Correction: Synergetic osteogenesis of extracellular vesicles and loading RGD colonized on 3D-printed titanium implants.

Correction for 'Synergetic osteogenesis of extracellular vesicles and loading RGD colonized on 3D-printed titanium implants' by Shiqing Ma et al., Biomater. Sci., 2022, 10, 4773-4784, https://doi.org/10.1039/D2BM00725H.

Overview of m6A and circRNAs in human cancers.

N6-methyladenosine (m6A), the richest post-transcriptional modification of RNA in eukaryotic cells, is dynamically installed/uninstalled by the RNA methylase complex ("writer") and demethylase ("eraser") and recognized by the m6A-binding protein ("reader"). M6A modification on RNA metabolism involves maturation, nuclear export, translation and splicing, thereby playing a critical role in cellular pathophysiology and disease processes. Circular RNAs (circRNAs) are a class of non-coding RNAs with a covalently closed loop structure. Due to its conserved and stable properties, circRNAs could participate in physiological and pathological processes through unique pathways. Despite the recent discovery of m6A and circRNAs remains in the initial stage, research has shown that m6A modifications are widespread in circRNAs and regulates circRNA metabolism, including biogenesis, cell localization, translation, and degradation. In this review, we describe the functional crosstalk between m6A and circRNAs, and illustrate their roles in cancer development. Moreover, we discuss the potential mechanisms and future research directions of m6A modification and circRNAs.

High-throughput microarray reveals the epitranscriptome-wide landscape of m6A-modified circRNA in oral squamous cell carcinoma.

BACKGROUND: Emerging transcriptome-wide high-throughput screenings reveal the landscape and functions of RNAs, such as circular RNAs (circRNAs), in human cancer. In addition, the post-transcriptional RNA internal modifications, especially N6-methyladenosine (m6A), greatly enrich the variety of RNAs metabolism. However, the m6A modification on circRNAs has yet to be addressed. RESULTS: Here, we report an epitranscriptome-wide mapping of m6A-modified circRNAs (m6A-circRNA) in oral squamous cell carcinoma (OSCC). Utilizing the data of m6A methylated RNA immunoprecipitation sequencing (MeRIP-seq) and m6A-circRNAs microarray, we found that m6A-circRNAs exhibited particular modification styles in OSCC, which was independent of m6A-mRNA. Besides, m6A modification on circRNAs frequently occurred on the long exons in the front part of the coding sequence (CDS), which was distinct from m6A-mRNA that in 3'-UTR or stop codon. CONCLUSION: In conclusion, our work preliminarily demonstrates the traits of m6A-circRNAs, which may bring enlighten for the roles of m6A-circRNAs in OSCC.

Methylated RNA Immunoprecipitation Sequencing Reveals the m6A Landscape in Oral Squamous Cell Carcinoma.

N6-methyladenosine (m6A) is the most common epigenetic modification existing in eukaryocyte transcripts. However, genes related to m6A modification in oral squamous cell carcinoma (OSCC) are still unclear. Here, methylated RNA immunoprecipitation sequencing (MeRIP-Seq) was performed to map the m6A landscape in OSCC and corresponding controls. The m6A peaks are always distributed in the junction of the 3'-untranslated regions (3'-UTRs) and the coding sequences (CDS) of mRNAs, as well as the entire genome of long noncoding RNA (lncRNA). Furthermore, enrichment analysis showed that differentially methylated genes were significantly enriched in NF-kappa B signaling pathway, Hedgehog signaling pathway, etc. In summary, our findings reveal the landscape of m6A modification on mRNAs and lncRNAs in OSCC, which may provide key clues for the precision-targeted therapy of OSCC.

Synergetic osteogenesis of extracellular vesicles and loading RGD colonized on 3D-printed titanium implants.

Titanium (Ti) and its alloys have been universally used as surgical implants, and the clinical need for modifying titanium surfaces to accelerate early stage osseointegration and prevent implant loosening is in huge demand. 3D printing technology is an accurate and controllable method to create titanium implants with complex nanostructures, which provide enough space to react and fit in the microenvironment of cells. Recently, extracellular vesicles (EVs) have attracted attention in promoting osteogenesis. The vesicles derived from bone marrow mesenchymal stem cells (BMSC-EVs) have been proved to pack osteogenic-relative RNAs thereby regulating the osteogenic differentiation and mineralization of the target BMSCs. Arg-Gly-Asp (RGD)-derived peptides are typical peptides used to improve cell attachment and proliferation in bone tissue engineering. A novel strategy is proposed to load RGD-derived peptides on EVs with a fusion peptide (EVsRGD) and colonize EVsRGD on the titanium surface via a specific bonding peptide. In this study, we verify that the presence of EVsRGD enables the realization of the synergetic effect of EVs and RGD, enhancing the osteogenic differentiation and mineralization of BMSCs in vitro, resulting in satisfactory osseointegration around implants in vivo.

The emerging roles of N6-methyladenosine (m6A)-modified long non-coding RNAs in human cancers.

N6-methyladenosine (m6A) epitranscriptional modifications widely exist in RNA, which play critical roles in RNA metabolism and biogenesis processes. Long non-coding RNAs (lncRNAs) are class of non-coding RNAs longer than 200 nucleotides without protein-coding ability. LncRNAs participate in a large number of vital biological progressions. With the great improvement of molecular biology, m6A and lncRNAs are attracting more attention from researchers and scholars. In this review, we overview the current status of m6A and lncRNAs based on the latest research, and propose some viewpoints for future research perspectives.

TERMINAL FLOWER 1 and TERMINAL FLOWER 1d respond to temperature and photoperiod signals to inhibit determinate growth in cucumber.

Plants monitor environmental cues to balance their vegetative and productive growth by optimizing their inflorescence architecture. TERMINAL FLOWER 1 (TFL1) and its orthologs regulate the inflorescence structure in cucumber, yet the mechanisms underlying their responses to environmental factors and the formation of terminal flowers remain elusive. Here, we performed map-based cloning to identify the gene that controls a season-dependent determinate growth phenotype and found that it was caused by the complete deletion of CsTFL1 in the genome of cucumber line WI1983Hde. In the CsTFL1 deletion plants (CsTFL1del ), determinate growth could be partially rescued by high-temperature and long-day conditions. The expressions of CsTFL1 and its ortholog CsTFL1d could be upregulated by long-day and high-temperature signals. Knockdown of CsTFL1d resulted in determinate growth and the formation of terminal flowers in WT. These results indicate that the induction of CsTFL1d expression by long-day and high-temperature might partially rescue determinate growth of CsTFL1del . Furthermore, biochemical analyses showed that CsTFL1d interacts directly with CsNOT2a, which indicated that CsTFL1d and CsTFL1 function via similar regulatory mechanism. Our data suggest that CsTFL1 and CsTFL1d co-contribute to inhibit determinate growth by responding to temperature and photoperiod signals. It provides mechanistic insights into how environmental cues sculpt the inflorescence architecture of cucumber.

A SNP of HD-ZIP I transcription factor leads to distortion of trichome morphology in cucumber (Cucumis sativus L.).

BACKGROUND: Trichomes are excellent model systems for the analysis of cell differentiation and play essential roles in plant protection. From cucumber inbred line 'WD1', we identified an EMS-induced trichome abnormally developing mutant, nps, which exhibited smaller, denser and no pyramid-shaped head trichomes. RESULTS: Using F2 and BC1 populations constructed from a cross between nps and '9930', the genetic analysis showed that the nps trait is controlled by a single recessive nuclear gene. We identified CsNps by map-based cloning with 576 individuals of the F2 population generated from the cross of nps and inbred line '9930'. The CsNps was located at a 13.4-kb genomic region on chromosome 3, which region contains three predicted genes. Sequence analysis showed that only one single nucleotide mutation (C → T) between 9930 and nps was found in the second exon of Csa3G748220, a plant-specific class I HD-Zip gene. The result of allelism test also indicated that nps is a novel allelic mutant of Mict (Micro-trichome). Thus, nps was renamed mict-L130F. By comparing the transcriptome of mict-L130F vs WD1 and 06-2 (mict) vs 06-1 (wildtype, near-isogenic line of 06-2), several potential target genes that may be related to trichome development were identified. CONCLUSIONS: Our results demonstrate that Mict-L130F is involved in the morphogenesis of trichomes. Map-based cloning of the Mict-L130F gene could promote the study of trichome development in cucumber.

A Positive Feedback Loop Mediated by CsERF31 Initiates Female Cucumber Flower Development: ETHYLENE RESPONSE FACTOR31 mediates a positive feedback loop that initiates female cucumber flower development.

Sex determination is a crucially important developmental event that is pervasive throughout nature and enhances the adaptation of species. Among plants, cucumber (Cucumis sativus L.) can generate both unisexual and bisexual flowers, and the sex type is mainly controlled by several 1-aminocyclopropane-1-carboxylic acid (ACC) synthases. However, the regulatory mechanism of these synthases remains elusive. Here, we used gene expression analysis, protein-DNA interaction assays and transgenic plants to study the function of a gynoecium-specific gene, ETHYLENE RESPONSE FACTOR31 (CsERF31), in female flower differentiation. We found that in a predetermined female flower, ethylene signalling activates CsERF31 by CsEIN3, and then CsERF31 stimulates CsACS2, which triggers a positive feedback loop to ensure female rather than bisexual flower development. A similar interplay is functionally conserved in melon (Cucumis melo L.). Knockdown of CsERF31 by RNAi causes defective bisexual flowers to replace female flowers. Ectopic expression of CsERF31 suppresses stamen development and promotes pistil development in male flowers, demonstrating that CsERF31 functions as a sex switch. Taken together, our data confirm that CsERF31 represents the molecular link between female-male determination and female-bisexual determination, and provide mechanistic insight into how ethylene promotes female flowers, rather than bisexual flowers, in cucumber sex determination.

A positive feedback loop mediated by CsERF31 initiates female cucumber flower development.

Sex determination is a crucially important developmental event that is pervasive throughout nature and enhances the adaptation of species. Among plants, cucumber (Cucumis sativus L.) can generate both unisexual and bisexual flowers, and the sex type is mainly controlled by several 1-aminocyclopropane-1-carboxylic acid synthases (CsACSs). However, the regulatory mechanism of these synthases remains elusive. Here, we used gene expression analysis, protein-DNA interaction assays, and transgenic plants to study the function of a gynoecium-specific gene, ETHYLENE RESPONSE FACTOR31 (CsERF31), in female flower differentiation. We found that in a predetermined female flower, ethylene signaling activates CsERF31 by CsEIN3, and then CsERF31 stimulates CsACS2, which triggers a positive feedback loop to ensure female rather than bisexual flower development. A similar interplay is functionally conserved in melon (Cucumis melo L.). Knockdown of CsERF31 by RNAi causes defective bisexual flowers to replace female flowers. Ectopic expression of CsERF31 suppresses stamen development and promotes pistil development in male flowers, demonstrating that CsERF31 functions as a sex switch. Taken together, our data confirm that CsERF31 represents the molecular link between female-male determination and female-bisexual determination, and provide mechanistic insight into how ethylene promotes female flowers, rather than bisexual flowers, in cucumber sex determination.

CsUFO is involved in the formation of flowers and tendrils in cucumber.

KEY MESSAGE: An unusual flower and tendril (uft) mutant in cucumber was caused by a mutation in Csa1G056950 encoding an F-box protein. Flowers and tendrils are important agronomic and yield traits of cucumber (Cucumis sativus L.). In this study, we identified an unusual flower and tendril (uft) mutant from an ethyl methanesulfonate (EMS) mutagenesis population. Genetic analysis revealed that the phenotype of the uft mutant was regulated by a single recessive nuclear gene. Map-based cloning and MutMap+ results demonstrated that Csa1G056950 (CsUFO), encoding an F-box protein, was the causal gene for the uft mutant phenotype of cucumber. A single nucleotide polymorphism (SNP) mutation (C to T) in the second exon of CsUFO resulted in premature translation termination. The expression level of CsUFO was significantly decreased in apical buds of the uft mutant compared with the wild-type (WT) WD1. Transcriptome analysis indicated that many genes for organ development were down-regulated in uft plants, suggesting CsUFO-associated networks that regulate flower and tendril development. These findings provide a new insight into understanding the molecular mechanisms of flower organogenesis in cucumber.

Study of micro-trichome (mict) reveals novel connections between transcriptional regulation of multicellular trichome development and specific metabolism in cucumber.

Trichomes that cover the epidermis of aerial plant organs play multiple roles in plant protection. Compared with a unicellular trichome in model plants, the development mechanism of the multicellular trichome is largely unclear. Notably, variations in trichome development are often accompanied by defects in the biosynthesis of cuticle and secondary metabolites; however, major questions about the interactions between developmental differences in trichomes and defects in metabolic pathways remain unanswered. Here, we characterized the glabrous mutant mict/csgl1/cstbh via combined metabolomic and transcriptomic analyses to extend our limited knowledge regarding multicellular trichome development and metabolism in cucumber. Mict was found to be explicitly expressed within trichome cells. Transcriptomic analysis indicated that genes involved in flavonoid and cuticle metabolism are significantly downregulated in mict mutants. Further metabolomic analysis confirmed that flavonoids, lipids, and cuticle compositions are dramatically altered in mict mutants. Additional studies revealed that Mict regulates flavonoid, lipid, and cuticle biosynthesis by likely directly binding to downstream functional genes, such as CsTT4, CsFLS1, CsCER26, and CsMYB36. These findings suggest that specific metabolic pathways (e.g., flavonoids and cuticle components) are co-regulated by Mict and provide insights into transcriptional regulation mechanisms of multicellular trichome development and its specific metabolism in cucumber.

Genome-Wide Identification and Characterization of the TCP Gene Family in Cucumber (Cucumis sativus L.) and Their Transcriptional Responses to Different Treatments.

TCP proteins are plant-specific transcription factors widely implicated in leaf morphogenesis and senescence, flowering, lateral branching, hormone crosstalk, and stress responses. However, the relationship between the transcription pattern of TCPs and organ development in cucumber has not been systematically studied. In this study, we performed a genome-wide identification of putative TCP genes and analyzed their chromosomal location, gene structure, conserved motif, and transcript expression. A total of 27 putative TCP genes were identified and characterized in cucumber. All 27 putative CsTCP genes were classified into class I and class II. Class I comprised 12 CsTCPs and Class II contained 15 CsTCPs. The 27 putative CsTCP genes were randomly distributed in five of seven chromosomes in cucumber. Four putative CsTCP genes were found to contain putative miR319 target sites. Quantitative RT-PCR revealed that 27 putative CsTCP genes exhibited different expression patterns in cucumber tissues and floral organ development. Transcript expression and phenotype analysis showed that the putative CsTCP genes responded to temperature and photoperiod and were induced by gibberellin (GA)and ethylene treatment, which suggested that CsTCP genes may regulate the lateral branching by involving in multiple signal pathways. These results lay the foundation for studying the function of cucumber TCP genes in the future.

The HD-ZIP IV transcription factor Tril regulates fruit spine density through gene dosage effects in cucumber.

Trichomes and fruit spines are important traits that directly affect the appearance quality and commercial value of cucumber (Cucumis sativus). Tril (Trichome-less), encodes a HD-Zip IV transcription factor that plays a crucial role in the initiation of trichomes and fruit spines, but little is known about the details of the regulatory mechanisms involved. In this study, analysis of tissue expression patterns indicated that Tril is expressed and functions in the early stages of organ initiation and development. Expression of Tril under the control of its own promoter (the TrilPro::Tril-3*flag fragment) could partly rescue the mutant phenotypes of tril, csgl3 (cucumber glabrous 3, an allelic mutant of tril), and fs1 (few spines 1, a fragment substitution in the Tril promoter region), providing further evidence that Tril is responsible for the initiation of trichomes and fruit spines. In lines with dense spine, fs1-type lines, and transgenic lines of different backgrounds containing the TrilPro::Tril-3*flag foreign fragment, spine density increased in conjunction with increases in Tril expression, indicating that Tril has a gene dosage effect on fruit spine density in cucumber. Numerous Spines (NS) is a negative regulatory factor of fruit spine density. Characterization of the molecular and genetic interaction between Tril and NS/ns demonstrated that Tril functions upstream of NS with respect to spine initiation. Overall, our results reveal a novel regulatory mechanism governing the effect of Tril on fruit spine development, and provide a reference for future work on breeding for physical quality in cucumber.

Cucumber CsTRY Negatively Regulates Anthocyanin Biosynthesis and Trichome Formation When Expressed in Tobacco.

The development of trichomes (spines) on cucumber fruits is an important agronomic trait. It has been reported that two MYB family members, CsMYB6 (Csa3G824850) and CsTRY (Csa5G139610) act as negative regulators of trichome or fruit spine initiation. To further study the functions of these two genes, we overexpressed them in tobacco, and found that the flowers and seed coats of transformants overexpressing CsTRY displayed an unexpected defect in pigmentation that was not observed in plants overexpressing CsMYB6. Moreover, the expression of key genes in the flavonoid synthesis pathway was repressed in CsTRY overexpressing plants, which resulted in the decrease of several important flavonoid secondary metabolites. In addition, CsTRY could interact with the AN1 homologous gene CsAN1 (Csa7G044190) in cucumber, which further confirmed that CsTRY not only regulates the development of fruit spines, but also functions in the synthesis of flavonoids, acting as the repressor of anthocyanin synthesis.

Mapping and identification of CsUp, a gene encoding an Auxilin-like protein, as a putative candidate gene for the upward-pedicel mutation (up) in cucumber.

BACKGROUND: Pedicel orientation can affect the female flower orientation and seed yield in cucumber. A spontaneous mutant possessing upward growth of pedicels was identified in the wild type inbred strain 9930 and named upward-pedicel (up). The morphological and genetic analyses of up were performed in this study. In order to clone the up gene, 933 F2 individuals and 524 BC1 individuals derived from C-8-6 (WT) and up were used for map-based cloning. RESULTS: up was mapped to a 35.2 kb physical interval on chromosome 1, which contains three predicted genes. Sequencing analysis revealed that a 5-bp deletion was found in the second exon of Csa1G535800, and it led to a frameshift mutation resulting in a premature stop codon. The candidate gene of CsUp (Csa1G535800) was further confirmed via genomic and cDNA sequencing in biparental and natural cucumber populations. Sequencing data showed that a 4-bp deletion was found in the sixth exon of Csa1G535800 in CGN19839, another inbred line, and there was also a mutation of an amino acid in Csa1G535800 that could contribute to the upward growth of pedicels in CGN19839. Moreover, it was found that Csa1G535800 exhibited strong expression in the pedicel of WT, suggesting its important role in development of pedicel orientation. Thus, Csa1G535800 was considered to be the candidate gene of CsUp. CONCLUSIONS: CsUp encodes an Auxilin-like protein and controls pedicel orientation in cucumber. The identification of CsUp may help us to understand the mechanism of pedicel orientation development and allow for investigation of novel functions of Auxilin-like proteins in cucumber.

Fiber-based monolithic columns for liquid chromatography.

Fiber-based monoliths for use in liquid chromatographic separations are defined by columns packed with aligned fibers, woven matrices, or contiguous fiber structures capable of achieving rapid separations of proteins, macromolecules, and low molecular weight components. A common denominator and motivating driver for this approach, first initiated 25 years ago, was reducing the cost of bioseparations in a manner that also reduced residence time of retained components while achieving a high ratio of mass to momentum transfer. This type of medium, when packed into a liquid chromatography column, minimized the fraction of stagnant liquid and resulted in a constant plate height for non-adsorbing species. The uncoupling of dispersion from eluent flow rate enabled the surface chemistry of the stationary phase to be considered separately from fluid transport phenomena and pointed to new ways to apply chemistry for the engineering of rapid bioseparations. This paper addresses developments and current research on fiber-based monoliths and explains how the various forms of this type of chromatographic stationary phase have potential to provide new tools for analytical and preparative scale separations. The different stationary phases are discussed, and a model that captures the observed constant plate height as a function of mobile phase velocity is reviewed. Methods that enable hydrodynamically stable fiber columns to be packed and operated over a range of mobile phase flow rates, together with the development of new fiber chemistries, are shown to provide columns that extend the versatility of liquid chromatography using monoliths, particularly at the preparative scale. Graphical Abstract Schematic representation of a sample mixture being separated by a rolled-stationary phase column, resulting separated peaks shown in the chromatogram.

Biological abatement of cellulase inhibitors.

Removal of enzyme inhibitors released during lignocellulose pretreatment is essential for economically feasible biofuel production. We tested bio-abatement to mitigate enzyme inhibitor effects observed in corn stover liquors after pretreatment with either dilute acid or liquid hot water at 10% (w/v) solids. Bio-abatement of liquors was followed by enzymatic hydrolysis of cellulose. To distinguish between inhibitor effects on enzymes and recalcitrance of the substrate, pretreated corn stover solids were removed and replaced with 1% (w/v) Solka Floc. Cellulose conversion in the presence of bio-abated liquors from dilute acid pretreatment was 8.6% (0.1x enzyme) and 16% (1x enzyme) higher than control (non-abated) samples. In the presence of bio-abated liquor from liquid hot water pretreated corn stover, 10% (0.1x enzyme) and 13% (1x enzyme) higher cellulose conversion was obtained compared to control. Bio-abatement yielded improved enzyme hydrolysis in the same range as that obtained using a chemical (overliming) method for mitigating inhibitors.