Porcine epidemic diarrhea (PED) caused by porcine epidemic diarrhea virus (PEDV), is an acute and highly infectious disease, resulting in substantial economic losses in the pig industry. Given that PEDV primarily infects the mucosal surfaces of the intestinal tract, it is crucial to improve the mucosal immunity to prevent viral invasion. Lactic acid bacteria (LAB) oral vaccines offer unique advantages and potential applications in combatting mucosal infectious diseases, making them an ideal approach for controlling PED outbreaks. However, traditional LAB oral vaccines use plasmids for exogenous protein expression and antibiotic genes as selection markers. Antibiotic genes can be diffused through transposition, transfer, or homologous recombination, resulting in the generation of drug-resistant strains. To overcome these issues, genome-editing technology has been developed to achieve gene expression in LAB genomes. In this study, we used the CRISPR-NCas9 system to integrate the PEDV S1 gene into the genome of alanine racemase-deficient Lactobacillus paracasei △Alr HLJ-27 (L. paracasei △Alr HLJ-27) at the thymidylate synthase (thyA) site, generating a strain, S1/△Alr HLJ-27. We conducted immunization assays in mice and piglets to evaluate the level of immune response and evaluated its protective effect against PEDV through challenge tests in piglets. Oral administration of the strain S1/△Alr HLJ-27 in mice and piglets elicited mucosal, humoral, and cellular immune responses. The strain also exhibited a certain level of resistance against PEDV infection in piglets. These results demonstrate the potential of S1/△Alr HLJ-27 as an oral vaccine candidate for PEDV control. KEY POINTS: • A strain S1/△Alr HLJ-27 was constructed as the candidate for an oral vaccine. • Immunogenicity response and challenge test was carried out to analyze the ability of the strain. • The strain S1/△Alr HLJ-27 could provide protection for piglets to a certain extent.

We examine the coordinated behavior of thousands of genes in cell fate transitions through genome expression as an integrated dynamical system using the concepts of self-organized criticality and coherent stochastic behavior. To quantify the effects of the collective behavior of genes, we adopted the flux balance approach and developed it in a new tool termed expression flux analysis (EFA). Here we describe this tool and demonstrate how its application to specific experimental genome-wide expression data provides new insights into the dynamics of the cell-fate transitions. Particularly, we show that in cell fate change, specific stochastic perturbations can spread over the entire system to guide distinct cell fate transitions through switching cyclic flux flow in the genome engine. Utilization of EFA enables us to elucidate a unified genomic mechanism for when and how cell-fate change occurs through critical transitions.

Herein, we provide a brief overview of complex systems theory approaches to investigate the genomic mechanism of cell-fate changes. Cell trajectories across the epigenetic landscape, whether in development, environmental responses, or disease progression, are controlled by extensively coordinated genome-wide gene expression changes. The elucidation of the mechanisms underlying these coherent expression changes is of fundamental importance in cell biology and for paving the road to new therapeutic approaches. In previous studies, we pointed at dynamic criticality as a plausible characteristic of genome-wide transition dynamics guiding cell fate. Whole-genome expression develops an engine-like organization (genome engine) in order to establish an autonomous dynamical system, capable of both homeostasis and transition behaviors. A critical set of genes behaves as a critical point (CP) that serves as the organizing center of cell-fate change. When the system is pushed away from homeostasis, the state change that occurs at the CP makes local perturbation spread over the genome, demonstrating self-organized critical (SOC) control of genome expression. Oscillating-Mode genes (which normally keep genome expression on pace with microenvironment fluctuations), when in the presence of an effective perturbative stimulus, drive the dynamics of synchronization, and thus guide the cell-fate transition.

Acinetobacter baylyi is an interesting model organism to investigate bacterial metabolism due to its vast repertoire of metabolic enzymes and ease of genetic manipulation. However, the study of gene expression in vitro is dependent on the availability of its RNA polymerase (RNAp), an essential enzyme in transcription. In this work, we developed a convenient method of producing the recombinant A. baylyi ADP1 RNA polymerase holoenzyme (RNApholo) in E. coli that yields 22 mg of a >96% purity protein from a 1-liter shake flask culture. We further characterized the A. baylyi ADP1 RNApholo kinetic profile using T7 Phage DNA as template and demonstrated that it is a highly transcriptionally active enzyme with an elongation rate of 24 nt/s and a termination efficiency of 94%. Moreover, the A. baylyi ADP1 RNApholo has a substantial sequence identity (∼95%) with the RNApholo from the human pathogen Acinetobacter baumannii. This protein can serve as a source of material for structural and biological studies towards advancing our understanding of genome expression and regulation in Acinetobacter species.

In the last fifty years, large efforts have been deployed in basic research, clinical oncology, and clinical trials, yielding an enormous amount of information regarding the molecular mechanisms of cancer and the design of effective therapies. The knowledge that has accumulated underpins the complexity, multifactoriality, and heterogeneity of cancer, disclosing novel landscapes in cancer biology with a key role of genome plasticity. Here, we propose that cancer onset and progression are determined by a stress-responsive epigenetic mechanism, resulting from the convergence of upregulation of LINE-1 (long interspersed nuclear element 1), the largest family of human retrotransposons, genome damage, nuclear lamina fragmentation, chromatin remodeling, genome reprogramming, and autophagy activation. The upregulated expression of LINE-1 retrotransposons and their protein products plays a key role in these processes, yielding an increased plasticity of the nuclear architecture with the ensuing reprogramming of global gene expression, including the reactivation of embryonic transcription profiles. Cancer phenotypes would thus emerge as a consequence of the unscheduled reactivation of embryonic gene expression patterns in an inappropriate context, triggering de-differentiation and aberrant proliferation in differentiated cells. Depending on the intensity of the stressing stimuli and the level of LINE-1 response, diverse degrees of malignity would be generated.

The purpose of our studies is to elucidate the nature of massive control of the whole genome expression with a particular emphasis on cell-fate change. The whole genome expression is coordinated by the emergence of a critical point (CP: a peculiar set of biphasic genes) with the genome acting as an integrated dynamical system. In response to stimuli, the genome expression self-organizes into local sub-, near-, and super-critical states, each exhibiting distinct collective behaviors with its center of mass acting as a local attractor, coexisting with the whole genome attractor (GA). The CP serves as the organizing center of cell-fate change, and its activation makes local perturbation to spread over the genome affecting GA. The activation of CP is in turn elicited by genes with elevated temporal variance (oscillating-mode genes), normally in charge to keep genome expression at pace with microenvironment fluctuations. When oscillation exceeds a given threshold, the CP synchronizes with the GA driving genome expression state transition. The expression synchronization wave invading the entire genome is fostered by the fusion-splitting dynamics of silencing pericentromere-associated heterochromatin domains and the consequent folding-unfolding transitions of transcribing euchromatin domains. The proposed mechanism is a unified step toward a time-evolutional transition theory of biological regulation.

The freshwater sponge Ephydatia muelleri and its Chlorella-like algal partner is an emerging model for studying animal: algal endosymbiosis. The sponge host is a tractable laboratory organism, and the symbiotic algae are easily cultured. We took advantage of these traits to interrogate questions about mechanisms that govern the establishment of durable intracellular partnerships between hosts and symbionts in facultative symbioses. We modified a classical experimental approach to discern the phagocytotic mechanisms that might be co-opted to permit persistent infections, and identified genes differentially expressed in sponges early in the establishment of endosymbiosis. We exposed algal-free E. muelleri to live native algal symbionts and potential food items (bacteria and native heat-killed algae), and performed RNA-Seq to compare patterns of gene expression among treatments. We found a relatively small but interesting suite of genes that are differentially expressed in the host exposed to live algal symbionts, and a larger number of genes triggered by host exposure to heat-killed algae. The upregulated genes in sponges exposed to live algal symbionts were mostly involved in endocytosis, ion transport, metabolic processes, vesicle-mediated transport, and oxidation-reduction. One of the host genes, an ATP-Binding Cassette transporter that is downregulated in response to live algal symbionts, was further evaluated for its possible role in the establishment of the symbiosis. We discuss the gene expression profiles associated with host responses to living algal cells in the context of conditions necessary for long-term residency within host cells by phototrophic symbionts as well as the genetic responses to sponge phagocytosis and immune-driven pathways.

Garlic (Allium sativum) is an important vegetable crop that is widely used in cooking and medicine. The greening phenomenon of garlic severely decreases the quality of garlic and hinders garlic processing. To study the mechanism of garlic greening, comprehensive full-length transcript sets were constructed. We detected the differences in greening between Pizhou (PZ) garlic and Laiwu (LW) garlic that were both stored at -2.5°C and protected from light at the same time. The results showed that 60,087 unigenes were respectively annotated to the NR, KEGG, GO, Pfam, eggNOG and Swiss Prot databases, and a total of 30,082 unigenes were annotated. The analysis of differential genes and differential proteins showed that PZ garlic and LW garlic had 923 differentially expressed genes (DEGs), of which 529 genes were up regulated and 394 genes were downregulated. Through KEGG and GO enrichment analysis, it was found that the most significant way of enriching DEGs was the phenylpropane metabolic pathway. Proteomics analysis found that there were 188 differentially expressed proteins (DAPs), 162 up-regulated proteins, and 26 down-regulated proteins between PZ garlic and LW garlic. The content of 10 proteins related to phenylpropanoid biosynthesis in PZ garlic was significantly higher than that of LW garlic. This study explored the mechanisms of garlic greening at a molecular level and further discovered that the formation of garlic green pigment was affected significantly by the phenylpropanoid metabolic pathway. This work provided a theoretical basis for the maintenance of garlic quality during garlic processing and the future development of the garlic processing industries.

Elucidation of the genomic mechanism that guides the cell-fate change is one of the fundamental issues of biology. We previously demonstrated that whole genome expression is coordinated by the emergence of a critical point at both the cell-population and single-cell levels through the physical principle of self-organized criticality. In this paper, we further examine the genomic mechanism that determines the cell-fate changes from embryo to cancer development. The state of the critical point, acting as the organizing center of the cell fate, determines whether the genome resides in a super- or sub-critical state. In the super-critical state, a specific stochastic perturbation can spread over the entire system through the \"genome engine\", an autonomous critical-control genomic system, whereas in the sub-critical state, the perturbation remains at a local level. The cell-fate changes when the genome becomes super-critical. We provide a consistent framework to develop a time-evolutional transition theory for the biological regulation of the cell-fate change.

The view of the cell nucleus has evolved from an isolated, static organelle to a dynamic structure integrated with other mechanical elements of the cell. Both dynamics and integration appear to contribute to a mechanical regulation of genome expression. Here, we review physical structures inside the nucleus at different length scales and the dynamic reorganization modulated by cellular forces. First, we discuss nuclear organization focusing on self-assembly and disassembly of DNA structures and various nuclear bodies. We then discuss the importance of connections from the chromatin fiber through the nuclear envelope to the rest of the cell as they relate to mechanobiology. Finally, we discuss how cell stimulation, both chemical and physical, can alter nuclear structures and ultimately cellular function in healthy cells and in some model diseases. The view of chromatin and nuclear bodies as mechanical entities integrated with force generation from the cytoskeleton combines polymer physics with cell biology and medicine.