SUMOylation Research Articles

SUMO enhances unfolding of SUMO–polyubiquitin-modified substrates by the Ufd1/Npl4/Cdc48 complex

The Ufd1/Npl4/Cdc48 complex is a universal protein segregase that plays key roles in eukaryotic cellular processes. Its functions orchestrating the clearance or removal of polyubiquitylated targets are established; however, prior studies suggest that the complex also targets substrates modified by the ubiquitin-like protein SUMO. Here, we show that interactions between Ufd1 and SUMO enhance unfolding of substrates modified by SUMO-polyubiquitin hybrid chains by the budding yeast Ufd1/Npl4/Cdc48 complex compared to substrates modified by polyubiquitin chains, a difference that is accentuated when the complex has a choice between these substrates. Incubating Ufd1/Npl4/Cdc48 with a substrate modified by a SUMO-polyubiquitin hybrid chain produced a series of single-particle cryo-EM structures that reveal features of interactions between Ufd1/Npl4/Cdc48 and ubiquitin prior to and during unfolding of ubiquitin. These results are consistent with cellular functions for SUMO and ubiquitin modifications and support a physical model wherein Ufd1/Npl4/Cdc48, SUMO, and ubiquitin conjugation pathways converge to promote clearance of proteins modified with SUMO and polyubiquitin.

SENP1 induced protein deSUMO modification increased the chemotherapy sensitivity of endometrial cancer side population cells

Objective: To inhibit the stemness maintenance potential of endometrial cancer and increase the sensitivity of endometrial cancer side population cells to chemotherapy drugs by inducing extensive deSUMOylation modification of proteins. Methods: Flow cytometry was used to sort and culture CD133(+) CD44(+) KLE endometrial cancer cell clone spheres. Protein expression level of small ubiquitin-related modifier 1 (SUMO1) and two stemness maintenance genes of tumor side population cells, octamer binding transcription factor-4 (Oct4) and sex determining region Y-box2 (Sox2), were detected by western blotting method. Lentivirus-mediated Sentrin/SUMO-specific proteases 1 (SENP1) gene was stably transfected into KLE side population cells. Western blotting was used to detect the protein expressions of SENP1, SUMO1, Oct4 and Sox2. The clone formation rate was compared between KLE side population cells with or without SENP1 overexpression. Flow cytometry was applied to detect cell cycle changes. 3-(4, 5-Dimethylthiazole-2)-2, 5-diphenyl-tetrazolium bromide (MTT) experiment and flow cytometry apoptosis method were used to detect the chemosensitivity of the side population of endometrial cancer cells to cisplatin. Tumor-bearing mouse models of endometrial cancer were established to detect the effect of SENP1 overexpression on the chemotherapy sensitivity of cisplatin. Results: Compared with CD133(-)CD44(-) KLE cells, CD133(+) CD44(+) KLE side population cells could form clonal spheres and express higher levels of SUMO1, Oct4 and Sox2 proteins (P<0.05). Compared with KLE side population cells that were not transfected with SENP1 gene, the expression level of SENP1 protein in KLE side population cells overexpressing SUMO1、Oct4 and Sox2 were lower. The clonal sphere formation rate was reduced from (25.67±5.44)% to (7.46±1.42)%, and cell cycle shifted from G(0)/G(1) phase to G(2) phase. IC(50) of cisplatin decreased from (55.46±6.14) μg/ml to (11.55±3.12) μg/ml, and cell apoptosis rate increased from (9.76±2.09)% to (16.79±3.44)%. Overexpression of SENP1 could reduce the tumorigenesis rate of KLE side population cells in vivo and increase their chemotherapy sensitivity to cisplatin (P<0.05). Conclusion: Overexpression of SENP1 can induce protein deSUMOylation modification, inhibit the stemness maintenance potential of endometrial cancer side population cells, and enhance their chemotherapy sensitivity, which provides a new reference for gene therapy of endometrial cancer.

SUMO1 regulates post-infarct cardiac repair based on cellular heterogeneity

Small ubiquitin-related modifier (SUMOylation) is a dynamic post-translational modification that maintains cardiac function and can protect against a hypertrophic response to cardiac pressure overload. However, the function of SUMOylation after myocardial infarction (MI) and the molecular details of heart cell responses to SUMO1 deficiency have not been determined. In this study, we demonstrated that SUMO1 protein was inconsistently abundant in different cell types and heart regions after MI. However, SUMO1 knockout significantly exacerbated systolic dysfunction and infarct size after myocardial injury. Single-nucleus RNA sequencing revealed the differential role of SUMO1 in regulating heart cells. Among cardiomyocytes, SUMO1 deletion increased the Nppa+Nppb+Ankrd1+ cardiomyocyte subcluster proportion after MI. In addition, the conversion of fibroblasts to myofibroblasts subclusters was inhibited in SUMO1 knockout mice. Importantly, SUMO1 loss promoted proliferation of endothelial cell subsets with the ability to reconstitute neovascularization and expressed angiogenesis-related genes. Computational analysis of ligand/receptor interactions suggested putative pathways that mediate cardiomyocytes to endothelial cell communication in the myocardium. Mice preinjected with cardiomyocyte-specific AAV-SUMO1, but not the endothelial cell-specific form, and exhibited ameliorated cardiac remodeling following MI. Collectively, our results identified the role of SUMO1 in cardiomyocytes, fibroblasts, and endothelial cells after MI. These findings provide new insights into SUMO1 involvement in the pathogenesis of MI and reveal novel therapeutic targets.

SUMO E3 ligase AtMMS21-dependent SUMOylation of AUXIN/INDOLE-3-ACETIC ACID 17 regulates auxin signaling.

Changes in plant auxin levels can be perceived and converted into cellular responses by auxin signal transduction. AUXIN/INDOLE-3-ACETIC ACID (Aux/IAA) proteins are auxin transcriptional inhibitors that play important roles in regulating auxin signal transduction. The stability of Aux/IAA proteins is important for transcription initiation and downstream auxin-related gene expression. Here, we report that the Aux/IAA protein IAA17 interacts with the small ubiquitin-related modifier (SUMO) E3 ligase METHYL METHANESULFONATE-SENSITIVE 21 (AtMMS21) in Arabidopsis (Arabidopsis thaliana). AtMMS21 regulated the SUMOylation of IAA17 at the K41 site. Notably, root length was suppressed in plants overexpressing IAA17, whereas the roots of K41-mutated IAA17 transgenic plants were not significantly different from wild-type roots. Biochemical data indicated that K41-mutated IAA17 or IAA17 in the AtMMS21 knockout mutant was more likely to be degraded compared with nonmutated IAA17 in wild-type plants. In conclusion, our data revealed a role for SUMOylation in the maintenance of IAA17 protein stability, which contributes to improving our understanding of the mechanisms of auxin signaling.

Biểu hiện gen fliC cải biến của Salmonella Dublin và tinh sạch flagellin tái tổ hợp

Flagellin is a potent trigger of innate immune responses that enhance adaptive immune responses. Flagellin has intrinsic adjuvant activity mediated through toll-like receptor (TLR 5) and is an attractive candidate for highly effective vaccine adjuvant. The N-terminal D0-D1 domain of flagellin is required for TLR5 recognition. In the present study, fliC gene from S. Dublin expressed in E. coli BL21 in the fusion form with SUMO protein. The 48 kDa fusion protein of SUMO-flagellin was synthesised in insoluble inclusion body. The overall yield of expressed SUMO - flagellin was 0.6 g/L. The recombinant protein solubilized in 2 M urea then was purified by using Ni-NTA column chromatography with purity about 95.4%. Flagellin will be cleaved from fusion protein and used to develop as a radiation countermeasure.

SUMO pathway is required for ribosome biogenesis

Ribosomes, acting as the cellular factories for protein production, are essential for all living organisms. Ribosomes are composed of both proteins and RNAs and are established through the coordination of several steps, including transcription, maturation of ribosomal RNA (rRNA), and assembly of ribosomal proteins. In particular, diverse factors required for ribosome biogenesis, such as transcription factors, small nucleolar RNA (snoRNA)-associated proteins, and assembly factors, are tightly regulated by various post-translational modifications. Among these modifications, small ubiquitin-related modifier (SUMO) targets lots of proteins required for gene expression of ribosomal proteins, rRNA, and snoRNAs, rRNA processing, and ribosome assembly. The tight control of SUMOylation affects functions and locations of substrates. This review summarizes current studies and recent progress of SUMOylation-mediated regulation of ribosome biogenesis. [BMB Reports 2022; 55(11): 535-540].

Bioinformatic Tools for Exploring the SUMO Gene Network: An Update.

Plant sumoylation research has seen significant advances in recent years, particularly since high-throughput proteomic strategies have enabled the discovery of more than one thousand SUMO targets. In the present chapter, we update the previously reported SUMO (small ubiquitin-related modifier) gene network (SGN) to its v4 iteration. SGN is a curated assembly of Arabidopsis thaliana genes that have been functionally associated with sumoylation, from SUMO pathway components to targets and interactors. The enclosed tutorial helps interpret and manage these datasets and details bioinformatic tools that can be used for in silico-based hypothesis generation. The latter include tools for sumoylation site prediction, comparative genomics, and gene network analysis.

SUMO Conjugation and SUMO Chain Formation by Plant Enzymes.

SUMO conjugation is a conserved process of eukaryotes, and essential in metazoa. Similar to ubiquitylation, a SUMO-activating enzyme links to the SUMO carboxyl-terminal Gly in a thioester bond, and a SUMO-conjugating enzyme accepts activated SUMO and can transfer it to substrates. Unlike ubiquitylation, this transfer can also occur, in an unspecified number of cases, in the absence of ligase-like enzymes. Different isoforms of SUMO are present in eukaryotic genomes. Saccharomyces cerevisiae has only one SUMO protein, humans have four, and Arabidopsis thaliana has eight, the main isoforms being SUMO1 and SUMO2 with about 95% identity. Functionally similar to human SUMO2 and SUMO3, Arabidopsis SUMO1 and 2 can be transferred to substrates as single moieties, but can also form SUMO chains, a process enhanced by chain-forming ligases. By combined action with SUMO chain recognizing ubiquitin ligases, chains can channel substrates into the ubiquitin-dependent degradation pathway.A method is described to sumoylate substrates and to generate SUMO chains, using plant enzymes produced in E. coli. In vitro SUMO chain formation may serve for further analysis of SUMO chain functions. It can also provide an easy-to-synthesize substrate for SUMO-specific proteases.

Promoting soluble expression of hybrid mussel foot proteins by SUMO-TrxA tags for production of mussel glue

Mussel foot proteins (Mfps) display application potential with strong adhesion, enabling mussels to adhere firmly to various surfaces. Mytilus galloprovincialis foot protein 3B (Mgfp-3B) exhibits this characteristic remarkably. However, it remains a challenge for further research due to the low soluble expression of heterologous production. In this study, a small ubiquitin-related modifier (SUMO) and thioredoxin A (TrxA), which catalyzed the proper folding of disulfide bridges, were selected to increase the soluble expression of mfps. An additional ribosome binding site was introduced between the molecular chaperones and Mgfp-3B (fp-3) to form a bicistronic translation-coupled expression vector for co-expression. The results revealed that the combination of SUMO-TrxA increased the soluble expression of fp-3 by 18.07 %. Furthermore, the SUMO-TrxA also boosted the soluble expression of hybrid mfps Mgfp-3B-Mfp-1 (fp-3-1) by 11.29 %, Mgfp-3B-Mgfp-3B (fp-3-3) by 19.91 %, and Mgfp-3B-Mgfp-5 (fp-3-5) by 14.03 %. Ultimately, by high cell density cultivation in a 5 L bioreactor, the yields of fp-3, fp-3-3, and fp-3-5 co-expressed with SUMO-TrxA reached 217.75 mg/L, 127.2 mg/L, and 97.28 mg/L, respectively. Consequently, soluble production of mfps holds great potential for the sustainable supply of protein adhesive materials.

Interaction of FOXO1 and SUMOylated PPARγ1 induced by hyperlipidemia and hyperglycemia favors vascular endothelial insulin resistance and dysfunction

PPARγ1 and FOXO1 are the key transcription factors that regulate insulin sensitivity. We previously found that a small ubiquitin-related modifier of PPARγ1 at K77 (SUMOylation) favored endothelial insulin resistance (IR) induced by high fat/high glucose (HF/HG) administration. However, whether and how the crosstalk between SUMOylated PPARγ1 and FOXO1 would mediate the development of the endothelial IR and dysfunction remains unclear. Here, we emphasize how PPARγ1-K77 SUMOylation would interact with FOXO1 and participate in the development of the endothelial IR and dysfunction. Our results show that the combination of HF/HG and PPARγ1-K77 SUMOylation exhibits a synergistic deteriorative effect on the endothelial IR and dysfunction, presenting decreased NO levels and elevated ET-1 levels, weakened PI3K/Akt/eNOS signaling, and impaired endothelium-dependent vasodilation function. The further researches reveal that PPARγ1-K77 SUMOylation readily interacts with FOXO1, and FOXO1 occupies the PPAR response element (PPRE) which is supposed to be occupied by PPARγ, thus resulting in the decrease of PPARγ1 transcription activity and the mitigation of the PI3K/Akt signaling. Moreover, the mitigation of the PI3K/Akt signaling promotes in turn the accumulation of FOXO1 in the nucleus where FOXO1 interacts with the SUMOylated PPARγ1, thus exerting a positive feedback effect on IR pathogenesis. The findings uncover a novel association between PPARγ1-K77 SUMOylation and FOXO1, which contributes to our understanding of the pathogenesis of endothelial IR and dysfunction and provides novel pharmacological targets for diabetic angiopathy.

Profiling Substrate Specificity of the SUMO Protease Ulp1 by the YESS-PSSC System to Advance the Conserved Mechanism for Substrate Cleavage.

SUMO modification is a vital post-translational regulation process in eukaryotes, in which the SUMO protease is responsible for the maturation of the SUMO precursor and the deconjugation of the SUMO protein from modified proteins by accurately cleaving behind the C-terminal Gly–Gly motif. To promote the understanding of the high specificity of the SUMO protease against the SUMO protein as well as to clarify whether the conserved Gly–Gly motif is strictly required for the processing of the SUMO precursor, we systematically profiled the specificity of the S. cerevisiae SUMO protease (Ulp1) on Smt3 at the P2–P1↓P1’ (Gly–Gly↓Ala) position using the YESS–PSSC system. Our results demonstrated that Ulp1 was able to cleave Gly–Gly↓ motif-mutated substrates, indicating that the diglycine motif is not strictly required for Ulp1 cleavage. A structural-modeling analysis indicated that it is the special tapered active pocket of Ulp1 conferred the selectivity of small residues at the P1–P2 position of Smt3, such as Gly, Ala, Ser and Cys, and only which can smoothly deliver the scissile bond into the active site for cleavage. Meanwhile, the P1’ position Ala of Smt3 was found to play a vital role in maintaining Ulp1’s precise cleavage after the Gly–Gly motif and replacing Ala with Gly in this position could expand Ulp1 inclusivity against the P1 and P2 position residues of Smt3. All in all, our studies advanced the traditional knowledge of the SUMO protein, which may provide potential directions for the drug discovery of abnormal SUMOylation-related diseases.

Discovery of a Dual SENP1 and SENP2 Inhibitor.

SUMOylation is a reversible post–translational modification (PTM) involving covalent attachment of small ubiquitin-related modifier (SUMO) proteins to substrate proteins. Dysregulation of SUMOylation and deSUMOylation results in cellular malfunction and is linked to various diseases, such as cancer. Sentrin-specific proteases (SENPs) were identified for the maturation of SUMOs and the deconjugation of SUMOs from their substrate proteins. Hence, this is a promising target tackling the dysregulation of the SUMOylation process. Herein, we report the discovery of a novel protein-protein interaction (PPI) inhibitor for SENP1-SUMO1 by virtual screening and subsequent medicinal chemistry optimization of the hit molecule. The optimized inhibitor ZHAWOC8697 showed IC50 values of 8.6 μM against SENP1 and 2.3 μM against SENP2. With a photo affinity probe the SENP target was validated. This novel SENP inhibitor represents a new valuable tool for the study of SUMOylation processes and the SENP-associated development of small molecule-based treatment options.

Expression strategies for the efficient synthesis of antimicrobial peptides in plastids

Antimicrobial peptides (AMPs) kill microbes or inhibit their growth and are promising next-generation antibiotics. Harnessing their full potential as antimicrobial agents will require methods for cost-effective large-scale production and purification. Here, we explore the possibility to exploit the high protein synthesis capacity of the chloroplast to produce AMPs in plants. Generating a large series of 29 sets of transplastomic tobacco plants expressing nine different AMPs as fusion proteins, we show that high-level constitutive AMP expression results in deleterious plant phenotypes. However, by utilizing inducible expression and fusions to the cleavable carrier protein SUMO, the cytotoxic effects of AMPs and fused AMPs are alleviated and plants with wild-type-like phenotypes are obtained. Importantly, purified AMP fusion proteins display antimicrobial activity independently of proteolytic removal of the carrier. Our work provides expression strategies for the synthesis of toxic polypeptides in chloroplasts, and establishes transplastomic plants as efficient production platform for antimicrobial peptides.

Small-molecule SUMO inhibition for biomarker-informed B-cell lymphoma therapy.

Aberrant activity of the SUMOylation pathway has been associated with MYC overexpression and poor prognosis in aggressive B-cell lymphoma (BCL) and other malignancies. Recently developed small-molecule inhibitors of SUMOylation (SUMOi) target the heterodimeric E1 SUMO activation complex (SAE1/UBA2). Here, we report that activated MYC signaling is an actionable molecular vulnerability in vitro and in a preclinical murine in vivo model of MYC-driven BCL. While SUMOi conferred direct effects on MYC-driven lymphoma cells, SUMO inhibition also resulted in substantial remodeling of various subsets of the innate and specific immunity in vivo. Specifically, SUMOi increased the number of memory B cells as well as cytotoxic and memory T cells, subsets that are attributed a key role within a coordinated anti-tumor immune response. In summary, our data constitute pharmacologic SUMOi as a powerful therapy in a subset of BCL causing massive remodeling of the normal B-cell and T-cell compartment.

GTPBP4 promotes hepatocellular carcinoma progression and metastasis via the PKM2 dependent glucose metabolism

Guanosine triphosphate binding protein 4 (GTPBP4) is a key regulator of cell cycle progression and MAPK activation. However, how its biological properties intersect with cellular metabolism in hepatocellular carcinoma (HCC) development remains poorly unexplained. Here, high GTPBP4 expression is found to be significantly associated with worse clinical outcomes in patients with HCC. Moreover, GTPBP4 upregulation is paralleled by DNA promoter hypomethylation and regulated by DNMT3A, a DNA methyltransferase. Additionally, both gain- and loss-of-function studies demonstrate that GTPBP4 promotes HCC growth and metastasis in vitro and in vivo. Mechanically, GTPBP4 can induce dimeric pyruvate kinase M2 (PKM2) formation through protein sumoylation modification to promote aerobic glycolysis in HCC. Notably, active GTPBP4 facilitates SUMO1 protein activation by UBA2, and acts as a linker bridging activated SUMO1 protein and PKM2 protein to induce PKM2 sumoylation. Furthermore, SUMO-modified PKM2 relocates from the cytoplasm to the nucleus may also could contribute to HCC progression through activating epithelial-mesenchymal transition (EMT) and STAT3 signaling pathway. Shikonin, a PKM2-specific inhibitor, can attenuate PKM2 dependent HCC glycolytic reprogramming, growth and metastasis promoted by GTPBP4, which offers a promising therapeutic candidate for HCC patients. Our findings indicate that GTPBP4-PKM2 regulatory axis plays a vital role in promoting HCC proliferation as well as metastasis by aerobic glycolysis and offer a promising therapeutic target for HCC patients.

Small ubiquitin-related modifier (SUMO)ylation of SIRT1 mediates (-)-epicatechin inhibited- differentiation of cardiac fibroblasts into myofibroblasts

Context (-)-Epicatechin (EPI) is a crucial substance involved in the protective effects of flavanol-rich foods. Previous studies have indicated EPI has a cardioprotective effect, but the molecular mechanisms in inhibition of cardiac fibrosis are unclear. Objective We evaluated the effect of EPI in preventing cardiac fibrosis and the underlying molecular mechanism related to the SIRT1-SUMO1/AKT/GSK3β pathway. Materials and methods Cardiac fibrosis mice model was established with transaortic constriction (TAC). Male C57BL/6 mice were randomly separated into 4 groups. Mice received 1 mg/kg/day of EPI or vehicle orally for 4 weeks. The acutely isolated cardiac fibroblasts were induced to myofibroblasts with 1 µM angiotensin II (Ang II). The cardiac function was measured with the ultrasonic instrument. Histological analysis of mice’s hearts was determined with H&E or Masson method. The protein level of fibrosis markers, SUMOylation of SIRT1, and AKT/GSK3β pathway were quantified by immunofluorescence and western blot. Results EPI treatment (1 mg/kg/day) could reverse the TAC-induced decline in LVEF (TAC, 61.28% ± 1.33% vs. TAC + EPI, 74.00% ± 1.64%), LVFS (TAC, 28.16% ± 0.89% vs. TAC + EPI, 37.18% ± 1.29%). Meantime, we found that 10 µM EPI blocks Ang II-induced transformation of cardiac fibroblasts into myofibroblasts. The underlying mechanism of EPI-inhibited myofibroblasts transformation involves activation of SUMOylation of SIRT1 through SP1. Furthermore, SUMOylation of SIRT1 inhibited Ang II-induced fibrogenic effect via the AKT/GSK3β pathway. Conclusion EPI plays a protective effect on cardiac fibrosis by regulating the SUMO1-dependent modulation of SIRT1, which provides a theoretical basis for use in clinical therapies.

SIZ1-mediated SUMOylation responds to multiple abiotic stresses in plants

SUMOylation is an important posttranslational modification of eukaryotes that is widely present in metabolic regulation processes under various stresses. SIZ1 is a SUMO E3 ligase that plays a key role in substrate-specific recognition and directly promotes the binding of SUMO proteins to target proteins. Here, we reviewed the characteristics of SUMO as well as SUMOylation and the role of the SUMO E3 ligase SIZ1 in plant adaptation to abiotic stresses such as abnormal temperature, drought, salinization, phosphorus and nitrogen deficiency, heavy metal and metalloid toxicity, light exposure and reactive oxygen species. This review will enhance our understanding of these stressors and provide a useful reference for appropriate crop improvement, especially in suboptimal environments. Moreover, we provide a reference for future research on the role of SUMOylation in plant responses to abiotic stress.

SUMOylation in Skeletal Development, Homeostasis, and Disease.

The modification of proteins by small ubiquitin-related modifier (SUMO) molecules, SUMOylation, is a key post-translational modification involved in a variety of biological processes, such as chromosome organization, DNA replication and repair, transcription, nuclear transport, and cell signaling transduction. In recent years, emerging evidence has shown that SUMOylation regulates the development and homeostasis of the skeletal system, with its dysregulation causing skeletal diseases, suggesting that SUMOylation pathways may serve as a promising therapeutic target. In this review, we summarize the current understanding of the molecular mechanisms by which SUMOylation pathways regulate skeletal cells in physiological and disease contexts.

DNA Damage Induction Alters the Expression of Ubiquitin and SUMO Regulators in Preimplantation Stage Pig Embryos.

DNA damage in early-stage embryos impacts development and is a risk factor for segregation of altered genomes. DNA damage response (DDR) encompasses a sophisticated network of proteins involved in sensing, signaling, and repairing damage. DDR is regulated by reversible post-translational modifications including acetylation, methylation, phosphorylation, ubiquitylation, and SUMOylation. While important regulators of these processes have been characterized in somatic cells, their roles in early-stage embryos remain broadly unknown. The objective of this study was to explore how ubiquitylation and SUMOylation are involved in the regulation of early development in porcine embryos by assessing the mRNA profile of genes encoding ubiquitination (UBs), deubiquitination (DUBs), SUMOylation (SUMOs) or deSUMOylation (deSUMOs) enzymes in oocyte and embryos at different stages of development, and to evaluate if the induction of DNA damage at different stages of embryo development would alter the mRNA abundance of these genes. Pig embryos were produced by in vitro fertilization and DNA damage was induced by ultraviolet (UV) light exposure for 10 s on days 2, 4 or 7 of development. The relative mRNA abundance of most UBs, DUBs, SUMOs, and deSUMOs was higher in oocytes and early-stage embryos than in blastocysts. Transcript levels for UBs (RNF20, RNF40, RNF114, RNF169, CUL5, DCAF2, DECAF13, and DDB1), DUBs (USP16), and SUMOs (CBX4, UBA2 and UBC9), were upregulated in early-stage embryos (D2 and/or D4) compared to oocytes and blastocysts. In response to UV-induced DNA damage, transcript levels of several UBs, DUBs, SUMOs, and deSUMOs decreased in D2 and D4 embryos, but increased in blastocysts. These findings revealed that transcript levels of genes encoding for important UBs, DUBs, SUMOs, and deSUMOs are regulated during early embryo development and are modulated in response to induced DNA damage. This study has also identified candidate genes controlling post-translational modifications that may have relevant roles in the regulation of normal embryo development, repair of damaged DNA, and preservation of genome stability in the pig embryo.

Small ubiquitin-related modifier-fused bacteriophage tail fiber protein with favorable aqueous solubility for lateral flow assay of Pseudomonas aeruginosa

Rapid and sensitive assay of pathogenic bacteria is critical for minimizing the risk of infectious diseases. Inspired by the interaction between bacteriophages and host bacteria, we obtained a gene sequence of tail fiber protein (TFP) from Pseudomonas aeruginosa (P. aeruginosa) bacteriophage. Then the gene sequence was used to express a recombinant TFP, which can act as a potential capture molecule for P. aeruginosa. Small ubiquitin-related modifier (SUMO) tag was fused onto the TFP fragment to overcome its unfavorable aqueous solubility. The obtained SUMO tag-fused TFP (STFP) can be uniformly distributed onto a nitrocellulose membrane to form a test line due to the improved aqueous solubility, which facilities fabrication of a lateral flow assay strip. Thus we developed a lateral flow assay method by using STFP as a capture molecule and AuCo nanoparticles-labeled aptamer as a signal tracer for point-of-care testing of P. aeruginosa. By using the test strip, P. aeruginosa can be semi quantified with color band and quantified with chemiluminescent (CL) signal catalyzed by AuCo nanoparticles. The concentration range for quantification is 3.3 × 102 CFU/mL to 3.3 × 107 CFU/mL. The test strip was applied to assay P. aeruginosa in different sample matrixes including cerebrospinal fluid, physiological salt solution, drinking water and pear juice. The results demonstrate the application potential of the STFP-based lateral flow assay for medical diagnosis, food and drug safety monitoring.