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Nuclei undergo dynamic shape changes during plant development, but the mechanism is unclear. In Arabidopsis, Sad1/UNC-84 (SUN) proteins, WPP domain-interacting proteins (WIPs), WPP domain-interacting tail-anchored proteins (WITs), myosin XI-i, and CROWDED NUCLEI 1 (CRWN1) have been shown to be essential for nuclear elongation in various epidermal cell types. It has been proposed that WITs serve as adaptors linking myosin XI-i to the SUN-WIP complex at the nuclear envelope (NE). Recently, an interaction between Arabidopsis SUN1 and SUN2 proteins and CRWN1, a plant analog of lamins, has been reported. Therefore, the CRWN1-SUN-WIP-WIT-myosin XI-i interaction may form a linker of the nucleoskeleton to the cytoskeleton complex. In this study, we investigate this proposed mechanism in detail for nuclei of Arabidopsis root hairs and trichomes. We show that WIT2, but not WIT1, plays an essential role in nuclear shape determination by recruiting myosin XI-i to the SUN-WIP NE bridges. Compared with SUN2, SUN1 plays a predominant role in nuclear shape. The NE localization of SUN1, SUN2, WIP1, and a truncated WIT2 does not depend on CRWN1. While crwn1 mutant nuclei are smooth, the nuclei of sun or wit mutants are invaginated, similar to the reported myosin XI-i mutant phenotype. Together, this indicates that the roles of the respective WIT and SUN paralogs have diverged in trichomes and root hairs, and that the SUN-WIP-WIT2-myosin XI-i complex and CRWN1 independently determine elongated nuclear shape. This supports a model of nuclei being shaped both by cytoplasmic forces transferred to the NE and by nucleoplasmic filaments formed under the NE. [ABSTRACT FROM AUTHOR]

Cnidarians living in symbiosis with photosynthetic cells—called zooxanthellae—are submitted to high oxygen levels generated by photosynthesis. To cope with this hyperoxic state, symbiotic cnidarians present a high diversity of superoxide dismutases (SOD) isoforms. To understand better the mechanism of resistance of cnidarian hosts to hyperoxia, we studied copper- and zinc-containing SOD (CuZnSOD) from Anemonia viridis, a temperate symbiotic sea anemone. We cloned two CuZnSOD genes that we call AvCuZnSODa and AvCuZnSODb. Their molecular analysis suggests that the AvCuZnSODa transcript encodes an extracellular form of CuZnSOD, whereas the AvCuZnSODb transcript encodes an intracellular form. Using in situ hybridization, we showed that both AvCuZnSODa and AvCuZnSODb transcripts are expressed in the endodermal and ectodermal cells of the sea anemone, but not in the zooxanthellae. The genomic flanking sequences of AvCuZnSODa and AvCuZnSODb revealed different putative binding sites for transcription factors, suggesting different modes of regulation for the two genes. This study represents a first step in the understanding of the molecular mechanisms of host animal resistance to permanent hyperoxia status resulting from the photosynthetic symbiosis. Moreover, AvCuZnSODa and AvCuZnSODb are the first SODs cloned from a diploblastic animal, contributing to the evolutionary understanding of SODs. [Copyright &y& Elsevier]

The nucleotide sequence of the abnx cDNA gene, which encodes an exo-arabinanase (Abnx) of Penicillium chrysogenum 31B, was determined. Abnx was found to be structurally distinct from known arabinan-degrading enzymes based on its amino acid sequence and a hydrophobic cluster analysis. The protein in the protein database with the highest similarity to Abnx was the Neurospora crassa conserved hypothetical protein. The abnx cDNA gene product expressed in Escherichia coli catalyzed the release of arabinobiose from α-1,5-L-arabinan. The activity of the recombinant Abnx towards a series of arabino-oligosaccharides, as expressed by kcat/Km value, was greatest with arabinohexaose. [Copyright &y& Elsevier]

FOXJ2 is a fork head transcriptional activator, the expression of which starts very early in embryonic development and it is distributed widely in the adult. Here, we describe the characterization of domains that are important for its function. FOXJ2 is localized constitutively at the nucleus of the cell. Two tyrosine residues and a stretch of basic amino acid residues at the N and C-terminal ends of the fork head domain, respectively, are important for its nuclear targeting. These residues are conserved strongly among all members of the fork head family, suggesting that they could be involved in the nuclear translocation mechanism of all fork head factors. In addition to the AB domain, we have found, at least, two other transactivation domains: Domain I, at the N terminus, and the H/P domain, rich in histidine and proline residues. Although the AB domain shows the strongest transactivation capacity, all three domains are required for full FOXJ2 transcriptional activity. Furthermore, a fourth region rich in proline and glutamine residues and with no intrinsic transactivation function, the P/Q domain, appears to play an important role in the FOXJ2-mediated transactivation mechanism. Although FOXJ2 can be phosphorylated in two serine residues, this post-translational modification did not appear to be essential for transactivation. Finally, we have found that the W2 wing of the fork head domain of FOXJ2 is dispensable for specific DNA binding, although it could have a weak stabilizing role for the DNA–FOXJ2 complex. [Copyright &y& Elsevier]

We report cloning and characterization of a 2.8 kb DNA fragment that suppressed the aerial mycelium-deficient phenotype of an amfR mutant of Streptomyces griseus when it was introduced on a high-copy-number plasmid. Nucleotide sequencing revealed that the cloned DNA fragment contained a part of a regulatory operon homologous to one of the conserved operons identified in the genome of Streptomyces coelicolor A3(2). The operon appeared to consist of 5 CDSs (rarA–E; restoration of aerial mycelium formation in an amfR mutant): rarA encoded a membrane protein with weak similarity to the histidine kinase of the two-component regulatory system; rarB and rarC products did not show marked similarity to other proteins with known function; rarD encoded a G-protein carrying two GTP-binding consensus sequences conserved in the eukaryotic Ras-like proteins; rarE product showed end-to-end homology to cytochrome P450. The 2.8 kb fragment contained a 5′-end incomplete rarA and complete rarB-D in the downstream from the promoter region of mel operon of the vector plasmid. Subcloning showed that the region containing rarA only is sufficient for the aerial mycelium-inducing activity. The truncation of rarA at its 5′ terminus was essential for the restoration activity, which implied that the mutated rarA product causes unusual signaling that directs the onset of morphogenesis without amfR function. Inactivation of both rarA in Streptomyces griseus and cvnD9, a rarD ortholog in S. coelicolor resulted in precocious and glucose-resistant formation of aerial mycelium and secondary metabolites, which suggested that the operon negatively regulates the onset of differentiation. S1 nuclease protection analysis showed that the transcriptional activity of the promoter preceding rarA is developmentally regulated in an amfR- and glucose-dependent manner. [Copyright &y& Elsevier]

Tubulin is one of the most widespread classes of multiprotein families and is well known to construct microtubules with two different subunits, α- and β-tubulin. In the course of genome analysis of Bombyx mori, we have constructed an EST database by large-scale sequencing of clones that were randomly selected from cDNA libraries of various tissues and organs belonging to different developmental stages. Using this EST database, we have identified four types of β-tubulin gene and three types of α-tubulin gene. Based on the analysis of deduced amino acid sequences, we have determined the phylogenetic relationships of tubulins between Bombyx and Drosophila melanogaster as well as two other moth species, suggesting that each tubulin is classified into at least three distinct subfamilies: a ubiquitously expressed one, a developmentally regulated one and a testis specific one. [Copyright &y& Elsevier]

The first animal polygalacturonase (PG, EC 2.1.15) encoding cDNA, Mi-pg-1, was cloned from the plant parasitic nematode Meloidogyne incognita. The enzymatic activity of MI-PG-1 was confirmed after heterologous expression in Escherichia coli. The presence of a predicted signal peptide on the MI-PG-1 sequence together with the specific localization of the transcripts of the Mi-pg-1 gene in the oesophageal glands of infective juveniles imply that MI-PG-1 could be secreted into plant tissues. The potential role of MI-PG-1 in parasitism is discussed. [Copyright &y& Elsevier]

The β2 subunit of the interleukin (IL)-12 receptor (IL-12Rβ2) has been shown to play an essential role in differentiation of T helper 1 (Th1) cells in the murine and human system, and antibodies raised against IL-12Rβ2 recognized this molecule on human Th1 but not Th2 cells. However, while the cytokines secreted by clones of murine cells allowed the definition of distinct T helper cell subsets, bovine clones with polarized Th1 and Th2 cytokine profiles were rarely found. This raised important questions about the regulation of immune responses in cattle. We therefore cloned bovine IL-12Rβ2 (boIL-12Rβ2) DNA complementary to RNA (cDNA) from the start codon to the 3′ end of the mRNA. Comparison of boIL-12Rβ2 cDNA with human and murine IL-12Rβ2 cDNA sequences revealed homologies of 85 and 78%, respectively. The deduced protein sequence showed the hallmark motifs of the cytokine receptor superfamily including the four conserved cysteine residues, the WSXWS motif and fibronectin domains in the extracellular part as well as a STAT4 binding site in the intracellular part of the molecule. Using real-time reverse transcription–polymerase chain reaction, upregulation of mRNA expression of this molecule could be demonstrated in cultured bovine lymph node cells stimulated with phytohemagglutinin. Furthermore, cells with upregulated boIL-12Rβ2 mRNA responded with enhanced expression of interferon γ to treatment with interleukin 12. [Copyright &y& Elsevier]

The Neuropeptide Y (NPY) family of neuropeptides exert their function through a family of heptahelical G-protein coupled receptors regulating essential physiological processes. A 97 base pair intron (intron IV) intervenes the coding sequence of the human NPY Y1 receptor (hY1) gene and was found frequently retained at variable levels in poly A+ mRNA isolated from multiple human tissues. When included in hY1 expression vectors, either in its natural position or 5′ of the hY1 cDNA, intron IV mediated a significant increase of both hY1 mRNA and corresponding functional receptor protein in transfected mammalian cells, implying an in vivo regulatory function of the endogenous intron. Our results further indicate that the nuclear history of the hY1 pre-mRNA influence ectopic hY1 production through post-transcriptional mechanisms and argues against intron IV acting as a transcriptional enhancer as well as the possibility that a putative hY1 related 5TM accessory protein encoded by the non-spliced hY1 mRNA would facilitate hY1 production on a post-translational level. [Copyright &y& Elsevier]

We report identification and characterization of Kv6.3, a novel member of the voltage-gated K+ channel. Reverse transcriptase-polymerase chain reaction analysis indicated that Kv6.3 was highly expressed in the brain. Electrophysiological studies indicated that homomultimeric Kv6.3 did not yield a functional voltage-gated ion channel. When Kv6.3 and Kv2.1 were co-expressed, the heteromultimeric channels displayed the decreased rate of deactivation compared to the homomultimeric Kv2.1 channels. Immunoprecipitation studies indicated that Kv6.3 bound with Kv2.1 in co-transfected cells. These results indicate that Kv6.3 is a novel member of the voltage-gated K+ channel which functions as a modulatory subunit. [Copyright &y& Elsevier]

CRISPR-dependent base editors enable direct nucleotide conversion without the introduction of double-strand DNA break or donor DNA template, thus expanding the CRISPR toolbox for genetic manipulation. However, designing guide RNAs (gRNAs) for base editors to enable gene correction or inactivation is more complicated than using the CRISPR system for gene disruption. Here, we present a user-friendly web tool named BEtarget dedicated to the design of gRNA for base editing. It is currently supported by 46 plant reference genomes and 5 genomes of non-plant model organisms. BEtarget supports the design of gRNAs with different types of protospacer adjacent motifs (PAM) and integrates various functions, including automatic identification of open reading frame, prediction of potential off-target sites, annotation of codon change, and assessment of gRNA quality. Moreover, the program provides an interactive interface for users to selectively display information about the desired target sites. In brief, we have developed a flexible and versatile web-based tool to simplify complications associated with the design of base editing technology. BEtarget is freely accessible at https://skl.scau.edu.cn/betarget/., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2022 The Author(s).)

Machine learning methods played a major role in improving the accuracy of predictions and classification of DNA (Deoxyribonucleic Acid) and protein sequences. In eukaryotes, Splice-site identification and prediction is though not a straightforward job because of numerous false positives. To solve this problem, here, in this paper, we represent a bidirectional Long Short Term Memory (LSTM) Recurrent Neural Network (RNN) based deep learning model that has been developed to identify and predict the splice-sites for the prediction of exons from eukaryotic DNA sequences. During the splicing mechanism of the primary mRNA transcript, the introns, the non-coding region of the gene are spliced out and the exons, the coding region of the gene are joined. This bidirectional LSTM-RNN model uses the intron features that start with splice site donor (GT) and end with splice site acceptor (AG) in order of its length constraints. The model has been improved by increasing the number of epochs while training. This designed model achieved a maximum accuracy of 95.5%. This model is compatible with huge sequential data such as the complete genome., Competing Interests: The authors declare no conflict of interest, financial or otherwise., (© 2022 The Author(s).)

The emergence of resistance to first-line antimalarials, including artemisinin, the last effective malaria therapy in some regions, stresses the urgent need to develop new effective treatments against this disease. The identification and validation of metabolic pathways that could be targeted for drug development may strongly contribute to accelerate this process. In this study, we use fully characterized specific inhibitors targeting glycan biosynthetic pathways as research tools to analyze their effects on the growth of the malaria parasite Plasmodium falciparum and to validate these metabolic routes as feasible chemotherapeutic targets. Through docking simulations using models predicted by AlphaFold, we also shed new light into the modes of action of some of these inhibitors. Molecules inhibiting N -acetylglucosaminyl-phosphatidylinositol de- N -acetylase (GlcNAc-PI de- N -acetylase, PIGL/GPI12) or the inositol acyltransferase (GWT1), central for glycosylphosphatidylinositol (GPI) biosynthesis, halt the growth of intraerythrocytic asexual parasites during the trophozoite stages of the intraerythrocytic developmental cycle (IDC). Remarkably, the nucleoside antibiotic tunicamycin, which targets UDP- N -acetylglucosamine:dolichyl-phosphate N -acetylglucosaminephosphotransferase (ALG7) and N -glycosylation in other organisms, induces a delayed-death effect and inhibits parasite growth during the second IDC after treatment. Our data indicate that tunicamycin induces a specific inhibitory effect, hinting to a more substantial role of the N -glycosylation pathway in P. falciparum intraerythrocytic asexual stages than previously thought. To sum up, our results place GPI biosynthesis and N -glycosylation pathways as metabolic routes with potential to yield much-needed therapeutic targets against the parasite., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2022 Published by Elsevier B.V. on behalf of Research Network of Computational and Structural Biotechnology.)

Introduction: Tumors are usually refractory to anti-cancer therapeutics under hypoxic conditions. However, the underlying molecular mechanism remains to be elucidated., Objectives: Our study intended to identify hypoxia inducible lncRNAs and their biological function in gastric cancer (GC)., Methods: Differentially expressed lncRNAs were determined by microarray analysis between GC cells exposed to hypoxia (1% O 2 ) and normoxia (21% O 2 ) for 24 h. The expression level of CBSLR was manipulated in several GC cell lines to perform molecular and biological analyses both in vitro and in vivo ., Results: We identified a hypoxia-induced lncRNA- CBSLR that protected GC cells from ferroptosis, leading to chem-resistance. Mechanically, CBSLR interacted with YTHDF2 to form a CBSLR /YTHDF2/CBS signaling axis that decreased the stability of CBS mRNA by enhancing the binding of YTHDF2 with the m6A-modified coding sequence (CDS) of CBS mRNA. Furthermore, under decreased CBS levels, the methylation of the ACSL4 protein was reduced, leading to protein polyubiquitination and degradation of ACSL4. This, in turn, decreased the pro-ferroptosis phosphatidylethanolamine (PE) (18:0/20:4) and PE (18:0/22:4) content and contributed to ferroptosis resistance. Notably, CBSLR is upregulated, whereas CBS is downregulated in GC tissues compared to matched normal tissues; and GC patients with high CBSLR /low CBS levels have a worse clinical outcome and a poorer response to chemotherapy., Conclusion: Our study reveals a novel mechanism in how HIF1α/ CBSLR modulates ferroptosis/chemoresistance in GC, illuminating potential therapeutic targets for refractory hypoxic tumors., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2022 The Authors. Published by Elsevier B.V. on behalf of Cairo University.)

Arcanobacterium haemolyticum can cause deep infections, including osteomyelitis. In this study, an automated system misidentified this causal agent as Cellulomonas species but 16 s rRNA sequencing correctly identified it as A. haemolyticum . Recognizing the capability of A. haemolyticum to establish the disease is of great importance to enable accurate diagnosis and begin the suitable antibiotic therapy. Here we present the first case of successfully treated A. haemolyticum infective osteomyelitis in a 64-year-old Saudi patient with diabetes mellitus type 2 and review the characteristics of this seldom pathogenic agent., Competing Interests: The authors report no declarations of interest., (© 2021 The Authors.)

Insect neuropeptides, play a central role in the control of many physiological processes. Based on an analysis of Nyssorhynchus albimanus brain transcriptome a neuropeptide precursor database of the mosquito was described. Also, we observed that adipokinetic hormone/corazonin-related peptide (ACP), hugin and corazonin encoding genes were differentially expressed during Plasmodium infection. Transcriptomic data from Ny. albimanus brain identified 29 pre-propeptides deduced from the sequences that allowed the prediction of at least 60 neuropeptides. The predicted peptides include isoforms of allatostatin C, orcokinin, corazonin, adipokinetic hormone (AKH), SIFamide, capa, hugin, pigment-dispersing factor, adipokinetic hormone/corazonin-related peptide (ACP), tachykinin-related peptide, trissin, neuropeptide F, diuretic hormone 31, bursicon, crustacean cardioactive peptide (CCAP), allatotropin, allatostatin A, ecdysis triggering hormone (ETH), diuretic hormone 44 (Dh44), insulin-like peptides (ILPs) and eclosion hormone (EH). The analysis of the genome of An. albimanus and the generated transcriptome, provided evidence for the identification of myosuppressin neuropeptide precursor. A quantitative analysis documented increased expression of precursors encoding ACP peptide, hugin and corazonin in the mosquito brain after Plasmodium berghei infection. This work represents an initial effort to characterize the neuropeptide precursors repertoire of Ny. albimanus and provides information for understanding neuroregulation of the mosquito response during Plasmodium infection., Competing Interests: The authors declare that they have no competing interests., (© 2021 The Authors.)

Whilst substantial research effort has been placed on understanding the interactions of plant proteins with their molecular partners, relatively few studies in plants - by contrast to work in other organisms - address how these interactions evolve. It is thought that ancestral proteins were more promiscuous than modern proteins and that specificity often evolved following gene duplication and subsequent functional refining. However, ancestral protein resurrection studies have found that some modern proteins have evolved de novo from ancestors lacking those functions. Intriguingly, the new interactions evolved as a consequence of just a few mutations and, as such, acquisition of new functions appears to be neither difficult nor rare, however, only a few of them are incorporated into biological processes before they are lost to subsequent mutations. Here, we detail the approach of ancestral sequence reconstruction (ASR), providing a primer to reconstruct the sequence of an ancestral gene. We will present case studies from a range of different eukaryotes before discussing the few instances where ancestral reconstructions have been used in plants. As ASR is used to dig into the remote evolutionary past, we will also present some alternative genetic approaches to investigate molecular evolution on shorter timescales. We argue that the study of plant secondary metabolism is particularly well suited for ancestral reconstruction studies. Indeed, its ancient evolutionary roots and highly diverse landscape provide an ideal context in which to address the focal issue around the emergence of evolutionary novelties and how this affects the chemical diversification of plant metabolism., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2021 The Author(s).)

Advanced sequencing technologies such as RNASeq provide the means for production of massive amounts of data, including transcriptome-wide expression levels of coding RNAs (mRNAs) and non-coding RNAs such as miRNAs, lncRNAs, piRNAs and many other RNA species. In silico analysis of datasets, representing only one RNA species is well established and a variety of tools and pipelines are available. However, attaining a more systematic view of how different players come together to regulate the expression of a gene or a group of genes requires a more intricate approach to data analysis. To fully understand complex transcriptional networks, datasets representing different RNA species need to be integrated. In this review, we will focus on miRNAs as key post-transcriptional regulators summarizing current computational approaches for miRNA:target gene prediction as well as new data-driven methods to tackle the problem of comprehensively and accurately dissecting miRNome-targetome interactions., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2021 The Authors.)

Defining genes that are essential for life has major implications for understanding critical biological processes and mechanisms. Although essential genes have been identified and characterised experimentally using functional genomic tools, it is challenging to predict with confidence such genes from molecular and phenomic data sets using computational methods. Using extensive data sets available for the model organism Caenorhabditis elegans , we constructed here a machine-learning (ML)-based workflow for the prediction of essential genes on a genome-wide scale. We identified strong predictors for such genes and showed that trained ML models consistently achieve highly-accurate classifications. Complementary analyses revealed an association between essential genes and chromosomal location. Our findings reveal that essential genes in C. elegans tend to be located in or near the centre of autosomal chromosomes; are positively correlated with low single nucleotide polymorphim (SNP) densities and epigenetic markers in promoter regions; are involved in protein and nucleotide processing; are transcribed in most cells; are enriched in reproductive tissues or are targets for small RNAs bound to the argonaut CSR-1. Based on these results, we hypothesise an interplay between epigenetic markers and small RNA pathways in the germline, with transcription-based memory; this hypothesis warrants testing. From a technical perspective, further work is needed to evaluate whether the present ML-based approach will be applicable to other metazoans (including Drosophila melanogaster ) for which comprehensive data sets (i.e. genomic, transcriptomic, proteomic, variomic, epigenetic and phenomic) are available., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2020 The Authors.)