An evolutionarily stable strategy to colonize spatially extended habitats.

The ability of a species to colonize newly available habitats is crucial to its overall fitness1-3. In general, motility and fast expansion are expected to be beneficial for colonization and hence for the fitness of an organism4-7. Here we apply an evolution protocol to investigate phenotypical requirements for colonizing habitats of different sizes during range expansion by chemotaxing bacteria8. Contrary to the intuitive expectation that faster is better, we show that there is an optimal expansion speed for a given habitat size. Our analysis showed that this effect arises from interactions among pioneering cells at the front of the expanding population, and revealed a simple, evolutionarily stable strategy for colonizing a habitat of a specific size: to expand at a speed given by the product of the growth rate and the habitat size. These results illustrate stability-to-invasion as a powerful principle for the selection of phenotypes in complex ecological processes.

SRSF2 is required for mRNA splicing during spermatogenesis.

BACKGROUND: RNA splicing plays significant roles in fundamental biological activities. However, our knowledge about the roles of alternative splicing and underlying mechanisms during spermatogenesis is limited. RESULTS: Here, we report that Serine/arginine-rich splicing factor 2 (SRSF2), also known as SC35, plays critical roles in alternative splicing and male reproduction. Male germ cell-specific deletion of Srsf2 by Stra8-Cre caused complete infertility and defective spermatogenesis. Further analyses revealed that deletion of Srsf2 disrupted differentiation and meiosis initiation of spermatogonia. Mechanistically, by combining RNA-seq data with LACE-seq data, we showed that SRSF2 regulatory networks play critical roles in several major events including reproductive development, spermatogenesis, meiotic cell cycle, synapse organization, DNA recombination, chromosome segregation, and male sex differentiation. Furthermore, SRSF2 affected expression and alternative splicing of Stra8, Stag3 and Atr encoding critical factors for spermatogenesis in a direct manner. CONCLUSIONS: Taken together, our results demonstrate that SRSF2 has important functions in spermatogenesis and male fertility by regulating alternative splicing.

Exploiting spatial dimensions to enable parallelized continuous directed evolution.

Current strategies to improve the throughput of continuous directed evolution technologies often involve complex mechanical fluid-controlling system or robotic platforms, which limits their popularization and application in general laboratories. Inspired by our previous study on bacterial range expansion, in this study, we report a system termed SPACE for rapid and extensively parallelizable evolution of biomolecules by introducing spatial dimensions into the landmark phage-assisted continuous evolution system. Specifically, M13 phages and chemotactic Escherichia coli cells were closely inoculated onto a semisolid agar. The phages came into contact with the expanding front of the bacterial range, and then comigrated with the bacteria. This system leverages competition over space, wherein evolutionary progress is closely associated with the production of spatial patterns, allowing the emergence of improved or new protein functions. In a prototypical problem, SPACE remarkably simplified the process and evolved the promoter recognition of T7 RNA polymerase (RNAP) to a library of 96 random sequences in parallel. These results establish SPACE as a simple, easy to implement, and massively parallelizable platform for continuous directed evolution in general laboratories.

Data-driven quantitative modeling of bacterial active nematics.

Active matter comprises individual units that convert energy into mechanical motion. In many examples, such as bacterial systems and biofilament assays, constituent units are elongated and can give rise to local nematic orientational order. Such "active nematics" systems have attracted much attention from both theorists and experimentalists. However, despite intense research efforts, data-driven quantitative modeling has not been achieved, a situation mainly due to the lack of systematic experimental data and to the large number of parameters of current models. Here, we introduce an active nematics system made of swarming filamentous bacteria. We simultaneously measure orientation and velocity fields and show that the complex spatiotemporal dynamics of our system can be quantitatively reproduced by a type of microscopic model for active suspensions whose important parameters are all estimated from comprehensive experimental data. This provides unprecedented access to key effective parameters and mechanisms governing active nematics. Our approach is applicable to different types of dense suspensions and shows a path toward more quantitative active matter research.

A single mutation attenuates both the transcription termination and RNA-dependent RNA polymerase activity of T7 RNA polymerase.

Transcription termination is one of the least understood processes of gene expression. As the prototype model for transcription studies, the single-subunit T7 RNA polymerase (RNAP) is known to respond to two types of termination signals, but the mechanism underlying such termination, especially the specific elements of the polymerase involved, is still unclear, due to a lack of knowledge with respect to the structure of the termination complex. Here we applied phage-assisted continuous evolution to obtain variants of T7 RNAP that can bypass the typical class I T7 terminator with stem-loop structure. Through in vivo selection and in vitro characterization, we discovered a single mutation (S43Y) that significantly decreased the termination efficiency of T7 RNAP at all transcription terminators tested. Coincidently, the S43Y mutation almost eliminates the RNA-dependent RNAP (RdRp) activity of T7 RNAP without impeding the major DNA-dependent RNAP (DdRp) activity of the enzyme. S43 is located in a hinge region and regulates the transformation between transcription initiation and elongation of T7 RNAP. Steady-state kinetics analysis and an RNA binding assay indicate that the S43Y mutation increases the transcription efficiency while weakening RNA binding of the enzyme. As an enzymatic reagent for in vitro transcription, the T7 RNAP S43Y mutant reduces the undesired termination in run-off RNA synthesis and produces RNA with higher terminal homogeneity.

4-Hydroxy Pyridones from Heterologous Expression and Cultivation of the Native Host.

4-Hydroxy pyridones are a class of fungi-derived polyketide-nonribosomal peptide products featuring a core of 4-hydroxy-2-pyridone which have a wide range of biological activities. Genome mining of in-house strains using polyketide synthase-nonribosomal peptide synthase as a query identified an endophyte Tolypocladium sp. 49Y, which possesses a potential 4-hydroxy pyridone biosynthetic gene cluster. Heterologous expression in Aspergillus oryzae NSAR1 revealed that this gene cluster is functional and able to produce a rare type of 4-hydroxy pyridones called tolypyridones (compounds 3 and 4). Tolypocladium sp. 49Y was grown in a variety of media which led to the isolation of six 4-hydroxy pyridones (5-10) and one pyrrolidone (11) from a rice culture, and compounds 3 and 9 showed antifungal activity. These latter compounds are different from those obtained by heterologous expression. This study shows that both heterologous expression and cultivation of the native host are complementary approaches to discover new natural products.

Monitoring and Landscape Dynamic Analysis of Alpine Wetland Area Based on Multiple Algorithms: A Case Study of Zoige Plateau.

As an important part of the wetland ecosystem, alpine wetland is not only one of the most important ecological water conservation areas in the Qinghai-Tibet Plateau region, but is also an effective regulator of the local climate. In this study, using three machine learning algorithms to extract wetland, we employ the landscape ecological index to quantitatively analyze the evolution of landscape patterns and grey correlation to analyze the driving factors of Zoige wetland landscape pattern change from 1995 to 2020. The following results were obtained. (1) The random forest algorithm (RF) performs best when dealing with high-dimensional data, and the accuracy of the decision tree algorithm (DT) is better. The performance of the RF and DT is better than that of the support vector machine algorithm. (2) The alpine wetland in the study area was degraded from 1995 to 2015, whereas wetland area began to increase after 2015. (3) The results of landscape analysis show the decrease in wetland area from 1995 to 2005 was mainly due to the fragmentation of larger patches into many small patches and loss of the original small patches, while the 2005 to 2015 decrease was caused by the loss of many middle patches and the decrease in large patches from the edge to the middle. The 2015 to 2020 increase is due to an increase in the number of smaller patches and recovery of original wetland area. (4) The grey correlation degree further shows that precipitation and evaporation are the main factors leading to the change in the landscape pattern of Zoige alpine wetland. The results are of great significance to the long-term monitoring of the Zoige wetland ecosystem.

Single molecule identification of homology-dependent interactions between long ssRNA and dsDNA.

Long non-coding RNAs (lncRNAs) are prominently associated with chromosomes in an ever-increasing diversity of roles. To provide further insight into the potential nature of these associations, we have explored, for the first time, the interaction of long single-stranded (ss) RNAs with cognate homologous double-stranded (ds) DNA in vitro Using magnetic tweezers, we measured the effects of ssRNA on force extension curves for dsDNA. We observe that the presence of ssRNA impedes the extension of dsDNA, specifically at low forces, dependent on homology between the RNA and DNA species, and dependent on ssRNA lengths (≥1 kb). The observed effect also depends on the concentration of ssRNA and is abolished by overstretching of the dsDNA. These findings show that significant homologous contacts can occur between long ssRNA and dsDNA in the absence of protein and that these contacts alter the mechanical properties of the dsDNA. We propose that long ssRNA interacts paranemically with long dsDNA via periodic short homologous interactions, e.g. mediated by RNA/DNA triplex-formation, and that dsDNA extension is impeded by formation of RNA secondary structure in the intervening unbound regions. Analogous interactions in vivo would permit lncRNAs to mediate the juxtaposition of two or more DNA regions on the same or different chromosomes.

Interrogating the Escherichia coli cell cycle by cell dimension perturbations.

Bacteria tightly regulate and coordinate the various events in their cell cycles to duplicate themselves accurately and to control their cell sizes. Growth of Escherichia coli, in particular, follows a relation known as Schaechter's growth law. This law says that the average cell volume scales exponentially with growth rate, with a scaling exponent equal to the time from initiation of a round of DNA replication to the cell division at which the corresponding sister chromosomes segregate. Here, we sought to test the robustness of the growth law to systematic perturbations in cell dimensions achieved by varying the expression levels of mreB and ftsZ We found that decreasing the mreB level resulted in increased cell width, with little change in cell length, whereas decreasing the ftsZ level resulted in increased cell length. Furthermore, the time from replication termination to cell division increased with the perturbed dimension in both cases. Moreover, the growth law remained valid over a range of growth conditions and dimension perturbations. The growth law can be quantitatively interpreted as a consequence of a tight coupling of cell division to replication initiation. Thus, its robustness to perturbations in cell dimensions strongly supports models in which the timing of replication initiation governs that of cell division, and cell volume is the key phenomenological variable governing the timing of replication initiation. These conclusions are discussed in the context of our recently proposed "adder-per-origin" model, in which cells add a constant volume per origin between initiations and divide a constant time after initiation.

Effects of the supplementation of distillers' grape residues on ruminal degradability, whole tract digestibility and nitrogen metabolism in sheep.

Five ruminally and duodenally cannulated Kazakh male lambs (30 ± 2.75 kg) maintained singly in a metabolic cage were used in a 5 × 5 Latin square experiment to investigate the effect of supplementing a ration with five different levels of distillers' grape residue (DGR) on ruminal degradability, whole tract digestibility and nitrogen (N) metabolism of growing lambs. The rations were isoenergetic and isonitrogenous and contained 0, 3.85, 7.70, 11.55 and 15.41% DGR (DM basis). Each experimental period lasted for 18 d: 10 d for adaptation to the dietary treatment and 8 d for faecal, urinary, ruminal and duodenal digesta sample collections. The outflow rate of ruminal digesta increased (p = 0.032) linearly with the increased level of dietary neutral detergent fibre content, caused by the supplementation of DGR. As a result, the effective degradability of dry matter and crude protein decreased significantly with the treatments. Although the dietary intake of N, duodenal flow of total N, and the endogenous N at the duodenum were not affected by experimental treatments, N fractions in the digesta were altered. Ruminal microbial N decreased (p < 0.01) linearly; in contrast, ruminal un-degradable protein increased linearly (p < 0.01) in response to the increased addition of DGR. Although there was no significant difference in faecal N among treatments, N retention was increased linearly (p = 0.014), owing to the remarkable reduction (p = 0.016) of urinary N excretion with an increasing level of DGR. The results indicate that the DGR has some potential benefits of increasing the supply of bypass protein and of improving the utilisation efficiency of N for sheep. Therefore, the supplementation of DGR in ruminant feeding is recommended at levels not exceeding 10% of the diet.

S-Click Reaction for Isotropic Orientation of Oxidases on Electrodes to Promote Electron Transfer at Low Potentials.

Electrochemical sensors are essential for point-of-care testing (POCT) and wearable sensing devices. Establishing an efficient electron transfer route between redox enzymes and electrodes is key for converting enzyme-catalyzed reactions into electrochemical signals, and for the development of robust, sensitive, and selective biosensors. We demonstrate that the site-specific incorporation of a novel synthetic amino acid (2-amino-3-(4-mercaptophenyl)propanoic acid) into redox enzymes, followed by an S-click reaction to wire the enzyme to the electrode, facilitates electron transfer. The fabricated biosensor demonstrated real-time and selective monitoring of tryptophan (Trp) in blood and sweat samples, with a linear range of 0.02-0.8 mm. Further developments along this route may result in dramatic expansion of portable electrochemical sensors for diverse health-determination molecules.

The comparison of minimally invasive percutaneous nephrolithotomy and retrograde intrarenal surgery for stones larger than 2 cm in patients with a solitary kidney: a matched-pair analysis.

PURPOSE: To compare the treatment outcomes between retrograde intrarenal surgery (RIRS) and minimally invasive percutaneous nephrolithotomy (MPCNL) for the management of stones larger than 2 cm in patients with solitary kidney. METHODS: Between December 2012 and March 2014, 53 patients with a solitary kidney suffering from urinary stones larger than 2 cm were treated with RIRS. The outcomes of these patients were compared to a cohort of similar solitary kidney stone patients who underwent MPCNL using a matched-pair analysis (1:1 scenario). Cases were matched sequentially using the following criteria: size, number and location of stones, age, BMI, gender and previous ipsilateral open surgery. RESULTS: A stone-free rate (SFR) of 43.4 % was achieved after a single procedure in patients treated with RIRS and a SFR of 71.70 % in patients treated with MPCNL (p = 0.003). The operative time for RIRS was longer (p = 0.002), but the median hospital stay was shorter (p < 0.001). Average drop in hemoglobin level was comparable in both groups (9.30 vs. 10.85 g/L, p = 0.35). The transfusion rate as well as the incidence of postoperative complications such as fever and urosepsis was not statistically different between these two groups. Major complications (Clavien score 3a-4a) occurred in 3.77 and 1.89 %, RIRS and MPCNL, respectively (p = 1.000). CONCLUSIONS: Patients with a solitary kidney suffering from stones larger than 2 cm in size who undergo MPCNL had a higher SFR than RIRS. The complications were comparable in both groups. Even though RIRS patients spent less time in hospital, this procedure might not be an effective treatment as MPCNL in solitary kidneys with larger and multiple stones.

Rumen Degradability and Post-ruminal Digestion of Dry Matter, Nitrogen and Amino Acids of Three Protein Supplements.

This study evaluated the in situ ruminal degradability, and subsequent small intestinal digestibility (SID) of dry matter, crude protein (CP), and amino acids (AA) of cottonseed meal (CSM), sunflower seed meal (SFSM) and distillers dried grains with solubles (DDGS) by using the modified three-step in vitro procedure. The ruminal degradability and subsequent SID of AA in rumen-undegradable protein (RUP-AA) varied among three protein supplements. The result show that the effective degradability of DM for SFSM, CSM, and DDGS was 60.8%, 56.4%, and 41.0% and their ruminal fermentable organic matter was 60.0%, 55.9%, and 39.9%, respectively. The ruminal degradable protein (RDP) content in CP for SFSM, CSM, and DDGS was 68.3%, 39.0%, and 32.9%, respectively, at the ruminal solid passage rate of 1.84%/h. The SFSM is a good source of RDP for rumen micro-organisms; however, the SID of RUP of SFSM was lower. The DDGS and CSM are good sources of RUP for lambs to digest in the small intestine to complement ruminal microbial AA of growing lambs. Individual RUP-AA from each protein source was selectively removed by the rumen micro-organisms, especially for Trp, Arg, His, and Lys (p<0.01). The SID of individual RUP-AA was different within specific RUP origin (p<0.01). Limiting amino acid was Leu for RUP of CSM and Lys for both RUP of SFSM and DDGS, respectively. Therefore, different protein supplements with specific limitations should be selected and combined carefully in growing lambs ration to optimize AA balance.

RCOR1 is targeted by miR-23b-3p to modulate growth, colony formation, migration, and invasion of prostate cancer cells.

OBJECTIVES: Prostate cancer holds the second-highest incidence rate among all male malignancies, with a noticeable scarcity of effective treatment approaches. The REST Corepressor 1 (RCOR1) protein exhibits elevated expression across various tumors, acting as an oncogene. Nevertheless, its functions and mechanisms in prostate cancer have yet to be documented. While miR-23 demonstrates reduced expression in prostate cancer, the downstream genes it regulates remain unclear. METHODS: RT-qPCR and Western blotting assays were utilized to elucidate the mRNA and protein levels of miR-23b-3p and RCOR1. The luciferase reporter assay was employed to unveil the targeting relationship between miR-23b-3p and RCOR1. Additionally, a CCK-8 assay demonstrated cell growth, while colony formation and Transwell assays were performed to observe clone formation, cell migration, and invasion. RESULTS: In this study, we observed substantial mRNA and protein levels of RCOR1 in prostate cancer cells such as DU145, PC3, and LNCap. RCOR1 overexpression enhanced the growth, colony formation, migration, and invasion of prostate cancer cells, whereas genetic silencing of RCOR1 suppressed these processes. Bioinformatics analysis identified miR-23b-3p as a potential regulator of RCOR1, and luciferase assays validated RCOR1 as a downstream target of miR-23b-3p. Increasing miR-23b-3p mimics diminished RCOR1's mRNA and protein levels, while raising miR-23b-3p levels boosted RCOR1's expression. Moreover, the stimulatory impact of RCOR1 on prostate cancer cell development could be countered by elevating miR-23b-3p mimics. CONCLUSION: In summary, our findings confirm that RCOR1 is indeed under the influence of miR-23, shedding light on the miR-23/RCOR1 pathway's role in prostate cancer development. This offers novel theoretical and experimental support for comprehending the underlying mechanisms of prostate cancer and for targeted therapeutic avenues.

The Quantification of Bacterial Cell Size: Discrepancies Arise from Varied Quantification Methods.

The robust regulation of the cell cycle is critical for the survival and proliferation of bacteria. To gain a comprehensive understanding of the mechanisms regulating the bacterial cell cycle, it is essential to accurately quantify cell-cycle-related parameters and to uncover quantitative relationships. In this paper, we demonstrate that the quantification of cell size parameters using microscopic images can be influenced by software and by the parameter settings used. Remarkably, even if the consistent use of a particular software and specific parameter settings is maintained throughout a study, the type of software and the parameter settings can significantly impact the validation of quantitative relationships, such as the constant-initiation-mass hypothesis. Given these inherent characteristics of microscopic image-based quantification methods, it is recommended that conclusions be cross-validated using independent methods, especially when the conclusions are associated with cell size parameters that were obtained under different conditions. To this end, we presented a flexible workflow for simultaneously quantifying multiple bacterial cell-cycle-related parameters using microscope-independent methods.

Enabling technology and core theory of synthetic biology.

Synthetic biology provides a new paradigm for life science research ("build to learn") and opens the future journey of biotechnology ("build to use"). Here, we discuss advances of various principles and technologies in the mainstream of the enabling technology of synthetic biology, including synthesis and assembly of a genome, DNA storage, gene editing, molecular evolution and de novo design of function proteins, cell and gene circuit engineering, cell-free synthetic biology, artificial intelligence (AI)-aided synthetic biology, as well as biofoundries. We also introduce the concept of quantitative synthetic biology, which is guiding synthetic biology towards increased accuracy and predictability or the real rational design. We conclude that synthetic biology will establish its disciplinary system with the iterative development of enabling technologies and the maturity of the core theory.

SRSF1-mediated alternative splicing is required for spermatogenesis.

Alternative splicing (AS) plays significant roles in a multitude of fundamental biological activities. AS is prevalent in the testis, but the regulations of AS in spermatogenesis is only little explored. Here, we report that Serine/arginine-rich splicing factor 1 (SRSF1) plays critical roles in alternative splicing and male reproduction. Male germ cell-specific deletion of Srsf1 led to complete infertility by affecting spermatogenesis. Mechanistically, by combining RNA-seq data with LACE-seq data, we showed that SRSF1 affected the AS of Stra8 in a direct manner and Dazl, Dmc1, Mre11a, Syce2 and Rif1 in an indirect manner. Our findings demonstrate that SRSF1 has crucial functions in spermatogenesis and male fertility by regulating alternative splicing.

Stripe formation in bacterial systems with density-suppressed motility.

Engineered bacteria in which motility is reduced by local cell density generate periodic stripes of high and low density when spotted on agar plates. We study theoretically the origin and mechanism of this process in a kinetic model that includes growth and density-suppressed motility of the cells. The spreading of a region of immotile cells into an initially cell-free region is analyzed. From the calculated front profile we provide an analytic ansatz to determine the phase boundary between the stripe and the no-stripe phases. The influence of various parameters on the phase boundary is discussed.

A GWR downscaling method to reconstruct high-resolution precipitation dataset based on GSMaP-Gauge data: A case study in the Qilian Mountains, Northwest China.

Accurate precipitation data are crucial for hydrological, meteorological, and ecological research. However, it is difficult to obtain high-precision and high-resolution spatiotemporal distributions of precipitation in remote mountain regions with complex topography and sparse rain gauges. In addition, the spatial resolutions of existing satellite precipitation products are too coarse to apply them in the mountain regions with great spatiotemporal heterogeneity. To overcome the disadvantage, downscaling satellite precipitation products has been an effectively method to develop high-resolution precipitation data. In this study, a geographically weighted regression (GWR) model, coupling with topographical and water vapor source variables filtered by stepwise regression analysis (SRA), is applied to downscale the GSMaP-Gauge precipitation products (0.1° × 0.1°) to obtain high-resolution (1 km × 1 km) precipitation from 2000 to 2020 at annual, seasonal, and monthly scales over the Qilian Mountains. Besides, the accuracy of the downscaled precipitation based on all meteorological stations and the stations at high altitude (i.e., over 3000 m) are validated. Furtherly, the spatiotemporal variations of precipitation are analyzed. The results show that: (1) the accuracy after downscaling has been improved comparing with that of the original data. The accuracy of precipitation simulated at high-altitude stations is lower than that at all stations; (2) The trend of precipitation before and after downscaling is consistent in space. The spatial distributions of precipitation at annual, spring, summer, autumn, and months from March to November are decreased from the southeast to the northwest; (3) The spatial variations of precipitation show an increasing trend in most areas (>50%) at different time scales, except for March and September. Along with the time, the annual precipitation shows an increasing trend with a slope of 3.83 over the last 20 years. These findings suggest that the GWR method can be applied effectively to downscale annual, seasonal, and monthly precipitation of GSMaP-Gauge products in the Qilian Mountains.

Multiplex base editing to convert TAG into TAA codons in the human genome.

Whole-genome recoding has been shown to enable nonstandard amino acids, biocontainment and viral resistance in bacteria. Here we take the first steps to extend this to human cells demonstrating exceptional base editing to convert TAG to TAA for 33 essential genes via a single transfection, and examine base-editing genome-wide (observing ~40 C-to-T off-target events in essential gene exons). We also introduce GRIT, a computational tool for recoding. This demonstrates the feasibility of recoding, and highly multiplex editing in mammalian cells.