HUH Endonuclease: A Sequence-specific Fusion Protein Tag for Precise DNA-Protein Conjugation.

Synthetic DNA-protein conjugates have found widespread applications in diagnostics and therapeutics, prompting a growing interest in developing chemical biology methodologies for the precise and site-specific preparation of covalent DNA-protein conjugates. In this review article, we concentrate on techniques to achieve precise control over the structural and site-specific aspects of DNA-protein conjugates. We summarize conventional methods involving unnatural amino acids and self-labeling proteins, accompanied by a discussion of their potential limitations. Our primary focus is on introducing HUH endonuclease as a novel generation of fusion protein tags for DNA-protein conjugate preparation. The detailed conjugation mechanisms and structures of representative endonucleases are surveyed, showcasing their advantages as fusion protein tag in sequence selectivity, biological orthogonality, and no requirement for DNA modification. Additionally, we present the burgeoning applications of HUH-tag-based DNA-protein conjugates in protein assembly, biosensing, and gene editing. Furthermore, we delve into the future research directions of the HUH-tag, highlighting its significant potential for applications in the biomedical and DNA nanotechnology fields.

Stratification of dissolved organic matter in the upper 5000 m water column in the western Pacific Ocean.

Marine dissolved organic matter (DOM) is a principal reservoir involved in biogeochemical cycles and exerts a pivotal influence on global carbon flux dynamics. In this study, excitation-emission matrix fluorescence spectroscopy combined with parallel factor analysis (EEM-PARAFAC) was conducted on 230 DOM samples collected from 21 sites between February and April 2022 in the Western Pacific Ocean (WPO). We identified five distinct fluorescence peaks (peaks B, T, A, C, and M), predominantly protein-like and humic-like components. These findings, marked by significant differences (p < 0.01) in fluorescence intensities and spectral indices, characterized the transformation of DOM with ocean depth, illustrating a transition from active to recalcitrant forms. Additionally, random forest analysis (RFA) identified depth as a key factor influencing marine dissolved organic carbon (DOC), with a 32.59% importance value. Correlations between hydrological and fluorescent parameters underscored the complexity of DOM sources and influencing processes. Overall, this work broadens our understanding of DOM variability in the upper 5000 m of the WPO, enhancing our knowledge of the marine environment's role in the global carbon cycle.

Responses of biological traits of macrobenthic fauna to a eutrophication gradient in a semi-enclosed bay, China.

The impacts of eutrophication on benthic ecological functions are of increasing concern in recent years. In order to assess the response of macrobenthic fauna to increasing eutrophication, two field sampling surveys were conducted during the summer (July-August 2020) and autumn (October-November 2020) from offshore, nearshore to estuarine sediments in Bohai Bay, northern China. Biological trait analysis was employed for the assessment of macrofaunal samples. The results indicated that there was an increase in the proportion of benthic burrowering or tube-dwelling sediment feeders and taxa with higher larval dispersal ability, but a decrease in the proportion of taxa showing high motility in areas with higher nutrient levels. Seasonal differences were also noted in the shift in biological traits, with a significantly lower similarity among the sampling areas in summer and a higher proportion of carnivorous taxa in autumn. The findings suggested that long-term disturbance can lead to the dominance of smaller body-sized benthic species and reduced sediment quality, impeding ecological recovery of benthic organisms under such harsh environment.

A snapshot on vertical variability of dissolved organic matter in the epilagic zone of the eastern Indian Ocean.

Marine dissolved organic matter (DOM) plays an important role in aquatic ecosystems and is an essential reservoir of organic carbon in the marine carbon cycle. In this study, seawater DOM samples from 33 stations were collected in spring 2022 (April to May, 20 stations) and autumn 2020 (October to November, 13 stations) to better characterize and compare DOM variability within 200 m water layer in the eastern Indian Ocean (EIO). Hydrological parameters, nutrients and spectroscopic indices were calculated and evaluated for two cruises. In addition, excitation emission matrix spectroscopy combined with parallel factor analysis (EEM-PARAFAC) was used to directly analyse seawater DOM samples. The sources and processes of DOM in the EIO were assessed by fluorescence index (FI), freshness index (ß/α), Biological index (BIX), and humification index (HIX). Three fluorescent components were identified in DOM samples from two cruises, including: C1 (tryptophan- and tyrosine-like), C2 (marine and/or terrestrial humic-like), and C3 (terrestrial humic-like). The components of C1 accounted for 39.45 % ± 8.79 %, C2 for 33.05 % ± 6.42 %, and C3 for 27.20 % ± 4.47 % within 200 m water layer. The intensity of the DOM fluorescence seems to varied due to seasonal differences. In particular, the source of the DOM fraction varied at <100 m layer, which may also be related to the structure of the microbial community. Further, there is a strong correlation between the depth of seawater and hydrographic parameters, fluorescence indices and fluorescence components. Nutrients (nitrate, dissolved inorganic phosphate, and dissolved silicate) and humic-like fractions are more likely to accumulate in the deeper layers of the ocean. Thus, these results provide some data support for the variability of DOM fractions on a vertical scale in the EIO.

Applying causal discovery to single-cell analyses using CausalCell.

Correlation between objects is prone to occur coincidentally, and exploring correlation or association in most situations does not answer scientific questions rich in causality. Causal discovery (also called causal inference) infers causal interactions between objects from observational data. Reported causal discovery methods and single-cell datasets make applying causal discovery to single cells a promising direction. However, evaluating and choosing causal discovery methods and developing and performing proper workflow remain challenges. We report the workflow and platform CausalCell (http://www.gaemons.net/causalcell/causalDiscovery/) for performing single-cell causal discovery. The workflow/platform is developed upon benchmarking four kinds of causal discovery methods and is examined by analyzing multiple single-cell RNA-sequencing (scRNA-seq) datasets. Our results suggest that different situations need different methods and the constraint-based PC algorithm with kernel-based conditional independence tests work best in most situations. Related issues are discussed and tips for best practices are given. Inferred causal interactions in single cells provide valuable clues for investigating molecular interactions and gene regulations, identifying critical diagnostic and therapeutic targets, and designing experimental and clinical interventions.

Spatial and temporal changes in the assembly mechanism and co-occurrence network of the chromophytic phytoplankton communities in coastal ecosystems under anthropogenic influences.

As a typical semiclosed coastal sea area in China, the ecological environment of Bohai Bay has been significantly disturbed by human activities. As primary producers, the chromophytic phytoplankton are the basis of ecosystems, especially in coastal ecosystems, and changes in the chromophytic phytoplankton community can affect the stability of the entire ecosystem. In this study, we investigated the effects of the human activity-induced spatial and temporal environmental heterogeneity on the community composition, diversity, assembly mechanisms, and co-occurrence networks of chromophytic phytoplankton in Bohai Bay during the wet season and the dry season. The results showed that in both seasons, there was obvious environmental heterogeneity between the nearshore area and the offshore area, and the nearshore areas were more affected by human disturbance. Although higher diversity was supported by the abundance of nutrients in nearshore areas, co-occurrence network analysis revealed that the chromophytic phytoplankton were less closely connected to each other in nearshore areas than in offshore areas due to chemical oxygen demand (COD), eutrophication index (EI), and dissolved inorganic nitrogen (DIN). The nearshore network was less stable than the offshore co-occurrence network in both seasons, which was related to the concentration of dissolved oxygen and COD. Both stochastic and deterministic processes dominated the assembly of the chromophytic phytoplankton communities, with different importance rankings of stochastic and deterministic processes in the nearshore and offshore areas. Drift dominated the assembly of the communities in nearshore areas, while variable selection dominated the assembly of the communities in offshore areas. DIN, EI, and COD, rather than geographic distance, were the main environmental factors affecting the phylogenetic turnover of the chromophytic phytoplankton. Our study showed that environmental heterogeneity caused by human disturbance had a greater impact on the chromophytic phytoplankton communities in Bohai Bay than natural factors such as temperature and salinity.

Variations in dissolved organic matter chemistry on a vertical scale in the eastern Indian Ocean.

Oceans cover approximately 71% of the Earth's surface area, which is why some people refer to the Earth as a large water sphere. Marine dissolved organic matter (DOM) constitutes the main carbon pool for biogeochemical cycles and plays an important role in global carbon dynamics. Here, the molecular composition and component characteristics of surface (5 m), deep chlorophyll maximum (DCM), and deep (2000 m) layer DOM in the eastern Indian Ocean (EIO) were investigated using Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) and three-dimensional fluorescence spectroscopy. Thousands of individual DOM formulas (approximately 3716-6986 formulas) were detected at 100-700 Da, showing a Gaussian distribution. The elements carbon (C), hydrogen (H), oxygen (O), nitrogen (N) and sulfur (S) were detected and constituted four formula classes in solid-phase extracted marine DOM samples. Furthermore, the order of the percent intensity of the formulas was CHO > CHNO > CHOS > CHNOS. Carboxylic-rich alicyclic molecule (CRAM) compounds, as part of recalcitrant DOM (RDOM), were detected at 61.32%-78.77% (by intensity). In addition, the concept of islands of stability (IOS, approximately 3.99%-11.22%) has been proposed in this study, representing the most stable components in the marine environment. Such molecular formulas as described above probably contribute to increased RDOM content in the EIO and potentially reflect enhanced accumulation or sequestration of RDOM in the deep layer. The variation in the spectroscopic indices (FI, ß/α, BIX, and HIX) and fluorescent components (C1 to C4) with depth indicates a shift from protein-like to humic-like components, leading to gradual aging of the water column. In brief, this study relies on data from marine DOM in the EIO to provide a molecular and chemical background for global models of marine DOM production, transformation and sequestration.

Distribution, source identification and ecological effects of aerosol dissolved nutrients in the Bohai Bay.

The atmospheric aerosols around the Bohai Bay are affected intensively by the surrounding industrial, shipping and other human activities. Although atmospheric dry deposition is an important way for nutrients to enter the Bohai Bay, few studies explore the distribution patterns, source and deposition fluxes of typical nutrients in aerosols and their impacts on the marine ecosystem. This paper explored the spatial-temporal distribution of typical aerosol nutrients in summer and autumn, and their source and ecological effects were illustrated further. The mean concentration of dissolved total phosphorus (DTP), dissolved total nitrogen (DTN), dissolved organic nitrogen (DON), dissolved inorganic nitrogen (DIN), ammonium (NH4-N), nitrate (NO3-N), nitrite (NO2-N), silicate (SiO3-Si), phosphate (PO4-P), and dissolved organic phosphorus (DOP) were 31.22, 847.22, 288.19, 559.77, 288.19, 304.00, 253.65, 2.12, 15.74 and 15.48 nmol/m3, respectively, while their fluxes were corresponding to 0.61, 8.36, 2.52, 4.90, 1.41, 2.49, 0.02, 0.04, 0.19 and 0.26 mmol/(m2 month). Typical aerosol nutrient concentrations in autumn were mostly higher than those in summer, with high values occurring mainly in the central region. The potential sources of pollution were mainly concentrated in Shandong and Mongolia, and the sources of pollution were mainly agriculture, dust and industry. The large N:P and N:Si ratios in the dry deposition likely exacerbated Si and P limitation in the water column. These results provided the data basis for evaluating the pollution status and revealed that the dry deposition of aerosol nutrients should not be neglected by the ecological environment in the Bohai Bay.

Species and functional diversity of marine macrobenthic community and benthic habitat quality assessment in semi-enclosed waters upon recovering from eutrophication, Bohai Bay, China.

This study investigated the structure and function of macrobenthic community in Bohai Bay upon improvement of water quality due to pollution abatement. A total of 166 species were collected in the summer and autumn sampling, with an increase in sensitive species recorded as compared to data from previous studies. While historical variations in species richness indicated signs of improvement in community structure, results of functional diversity indices revealed that the macrobenthic community in Bohai Bay was still in an early stage of recovery. From BIO-ENV analysis, habitat instability may hinder how community responded to water quality improvement. Results of the benthic habitat quality assessment also indicated that the ecological status in most areas of Bohai Bay was classified as good, while a few estuarine regions were categorized in a poor status.

Fasim-LongTarget enables fast and accurate genome-wide lncRNA/DNA binding prediction.

Many long noncoding RNAs (lncRNAs) can bind to DNA sequences proximal and distal to abundant genes, thereby regulating gene expression by recruiting epigenomic modification enzymes to binding sites. Because a lncRNA's target genes scattering in a genome have correlated functions, epigenetic analyses should often be genome-wide on both genome and transcriptome levels. Multiple tools have been developed for predicting lncRNA/DNA binding, but fast and accurate genome-wide prediction remains a challenge. Here we report Fasim-LongTarget (a revised version of LongTarget), compare its performance with TDF and LongTarget using the experimental data of the lncRNA MEG3, NEAT1, and MALAT1, and describe a case of genome-wide prediction. Fasim-LongTarget is as accurate as LongTarget and more accurate than TDF and is 200 times faster than LongTarget, making accurate genome-wide prediction feasible. The code is available on the Github website (https://github.com/LongTarget/Fasim-LongTarget), and the online service is available on the LongTarget website (https://lncRNA.smu.edu.cn).

Pipelines for cross-species and genome-wide prediction of long noncoding RNA binding.

Abundant long, noncoding RNAs (lncRNAs) in mammals can bind to DNA sequences and recruit histone- and DNA-modifying enzymes to binding sites to epigenetically regulate target genes. However, most lncRNAs' binding motifs and target sites are unknown. The large numbers of lncRNAs and target sites in the whole genome make it infeasible to examine lncRNA binding to DNA purely experimentally. Here, we report a protocol for lncRNA/DNA-binding analysis that is built upon a database containing the GENCODE-annotated human and mouse lncRNAs, the orthologs of these lncRNAs in 17 mammals, and the genome sequences of the 17 mammals. Cross-species and genome-wide lncRNA/DNA-binding analysis begins with and is driven by database search. The predicted DNA-binding motifs and binding sites answer the general question of which lncRNAs may epigenetically regulate which genes, and can be used to identify potential sites for genome and epigenome editing. To use the protocol, preliminary knowledge of the base-pairing rules that guide the binding of noncoding RNAs to DNA to form triplexes, as well as the skills required to use the UCSC Genome Browser, are needed. A genome-wide prediction takes from 2 to 10 d, and the results are sent to users automatically by e-mail. The platform is updated continuously, making it possible to study more lncRNAs and larger genomic regions in less computational time.