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College students' diverse outcomes are said to be influenced by their motivation for their studies. Academic motivation is the desire or interest students have in participating in their education and college experience. Research has repeatedly shown that academically driven students have a tendency to value education and college, to like learning and learningrelated activities. Academic motivation is an essential learning indicator. The purpose of this study was to determine academic motivation in terms of gender and family structure among junior college students. The participants of the study consisted of 112 junior college students. The Academic Motivation Scale developed by Vallerand et al., 1992 was used to measure students' academic motivation level. The results revealed that no significant differences were found in academic motivation according to gender and family structure

Background Functional genomics uses unbiased systematic genome-wide gene disruption or analyzes natural variations such as gene expression profiles of different tissues from multicellular organisms to link gene functions to particular phenotypes. Functional genomics approaches are of particular importance to identify large sets of genes that are specifically important for a particular biological process beyond known candidate genes, or when the process has not been studied with genetic methods before. Results Here, we present a large set of genes whose disruption interferes with the function of the odoriferous defensive stink glands of the red flour beetle Tribolium castaneum. This gene set is the result of a large-scale systematic phenotypic screen using RNA interference applied in a genome-wide forward genetics manner. In this first-pass screen, 130 genes were identified, of which 69 genes could be confirmed to cause phenotypic changes in the glands upon knock-down, which vary from necrotic tissue and irregular reservoir size to irregular color or separation of the secreted gland compounds. Gene ontology analysis revealed that many of those genes are encoding enzymes (peptidases and cytochromes P450) as well as proteins involved in membrane trafficking with an enrichment in lysosome and mineral absorption pathways. The knock-down of 13 genes caused specifically a strong reduction of para-benzoquinones in the gland reservoirs, suggesting a specific function in the synthesis of these toxic compounds. Only 14 of the 69 confirmed gland genes are differentially overexpressed in stink gland tissue and thus could have been detected in a transcriptome-based analysis. However, only one out of eight genes identified by a transcriptomics approach known to cause phenotypic changes of the glands upon knock-down was recognized by this phenotypic screen, indicating the limitation of such a non-redundant first-pass screen. Conclusion Our results indicate the importance of combining diverse and independent methodologies to identify genes necessary for the function of a certain biological tissue, as the different approaches do not deliver redundant results but rather complement each other. The presented phenotypic screen together with a transcriptomics approach are now providing a set of close to hundred genes important for odoriferous defensive stink gland physiology in beetles.

About 30% of all small molecular drugs are organic cations (OCs). If these are more or less hydrophilic, they require membrane transporters to pass through biological membranes. Here, the proton-organic cation (Hsup+/supOC) antiporter may play a physiologically most relevant role, particularly concerning passage through the blood-brain barrier. Membrane transport of about 70 OCs is significantly enhanced by this Hsup+/supOC antiporter. Surprisingly still today the gene coding for this antiporter was not yet identified. However, the Hsup+/supOC antiporter is characterized by concentration- and pH-dependent uptake, antiport with another OC, and susceptibility to inhibition by specific inhibitors. Moreover, in the studied tissues and cell types, transport is not mediated by already well-known organic cation transporters. The review explains the typically used assays to identify potential substrates of the Hsup+/supOC antiporter. Thus far, the gene encoding for this transporter has not yet been identified, but a better understanding of this protein may be most relevant because it may affect the pharmacokinetics of up to 10% of all low molecular substances. This review summarizes the known functional characteristics of the Hsup+/supOC antiporter, its cell and tissue expression, and its substrate spectrum. Summarizing the features of the substrates of the Hsup+/supOC antiporter may even suggest that for OCs, good penetration through the blood-brain barrier is almost synonymous with being a substrate of the Hsup+/supOC antiporter. In clinical studies, pharmacokinetics of typical substrates of the antiporter showed outstanding between-subject variability.

Background Although psychostimulants apparently do cross the BBB, it is poorly understood how these hydrophilic and positively charged molecules can pass the blood-brain barrier (BBB). That may be mediated by a genetically still uncharacterized H+/OC antiporter with high activity at the BBB. Methods We studied the uptake of 16 psychostimulants and hallucinogens with hCMEC/D3 cells using the prototypic inhibitor imipramine (cis-inhibition), exchange transport with diphenhydramine and clonidine (trans-stimulation), proton dependency of the uptake, and we characterized the concentration-dependent uptake. Results Cell uptake of methylenedioxyamphetamines, amphetamines and dimethyltryptamine (DMT) were strongly inhibited (to about 10% of the controls) by imipramine and diphenhydramine, whereas uptake of cathine was only weakly inhibited and mescaline not significantly. Amphetamine, methylamphetamine, para-Methoxy-N-methylamphetamine (PMMA), Methylenedioxymethamphetamine (MDMA), phentermine and DMT exhibited the highest exchange after preloading with diphenhydramine with only 5.5%, 5.2%, 7.8%, 6%, 1.9%, 7.6% remaining in the cells. Less and no exchange were seen with cathine and mescaline, respectively. Dependence on intracellular pH was most pronounced with the methylendioxyamphetamines while uptake of cathine, DOI and cocaine were only moderately affected and mescaline not at all. Conclusion Except for mescaline, all psychostimulants studied here were substrates of the H+/OC antiporter, implicating a strong need for a better characterization of this transport protein.

Background Most of the known genes required for developmental processes have been identified by genetic screens in a few well-studied model organisms, which have been considered representative of related species, and informative—to some degree—for human biology. The fruit fly Drosophila melanogaster is a prime model for insect genetics, and while conservation of many gene functions has been observed among bilaterian animals, a plethora of data show evolutionary divergence of gene function among more closely-related groups, such as within the insects. A quantification of conservation versus divergence of gene functions has been missing, without which it is unclear how representative data from model systems actually are. Results Here, we systematically compare the gene sets required for a number of homologous but divergent developmental processes between fly and beetle in order to quantify the difference of the gene sets. To that end, we expanded our RNAi screen in the red flour beetle Tribolium castaneum to cover more than half of the protein-coding genes. Then we compared the gene sets required for four different developmental processes between beetle and fly. We found that around 50% of the gene functions were identified in the screens of both species while for the rest, phenotypes were revealed only in fly (~ 10%) or beetle (~ 40%) reflecting both technical and biological differences. Accordingly, we were able to annotate novel developmental GO terms for 96 genes studied in this work. With this work, we publish the final dataset for the pupal injection screen of the iBeetle screen reaching a coverage of 87% (13,020 genes). Conclusions We conclude that the gene sets required for a homologous process diverge more than widely believed. Hence, the insights gained in flies may be less representative for insects or protostomes than previously thought, and work in complementary model systems is required to gain a comprehensive picture. The RNAi screening resources developed in this project, the expanding transgenic toolkit, and our large-scale functional data make T. castaneum an excellent model system in that endeavor.

Most gene functions were detected by screens in very few model organisms but it has remained unclear how comprehensive these data are. Here, we expanded our RNAi screen in the red flour beetleTribolium castaneumto cover more than half of the protein-coding genes and we compared the gene sets involved in several processes between beetle and fly.We find that around 50 % of the gene functions are detected in both species while the rest was found only in fly (~10%) or beetle (~40%) reflecting both technical and biological differences. We conclude that work in complementary model systems is required to gain a comprehensive picture on gene functions documented by the annotation of novel GO terms for 96 genes studied here. The RNAi screening resources developed in this project, the expanding transgenic tool-kit and our large-scale functional data makeT. castaneuman excellent model system in that endeavor.

The solidification of metals continues to be a phenomenon of great interest to physicists, metallurgists, casting engineers, and software developers. It is a non-linear transient phenomenon, posing a challenge in terms of modeling and analysis. During the solidification of a casting in a mould, the heat-transfer between the casting and the mould plays a vital role. This paper attempts to study heat flow within the casting, as well as from the casting to the mould, and finally obtains the temperature history of some points inside the casting. The most important instant of time is when the hottest region inside the casting is solidifying. ProCAST software has been used to obtain the temperature distribution in the casting process by performing Transient Thermal Analysis. In this research work, solidification of lead-2wt%antimony alloy has been carried out in the different sizes of metallic mold to predict the formation of shrinkage during solidification. Theoretical results have been validated experimentally for a particular case of lead-2wt%antimony alloy solidification. Results obtained by simulation software are compared with the experimental reading of temperature and found to be in good agreement. Voids appeared at the top and isolated area of castings for the defect-free direct method used in this study.

Overexpression of single genes in mammalian cells is widely used to investigate protein function in basic and applied biosciences and in drug research. A better understanding of interactions of two proteins is an important next step in the advancement of our understanding of complex biological systems. However, simultaneous and robust overexpression of two or more genes is challenging. The Flp-In system integrates a vector into cell lines at a specific genomic locus, but has not been used for integration of more than one gene. Here we present a modification of the Flp-In system that enables the simultaneous targeted integration of two genes. We describe the modification and generation of the vectors required and give the complete protocol for transfection and validation of correct genomic integration and expression. We also provide results on the stability and reproducibility, and we functionally validated this approach with a pharmacologically relevant combination of a membrane transporter facilitating drug uptake and an enzyme mediating drug metabolism. Open-Access-Publikationsfonds 2020 peerReviewed

Significance One of the first crucial steps of animal development is to distinguish the anterior versus the posterior pole of the embryo, i.e., the AP axis. If this process fails, embryos may develop two mirror image tails or heads. In the fly Drosophila, the mother provides the signals required for AP axis formation, while in vertebrates, gene activity of the embryo is required as well. We identified two genes whose knockdown leads to double-tail phenotypes in the beetle Tribolium, representing the insect-typical short-germ embryogenesis. Intriguingly, embryo polarity depends on zygotic gene activities and Wnt signaling. Hence, short-germ insect axis formation is more similar to vertebrates than the mechanism employed by Drosophila., The distinction of anterior versus posterior is a crucial first step in animal embryogenesis. In the fly Drosophila, this axis is established by morphogenetic gradients contributed by the mother that regulate zygotic target genes. This principle has been considered to hold true for insects in general but is fundamentally different from vertebrates, where zygotic genes and Wnt signaling are required. We investigated symmetry breaking in the beetle Tribolium castaneum, which among insects represents the more ancestral short-germ embryogenesis. We found that maternal Tc-germ cell-less is required for anterior localization of maternal Tc-axin, which represses Wnt signaling and promotes expression of anterior zygotic genes. Both RNAi targeting Tc-germ cell-less or double RNAi knocking down the zygotic genes Tc-homeobrain and Tc-zen1 led to the formation of a second growth zone at the anterior, which resulted in double-abdomen phenotypes. Conversely, interfering with two posterior factors, Tc-caudal and Wnt, caused double-anterior phenotypes. These findings reveal that maternal and zygotic mechanisms, including Wnt signaling, are required for establishing embryo polarity and induce the segmentation clock in a short-germ insect.

Although muscle development has been widely studied inDrosophila melanogasterthere are still many gaps in our knowledge, and it is not known to which extent this knowledge can be transferred to other insects. To help in closing these gaps we participated in a large-scale RNAi screen that used the red flour beetle,Tribolium castaneum, as a screening platform. The effects of systemic RNAi were screened upon double-stranded RNA injections into appropriate muscle-EGFP tester strains. Injections into pupae were followed by the analysis of the late embryonic/early larval muscle patterns, and injections into larvae by the analysis of the adult thoracic muscle patterns. Herein we describe the results of the first-pass screens with pupal and larval injections, which covered ~8,500 and ~5,000 genes, respectively, of a total of ~16,500 genes of theTriboliumgenome. Apart from many genes known fromDrosophilaas regulators of muscle development, a collection of genes previously unconnected to muscle development yielded phenotypes in larval body wall and leg muscles as well as in indirect flight muscles. We then present the main candidates from the pupal injection screen that remained after being processed through a series of verification and selection steps. Further, we discuss why distinct though overlapping sets of genes are revealed by theDrosophilaandTriboliumscreening approaches.

Background Small RNAs (sRNAs) are important modulators of gene expression in bacteria. Regulation by sRNAs is yet to be established in Bacillus anthracis. Here, regulation and RNA-binding properties of Hfq-like RNA chaperones in B. anthracis are investigated. Methods Transcript levels were measured by RT-PCR. Proteins were cloned, purified, and their ability to bind sRNA was seen by EMSA. Regulators of Hfq1 were identified by in silico analysis and validated by EMSA. Conserved sRNAs were identified by homology search and their ability to bind Hfq1 was seen by EMSA. Residues crucial for sRNA binding were identified by mutational studies. Results hfq1 and hfq3 showed expression during exponential phase on BHI medium, while hfq2 showed no expression. Hfq1 and Hfq3 formed hexamer and sRNA–Hfq complex, not Hfq2. In silico prediction and EMSA confirmed AbrB binding to the promoter of Hfq1. Homology search identified 7 sRNAs in B. anthracis. The sRNA CsfG showed binding to Hfq1 via residues Y24, F29, Q30, R32, K56, and H57. Conclusions Hfq1 and Hfq3 can function as RNA chaperones in B. anthracis. The transition phase repressor AbrB might be responsible for the growth-dependent expression of Hfq1. The sporulation-specific sRNA CsfG binds to Hfq1 via its distal surface and requires an intact hexameric structure for forming CsfG-Hfq1 complex. General significance This is the first report demonstrating the regulation and functional properties of Hfq-like RNA chaperones in B. anthracis and paves the path towards establishing sRNA-based regulation in B. anthracis.