role of the SWI3B subunit of the SWI/SNF chromatin remodeling complex in gibberellin signaling
Get PDFThe Arabidopsis ERECTA family (ERf) of leucine-rich repeat receptor-like kinases (LRR-RLKs) comprising
ERECTA (ER), ERECTA-LIKE 1 (ERL1), and ERECTA-LIKE 2 (ERL2) controls epidermal patterning, inflorescence
architecture, and stomata development and patterning. These proteins are reported to be plasma membrane associated. Here we show that the er/erl1/erl2 mutant exhibits impaired gibberellin (GA) biosynthesis
and perception alongside broad transcriptional changes. The ERf kinase domains were found to localize to
the nucleus where they interact with the SWI3B subunit of the SWI/SNF chromatin remodeling complex
(CRCs). The er/erl1/erl2 mutant exhibits reduced SWI3B protein level and affected nucleosomal chromatin
structure. Similar to swi3c and brm plants with inactivated subunits of SWI/SNF CRCs, it also does not
accumulate DELLA RGA and GAI proteins. The ER kinase phosphorylates SWI3B in vitro, and the inactiva�tion of all ERf proteins leads to the decreased phosphorylation of SWI3B protein in vivo. The identified cor�relation between DELLA overaccumulation and SWI3B proteasomal degradation, and the physical
interaction of SWI3B with DELLA proteins indicate an important role of SWI3B-containing SWI/SNF CRCs in
gibberellin signaling. Co-localization of ER and SWI3B on GID1 (GIBBERELLIN INSENSITIVE DWARF 1)
DELLA target gene promoter regions and abolished SWI3B binding to GID1 promoters in er/erl1/erl2 plants
supports the conclusion that ERf-SWI/SNF CRC interaction is important for transcriptional control of GA
receptors. Thus, the involvement of ERf proteins in the transcriptional control of gene expression, and
observed similar features for human HER2 (epidermal growth family receptor member), indicate an exciting
target for further studies of evolutionarily conserved non-canonical functions of eukaryotic membrane
receptors
Fucose as a nutrient ligand for Dikarya and a building block of early diverging lineages
Get PDFFucose is a deoxyhexose sugar present and studied in mammals. The process of fucosylation has been the primary focus in studies relating to fucose in animals due to the presence of fucose in Lewis antigens. Very few studies have reported its presence in Fungi, mostly in Mucoromycotina. The constitution of 25% and 12% of this sugar in the carbohydrates of cell wall in the respective Umbelopsis and Mucorales strains boosts the need to bridge the gap of knowledge on fucose metabolism across the fungal tree of life. In the absence of a network map involving fucose proteins, we carried out an in-silico approach to construct the fucose metabolic map in Fungi. We analyzed the taxonomic distribution of 85 protein families in Fungi including diverse early diverging fungal lineages. The expression of fucose-related protein-coding genes proteins was validated with the help of transcriptomic data originating from representatives of early diverging fungi. We found proteins involved in several metabolic activities apart from fucosylation such as synthesis, transport and binding. Most of the identified protein families are shared with Metazoa suggesting an ancestral origin in Opisthokonta. However, the overall complexity of fucose metabolism is greater in Metazoa than in Fungi. Massive gene loss has shaped the evolutionary history of these metabolic pathways, leading to a repeated reduction of these pathways in most yeast-forming lineages. Our results point to a distinctive mode of utilization of fucose among fungi belonging to Dikarya and the early diverging lineages. We speculate that, while Dikarya used fucose as a source of nutrients for metabolism, the early diverging group of fungi depended on fucose as a building block and signaling compound
Strand specificity of ribonucleotide excision repair in Escherichia coli
Get PDFIn Escherichia coli, replication of both strands of genomic DNA is carried out by a single replicase—DNA polymerase III holoenzyme (pol III HE). However, in certain genetic backgrounds, the low-fidelity TLS polymerase, DNA polymerase V (pol V) gains access to undamaged genomic DNA where it promotes elevated levels of spontaneous mutagenesis preferentially on the lagging strand. We employed active site mutants of pol III (pol IIIα_S759N) and pol V (pol V_Y11A) to analyze ribonucleotide incorporation and removal from the E. coli chromosome on a genome-wide scale under conditions of normal replication, as well as SOS induction. Using a variety of methods tuned to the specific properties of these polymerases (analysis of lacI mutational spectra, lacZ reversion assay, HydEn-seq, alkaline gel electrophoresis), we present evidence that repair of ribonucleotides from both DNA strands in E. coli is unequal. While RNase HII plays a primary role in leading-strand Ribonucleotide Excision Repair (RER), the lagging strand is subject to other repair systems (RNase HI and under conditions of SOS activation also Nucleotide Excision Repair). Importantly, we suggest that RNase HI activity can also influence the repair of single ribonucleotides incorporated by the replicase pol III HE into the lagging strand
What can be lost? Genomic perspective on the lipid metabolism of Mucoromycota
Get PDFMucoromycota is a phylum of early diverging fungal (EDF) lineages, of mostly plant-associated terrestrial fungi. Some
strains have been selected as promising biotechnological organisms due to their ability to produce polyunsaturated
fatty acids and efficient conversion of nutrients into lipids. Others get their lipids from the host plant and are unable
to produce even the essential ones on their own. Following the advancement in EDF genome sequencing, we carried
out a systematic survey of lipid metabolism protein families across different EDF lineages. This enabled us to explore
the genomic basis of the previously documented ability to produce several types of lipids within the fungal tree
of life. The core lipid metabolism genes showed no significant diversity in distribution, however specialized lipid meta-
bolic pathways differed in this regard among different fungal lineages. In total 165 out of 202 genes involved in lipid
metabolism were present in all tested fungal lineages, while remaining 37 genes were found to be absent in some
of fungal lineages. Duplications were observed for 69 genes. For the first time we demonstrate that ergosterol
is not being produced by several independent groups of plant-associated fungi due to the losses of different ERG
genes. Instead, they possess an ancestral pathway leading to the synthesis of cholesterol, which is absent in other
fungal lineages. The lack of diacylglycerol kinase in both Mortierellomycotina and Blastocladiomycota opens the ques-
tion on sterol equilibrium regulation in these organisms. Early diverging fungi retained most of beta oxidation com-
ponents common with animals including Nudt7, Nudt12 and Nudt19 pointing at peroxisome divergence in Dikarya.
Finally, Glomeromycotina and Mortierellomycotina representatives have a similar set of desaturases and elongases
related to the synthesis of complex, polyunsaturated fatty acids pointing at an ancient expansion of fatty acid metab-
olism currently being explored by biotechnological studies
Impairment of the non-catalytic subunit Dpb2 of DNA Pol ɛ results in increased involvement of Pol δ on the leading strand
Get PDFThe generally accepted model assumes that leading strand synthesis is performed by Pol ε, while lagging-strand
synthesis is catalyzed by Pol δ. Pol ε has been shown to target the leading strand by interacting with the CMG
helicase [Cdc45 Mcm2–7 GINS(Psf1–3, Sld5)]. Proper functioning of the CMG-Pol ε, the helicase-polymerase
complex is essential for its progression and the fidelity of DNA replication. Dpb2p, the essential non-catalytic subunit of Pol ε plays a key role in maintaining the correct architecture of the replisome by acting as a link between Pol ε and the CMG complex. Using a temperature-sensitive dpb2–100 mutant previously isolated in our laboratory, and a genetic system which takes advantage of a distinct mutational signature of the Pol δ-L612M variant which allows detection of the involvement of Pol δ in the replication of particular DNA strands we show that in yeast cells with an impaired Dpb2 subunit, the contribution of Pol δ to the replication of the leading strand is significantly increased