Helper Factors Research Articles

Research on the Influencing Factors of College Students' Deep and Meaningful Learning in Blended Learning Mode

This study explores the influencing factors of deep and meaningful learning in blended learning modes and their interrelationships. The sample comprises 397 college students from a university in Sichuan Province, selected through random sampling. Data collection utilized a questionnaire based on Bandura's ternary interaction theory, covering dimensions such as learners, helpers, environment, and interaction. Hypotheses were formulated based on literature research, and a survey was developed using established scales. Quantitative analysis was conducted using SPSS and AMOS software. The mean, standard deviation, variance, skewness, and kurtosis values were within reasonable ranges. The latent variables of the model exhibited sound convergent validity, with SFL ranging from 0.807 to 0.965, AVE from 0.697 to 0.946, and CR from 0.919 to 0.946. Model fit indices indicated acceptable fit (CMIN/DF: 2.303, NFI: 0.966, CFI: 0.980, RMSEA: 0.58, RMR: 0.008, PNFI: 0.789). Through path analysis, the study optimized the model, resulting in the final structural equation model (SEM). The findings suggest that (1) learner, environmental, and interaction factors positively influence deep and meaningful learning, while helper factors show a negative correlation; (2) learner, interaction, and helper factors mediate the environment's impact on deep and meaningful learning; and (3) environmental factors have the most significant influence on helper factors, followed by interaction and learner factors. Helpers play a significant role in enhancing deep understanding for learners. These insights guide effective strategies in promoting deep and meaningful learning in blended learning environments.

Educational and Instructional Technology in EFL/ESL Classrooms: A Literature Review

Technology in education has a significant impact on how new and creative types of assistance are offered to instructors, students, and the learning process as a whole. This review paper presents a literature review on educational and instructional technology in EFL/ESL classrooms. Utilizing the qualitative method, which relies solely on the collection of materials and findings from previous studies, such as books, papers, and articles, was employed. In addition, this review paper seeks to make the perspectives of earlier academics more visible and distinct. The study sheds light on whether technology enhances and influences the vocabulary, grammar, speaking, and reading skills of students. Based on the existing literature, technology in education is promising and a vital tool, especially in language learning. The findings have shown that there is a strong positive correlation between technology and language learning. Also, technology is the best helper, facilitator, and motivational factor in the classroom environment. Teachers are encouraged to use technology in teaching language skills, especially CALL, and wait to see positive results from their students. Finally, it is also advised that teachers educate themselves to become proficient in using different technology tools with their students. Ultimately, doing so, will lead them to enjoy teaching challenging subjects to their students using technology.

Herpes Simplex Virus 1 Coinfection Modifies Adeno-associated Virus Genome End Recombination.

ABSTRACTWild-type adeno-associated virus (AAV) can only replicate in the presence of helper factors, which can be provided by coinfecting helper viruses such as adenoviruses and herpesviruses. The AAV genome consists of a linear, single-stranded DNA (ssDNA), which is converted into different molecular structures within the host cell. Using high-throughput sequencing, we found that herpes simplex virus 1 (HSV-1) coinfection leads to a shift in the type of AAV genome end recombination. In particular, open-end inverted terminal repeat (ITR) recombination was enhanced, whereas open-closed ITR recombination was reduced in the presence of HSV-1. We demonstrate that the HSV-1 protein ICP8 plays an essential role in HSV-1-mediated interference with AAV genome end recombination, indicating that the previously described ICP8-driven mechanism of HSV-1 genome recombination may be underlying the observed changes. We also provide evidence that additional factors, such as products of true late genes, are involved. Although HSV-1 coinfection significantly changed the type of AAV genome end recombination, no significant change in the amount of circular AAV genomes was identified.IMPORTANCE Adeno-associated virus (AAV)-mediated gene therapy represents one of the most promising approaches for the treatment of genetic diseases. Currently, various GMP-compatible production methods can be applied to manufacture clinical-grade vector, including methods that employ helper factors derived from herpes simplex virus 1 (HSV-1). Yet, to date, we do not fully understand how HSV-1 interacts with AAV. We observed that HSV-1 modulates AAV genome ends similarly to the genome recombination events observed during HSV-1 replication and postulate that further improvements of the HSV-1 production platform may enhance packaging of the recombinant AAV particles.

ICTV Virus Taxonomy Profile: Nanoviridae.

Nanoviridae is a family of plant viruses (nanovirids) whose members have small isometric virions and multipartite, circular, single-stranded (css) DNA genomes. Each of the six (genus Babuvirus) or eight (genus Nanovirus) genomic DNAs is 0.9–1.1 kb and is separately encapsidated. Many isolates are associated with satellite-like cssDNAs (alphasatellites) of 1.0–1.1 kb. Hosts are eudicots, predominantly legumes (genus Nanovirus), and monocotyledons, predominantly in the order Zingiberales (genus Babuvirus). Nanovirids require a virus-encoded helper factor for transmission by aphids in a circulative, non-propagative manner. This is a summary of the ICTV Report on the family Nanoviridae, which is available at ictv.global/report/nanoviridae.

Systematic use of synthetic 5'-UTR RNA structures to tune protein translation improves yield and quality of complex proteins in mammalian cell factories.

Predictably regulating protein expression levels to improve recombinant protein production has become an important tool, but is still rarely applied to engineer mammalian cells. We therefore sought to set-up an easy-to-implement toolbox to facilitate fast and reliable regulation of protein expression in mammalian cells by introducing defined RNA hairpins, termed ‘regulation elements (RgE)’, in the 5′-untranslated region (UTR) to impact translation efficiency. RgEs varying in thermodynamic stability, GC-content and position were added to the 5′-UTR of a fluorescent reporter gene. Predictable translation dosage over two orders of magnitude in mammalian cell lines of hamster and human origin was confirmed by flow cytometry. Tuning heavy chain expression of an IgG with the RgEs to various levels eventually resulted in up to 3.5-fold increased titers and fewer IgG aggregates and fragments in CHO cells. Co-expression of a therapeutic Arylsulfatase-A with RgE-tuned levels of the required helper factor SUMF1 demonstrated that the maximum specific sulfatase activity was already attained at lower SUMF1 expression levels, while specific production rates steadily decreased with increasing helper expression. In summary, we show that defined 5′-UTR RNA-structures represent a valid tool to systematically tune protein expression levels in mammalian cells and eventually help to optimize recombinant protein expression.

The Interplay between Adeno-Associated Virus and its Helper Viruses.

The adeno-associated virus (AAV) is a small, nonpathogenic parvovirus, which depends on helper factors to replicate. Those helper factors can be provided by coinfecting helper viruses such as adenoviruses, herpesviruses, or papillomaviruses. We review the basic biology of AAV and its most-studied helper viruses, adenovirus type 5 (AdV5) and herpes simplex virus type 1 (HSV-1). We further outline the direct and indirect interactions of AAV with those and additional helper viruses.

Aphid transmission of nanoviruses.

The genus Nanovirus consists of plant viruses that predominantly infect legumes leading to devastating crop losses. Nanoviruses are transmitted by various aphid species. The transmission occurs in a circulative nonpropagative manner. It was long suspected that a virus-encoded helper factor would be needed for successful transmission by aphids. Recently, a helper factor was identified as the nanovirus-encoded nuclear shuttle protein (NSP). The mode of action of NSP is currently unknown in contrast to helper factors from other plant viruses that, for example, facilitate binding of virus particles to receptors within the aphids' stylets. In this review, we are summarizing the current knowledge about nanovirus-aphid vector interactions.

Ran-binding protein 3 is associated with human spermatogenesis and male infertility.

Ran-binding protein 3 (RanBP3) is a Ran-interacting protein, which participates in the Ran GTPase system in cancer cell biology. However, the expression pattern and physiological role of RanBP3 remain largely unknown. In this study, we found that RanBP3 was expressed in human testes and localised to spermatogonium and spermatocyte of germ cells. In subcellular structure, its localisation is in the nucleus and cytoplasm. Interestingly, compared with normal groups, RanBP3 expression was lower in groups of patients with Maturation Arrest (MA) and Sertoli cell-only syndrome (SCO) when considered by the Johnson Score. RanBP3 expression in the MA group and SCO groups was dramatically lower than that in the normal control group. Studies have shown that RanBP3, which is one of the helper factors of Ran, is mainly participate in the nucleocytoplasmic transport of cells. RanBP3 helps Ran to achieve some functions such as nucleocytoplasmic transport, spindle assembly during mitosis and nuclear assembly after mitosis. Consequent changes in the expression of RanBP3 may associate with human spermatogenesis disorders and male infertility. The identification and characterisation of RanBP3 enhances our understanding of the molecular mechanisms underpinning its function in human spermatogenesis and male infertility.

High-level expression of Thermomyces dupontii thermophilic lipase in Pichia pastoris via combined strategies.

In this study, a combined strategy was used to improve the production of Thermomyces dupontii lipase (TDL) in Pichia pastoris. First, the native gene of TDL was optimized based on the codon usage of P. pastoris, ligated to pPICZαA and transformed in P. pastoris X33. A recombinant strain designated X33-T23 with the highest activity (1020 U/mL in shake flasks) amongst 216 recombinant colonies was selected for further investigations. To further increase the production of TDL, nine different secretion helper factor genes were transformed in the recombinant strain, X33-T23. The recombinant strain co-expression with the gene encoding protein disulfide isomerase, designated X33-T23-PDI, exhibited the highest activity in shake flasks (1760 U/mL) and in 5L bioreactor (57521 U/mL) which were 1.67- and 1.46-fold higher, respectively, than for strain X33-T23. Additionally, the optimization of the inducers (temperature and pH) for the recombinant strain X33-T23-PDI in 5L bioreactor produced, as expected, much higher lipase activity (81203 U/mL). The results of this study will provide an effective method to produce TDL and give some clues on how to improve production of heterologous proteins in P. pastoris.

PENGARUH PIJAT PERINEUM SELAMA MASA KEHAMILAN TERHADAP KEJADIAN RUPTURA PERINEUM SPONTAN DI PMB SHINTA NUR ROCHMAYANTI, SSiT., M.Kes

Some of the factors that cause perineal rupture consist of maternal factors, fetal factors, and helper factors. A method of reduce perineal rupture is a perinel massage to promote heatlh, blood flow and elasticity of the perineum. Based on preliminary study at PMB Shinta Nur Rochmayanti, SSiT., M.Kes from June to September of 2017 there were 20 births per month, and 70% had spontaneous birthrate while 30% had no perineal rupture. The purpose of this study was to determine the effect of perineal massage during pregnancy on the incidence of Rupture Perineum during delivery at the Midwives Practice Midwife Shinta Nur Rochmayanti, SSiT., M.Kes year 2018. The objektive of this study was to determine the effect of perineal massage on the incidence of perineal rupture in pregnant period during childbirth in independent practice of midwifery in PMB Shinta Nur Rochmayanti, SSiT., M.Kes year 2018.This study is Quasi Experimental study with Post Test Only Control Group Design method. The study was conducted in midwife Mandiri Practice (PMB) Shinta Nur Rochmayanti, SSiT., M.Kes Surabaya in 2018 with subject of 28 pregnant women 36 weeks pregnancy consisting of 14 intervention group and 14 control group. The study was conducted from December 2017 until April 2018, then the data were analyzed by Chi Square test. The incidence of Rupture Perineum in the intervention group after perineal massage was only 14,3% while in the control group 64,3%. The results showed that there was an effect of perineal massage during pregnancy on the incidence of perineum rupture (p <0.05). This study concluded that perineal massage during pregnancy affects the incidence of perineal rupture at delivery.Keywords: perineal massage, pregnant periode, perineal rupture

Identification of novel factors enhancing recombinant protein production in multi-copy Komagataella phaffii based on transcriptomic analysis of overexpression effects

The methylotrophic yeast Komagataella phaffii (Pichia pastoris) has been developed into a highly successful system for heterologous protein expression in both academia and industry. However, overexpression of recombinant protein often leads to severe burden on the physiology of K. phaffii and triggers cellular stress. To elucidate the global effect of protein overexpression, we set out to analyze the differential transcriptome of recombinant strains with 12 copies and a single copy of phospholipase A2 gene (PLA2) from Streptomyces violaceoruber. Through GO, KEGG and heat map analysis of significantly differentially expressed genes, the results indicated that the 12-copy strain suffered heavy cellular stress. The genes involved in protein processing and stress response were significantly upregulated due to the burden of protein folding and secretion, while the genes in ribosome and DNA replication were significantly downregulated possibly contributing to the reduced cell growth rate under protein overexpression stress. Three most upregulated heat shock response genes (CPR6, FES1, and STI1) were co-overexpressed in K. phaffii and proved their positive effect on the secretion of reporter enzymes (PLA2 and prolyl endopeptidase) by increasing the production up to 1.41-fold, providing novel helper factors for rational engineering of K. phaffii.

Recycling of a selectable marker with a self-excisable plasmid in Pichia pastoris

Pichia pastoris is a widely used heterologous protein production workhorse. However, with its multiple genetic modifications to solve bottlenecks for heterologous protein productivity, P. pastoris lacks selectable markers. Existing selectable marker recycling plasmids have drawbacks (e.g., slow growth and conditional lethality). Here, zeocin-resistance marker recycling vectors were constructed using the Cre/loxP recombination system. The vectors were used to (i) knock in heterologous phytase, xylanase and lipase expression cassettes, (ii) increase the phytase, xylanase and lipase gene copy number to 13, 5, and 5, respectively, with vector introduction and (iii) engineer the secretion pathway by co-overexpressing secretion helper factors (Sly1p and Sec1p) without introducing selectable markers, giving a phytase field of 0.833 g/L. The vectors allow selectable marker recycling and would be a useful tool to engineer P. pastoris for high heterologous protein productivity.

Cell Cycle-Dependent Expression of Adeno-Associated Virus 2 (AAV2) Rep in Coinfections with Herpes Simplex Virus 1 (HSV-1) Gives Rise to a Mosaic of Cells Replicating either AAV2 or HSV-1

Adeno-associated virus 2 (AAV2) depends on the simultaneous presence of a helper virus such as herpes simplex virus 1 (HSV-1) for productive replication. At the same time, AAV2 efficiently blocks the replication of HSV-1, which would eventually limit its own replication by diminishing the helper virus reservoir. This discrepancy begs the question of how AAV2 and HSV-1 can coexist in a cell population. Here we show that in coinfected cultures, AAV2 DNA replication takes place almost exclusively in S/G2-phase cells, while HSV-1 DNA replication is restricted to G1 phase. Live microscopy revealed that not only wild-type AAV2 (wtAAV2) replication but also reporter gene expression from both single-stranded and double-stranded (self-complementary) recombinant AAV2 vectors preferentially occurs in S/G2-phase cells, suggesting that the preference for S/G2 phase is independent of the nature of the viral genome. Interestingly, however, a substantial proportion of S/G2-phase cells transduced by the double-stranded but not the single-stranded recombinant AAV2 vectors progressed through mitosis in the absence of the helper virus. We conclude that cell cycle-dependent AAV2 rep expression facilitates cell cycle-dependent AAV2 DNA replication and inhibits HSV-1 DNA replication. This may limit competition for cellular and viral helper factors and, hence, creates a biological niche for either virus to replicate.IMPORTANCE Adeno-associated virus 2 (AAV2) differs from most other viruses, as it requires not only a host cell for replication but also a helper virus such as an adenovirus or a herpesvirus. This situation inevitably leads to competition for cellular resources. AAV2 has been shown to efficiently inhibit the replication of helper viruses. Here we present a new facet of the interaction between AAV2 and one of its helper viruses, herpes simplex virus 1 (HSV-1). We observed that AAV2 rep gene expression is cell cycle dependent and gives rise to distinct time-controlled windows for HSV-1 replication. High Rep protein levels in S/G2 phase support AAV2 replication and inhibit HSV-1 replication. Conversely, low Rep protein levels in G1 phase permit HSV-1 replication but are insufficient for AAV2 replication. This allows both viruses to productively replicate in distinct sets of dividing cells.

Overexpressing target helper genes enhances secretion and glycosylation of recombinant proteins in Pichia pastoris under simulated microgravity

In this study, the potential helper genes were identified through the data analysis of transcriptomic and proteomic profiling in recombinant Pichia pastoris cultured under simulated microgravity (SMG). Co-expressing of four genes PRX1, YAP1, AHA1, and YPT6, involved in the oxidative stress response and protein folding, exhibited promising helper factor effects on the recombinant protein yields in engineered P. pastoris, respectively. When two of the above genes were co-expressed simultaneously, β-glucuronidase (PGUS) specific activity was further increased by 30.3-50.6% comparing with that of single helper gene, particularly when the oxidative stress response and protein folding genes were both present in the combinations. In addition, co-expressing co-chaperone AHA1 and transcription factor YAP1 not only enhanced PGUS secretion, but also affected its glycosylation. Thus, through deep "omics" analysis of SMG effects, our results provided combined impact of new helper factors to improve the efficacy of recombinant protein secretion and glycosylation in engineered P. pastoris.

Effects of co-overexpression of secretion helper factors on the secretion of a HSA fusion protein (IL2-HSA) in pichia pastoris.

Pichia pastoris is generally considered as an expression host for heterologous proteins with the coding gene under control of the alcohol oxidase 1 (AOX1) promoter. The secretion of heterologous proteins in P. pastoris can be potentially affected by many factors. Based on our previous results, the secretion levels of human albumin (HSA) fusion protein IL2-HSA were only around 500 mg/L or less in fermentor cultures, which decreased more than 50% compared with that of HSA (>1 g/L). In this study, we selected five potential secretion helper factors, in which Ero1, Pdi1 and Kar2 were involved in protein folding and Sec1 and Sly1 were involved in vesicle trafficking. We evaluated the possible effects of individual overexpression of these secretion helper factors on the secretion of IL2-HSA in P. pastoris. Constitutive overexpression of the five selected secretion factors did not have an obvious negative effect on cell growth of the IL2-HSA secreting strain. Individual co-overexpression of Ero1, Kar2, Pdi1, Sec1 and Sly1 improved the secretion level of IL2-HSA to ~2.3-, 1.9-, 2.2-, 2.5- and 1.9-fold that in the control strain respectively in shake flasks. We evaluated the changes in mRNA and protein levels of the intracellular IL2-HSA, as well as the secretion helper factor genes in the co-overexpressing strains. Our results indicated that manipulating the expression level of ER resident protein Pdi1, Ero1, Kar2 and SM protein Sec1 and Sly1 could improve the secretion level of IL2-HSA fusion protein in P. pastoris, which provided new candidates for combinatorial engineering in future study. Copyright © 2016 John Wiley & Sons, Ltd.

CIB1 and CIB2 are HIV-1 helper factors involved in viral entry.

HIV-1 relies on the host-cell machinery to accomplish its replication cycle, and characterization of these helper factors contributes to a better understanding of HIV-host interactions and can identify potential novel antiviral targets. Here we explored the contribution of CIB2, previously identified by RNAi screening as a potential helper factor, and its homolog, CIB1. Knockdown of either CIB1 or CIB2 strongly impaired viral replication in Jurkat cells and in primary CD4+ T-lymphocytes, identifying these proteins as non-redundant helper factors. Knockdown of CIB1 and CIB2 impaired envelope-mediated viral entry for both for X4- and R5-tropic HIV-1, and both cell-free and cell-associated entry pathways were affected. In contrast, the level of CIB1 and CIB2 expression did not influence cell viability, cell proliferation, receptor-independent viral binding to the cell surface, or later steps in the viral replication cycle. CIB1 and CIB2 knockdown was found to reduce the expression of surface molecules implicated in HIV-1 infection, including CXCR4, CCR5 and integrin α4β7, suggesting at least one mechanism through which these proteins promote viral infection. Thus, this study identifies CIB1 and CIB2 as host helper factors for HIV-1 replication that are required for optimal receptor-mediated viral entry.

CIB1 and CIB2 are HIV-1 helper factors involved in envelope-mediated viral entry

CIB1 and CIB2 are HIV-1 helper factors involved in envelope-mediated viral entry

Novel helper factors influencing recombinant protein production in Pichia pastoris based on proteomic analysis under simulated microgravity.

Microgravity and simulated microgravity (SMG) have quite significant effects on numerous microbial cellular processes. The effects of SMG on the production of recombinant proteins and transcription profiling in prokaryotic and eukaryotic expression host have been investigated. The present study showed that SMG significantly enhanced the specific productivities and activities of the reporter enzymes PGUS and AtXYN that were expressed in recombinant Pichia pastoris. Proteomic profiling revealed that 21 proteins were significantly up-regulated and 35 proteins were drastically down-regulated at the stationary phase, when the recombinant P. pastoris responded to SMG. Six strongly up-regulated genes, TPX, FBA, PGAM, ENO, SBA1, and AKR-E, involved in the oxidative stress response, methanol metabolism, glycolytic pathway, and protein folding, were selected to analyze their impacts on recombinant protein production by co-overexpression in the shaker flask fermentation. The co-overexpressed strains, particularly TPX, FBA, and PGAM, demonstrated promising results with approximately 2.46-fold, 1.58-fold, and 1.33-fold increases in the specific yields of PGUS compared to the control after 48h of methanol induction, respectively. In the meantime, the corresponding PGUS specific activities were increased by 2.33-fold, 2.09-fold, and 1.32-fold, respectively. Thiol peroxidase (TPX), which is involved in the oxidative stress response, significantly influenced the transcriptional levels of the reporter gene PGUS. The present study provides valuable information for further exploration of the molecular mechanism of P. pastoris response to SMG and facilitates simulated microgravity for finding novel helper factors to rationally engineer the strains in normal fermentation by using proteomic studies.

Ly108 expression distinguishes subsets of invariant NKT cells that help autoantibody production and secrete IL-21 from those that secrete IL-17 in lupus prone NZB/W mice

Lupus is a systemic autoimmune disease characterized by anti-nuclear antibodies in humans and genetically susceptible NZB/W mice that can cause immune complex glomerulonephritis. T cells contribute to lupus pathogenesis by secreting pro-inflammatory cytokines such as IL-17, and by interacting with B cells and secreting helper factors such as IL-21 that promote production of IgG autoantibodies. In the current study, we determined whether purified NKT cells or far more numerous conventional non-NKT cells in the spleen of NZB/W female mice secrete IL-17 and/or IL-21 after TCR activation in vitro, and provide help for spontaneous IgG autoantibody production by purified splenic CD19+ B cells. Whereas invariant NKT cells secreted large amounts of IL-17 and IL-21, and helped B cells, non-NKT cells did not. The subset of IL-17 secreting NZB/W NKT cells expressed the Ly108loCD4−NK1.1− phenotype, whereas the IL-21 secreting subset expressed the Ly108hiCD4+NK1.1− phenotype and helped B cells secrete a variety of IgG anti-nuclear antibodies. α-galactocylceramide enhanced the helper activity of NZB/W and B6.Sle1b NKT cells for IgG autoantibody secretion by syngeneic B cells. In conclusion, different subsets of iNKT cells from mice with genetic susceptibility to lupus can contribute to pathogenesis by secreting pro-inflammatory cytokines and helping autoantibody production.

Truncation and Sequence Shuffling of Segment 6 Generate Replication-Competent Neuraminidase-Negative Influenza H5N1 Viruses

Influenza viruses are highly genetically variable and escape from immunogenic pressure by antigenic changes in their surface proteins, referred to as "antigenic drift" and "antigenic shift." To assess the potential genetic plasticity under strong selection pressure, highly pathogenic avian influenza virus (HPAIV) of subtype H5N1 was passaged 50 times in embryonated chicken eggs in the presence of a neutralizing, polyclonal chicken serum. The resulting mutant acquired major alterations in the neuraminidase (NA)-encoding segment. Extensive deletions and rearrangements were detected, in contrast to only 12 amino acid substitutions within all other segments. Interestingly, this new neuraminidase segment resulted from complex sequence shuffling and insertion of a short fragment originating from the PA segment. Characterization of that novel variant revealed a loss of the neuraminidase protein and enzymatic activity, but its replication efficiency remained comparable to that of the wild type. Using reverse genetics, a recombinant virus consisting of the wild-type backbone and the shortened NA segment could be generated; however, generation of this recombinant virus required the polybasic hemagglutinin cleavage site. Two independent repetitions starting with egg passage 30 in the presence of alternative chicken-derived immune sera selected mutants with similar but different large deletions within the NA segment without any neuraminidase activity, indicating a general mechanism. In chicken, these virus variants were avirulent, even though the HPAIV polybasic hemagglutinin cleavage site was still present. Overall, the variants reported here are the first HPAIV H5N1 strains without a functional neuraminidase shown to grow efficiently without any helper factor. These novel HPAIV variants may facilitate future studies shedding light on the role of neuraminidase in virus replication and pathogenicity.