Transcription factor Sp2: Molecular characterization and generation of Sp2 gene targeted mice

Summary The Sp family of transcription factors is characterised by its DNA-binding domain, an array of three conserved C2H2 zinc fingers. As a consequence of the conserved DNA binding motif, Sp members recognize GC (GGGGCGGGG) and GT (GGTGTGGGG) boxes with similar specificity and affinity. GC and GT boxes are im-portant for the expression of many different ubiquitous as well as tissue-specific cellular and viral genes. To date, nine members of the Sp family (Sp1 - Sp9) have been identified. In addition to their DNA-binding domain, Sp1 - Sp4 also share other structural features like two glutamine-rich transactivation domains and two serine/threonine-rich regions. Molecular and functional properties have been de-scribed for Sp1, Sp3 and Sp4. Mouse deletion mutants, which have been gener-ated for these factors, demonstrate their manifold function and essential impor-tance for mammalian development. Since the cloning of Sp2, which is the less conserved factor among Sp1 - Sp4, no reports about its function, neither in vitro nor in vivo have been published. Therefore, the aim of this thesis work was to unravel Sp2 function by two parallel approaches: a functional molecular characterization (including expression, transactivation and DNA binding studies) and the generation of Sp2 gene targeted mice. To study the Sp2 protein at the molecular level, Sp2-specific rabbit polyclonal anti-bodies were generated. Sp2 protein, which is exclusively localized to the nucleus, was detected in all analyzed cell lines and adult mouse tissues, although in differ-ent amounts. This favours at least a widely expression of the transcription factor Sp2. To explore Sp2 transactivation properties, reporter assays were performed with full-length Sp2 protein as well as various Sp2 deletion mutants using different GC- and GT-box-containing promoters. Unlike transcription factor Sp1, which is a strong activator, Sp2 proteins did not activate reporter gene expression. Also, when fusing Sp2 deletions to a heterologous Gal4 DNA binding domain, no activation was detectable. In addition, the DNA binding capacity and specificity of full-length Sp2 protein and a series of Sp2 deletion mutants were investigated by Electropho-retic Mobility Shift Assays. Full-length Sp2 protein was not able to bind to DNA, nei-ther to GC boxes (the “classical” Sp1 binding site) and GC box variants, nor to other DNA binding sequences like GT and CT boxes. However, when deleting the N-terminal amino acids 1-179, GC box binding was possible. These results suggest that the DNA binding activity is regulated in vivo. To unravel the physiological func-tion of transcription factor Sp2, targeted mice were generated. These mice are not viable; they die shortly before or after birth. Whereas Sp2-targeted embryos develop normal until day E12.5, day E18.5 embryos are characterized by a strongly reduced body size and weight, however with strong variations. These results demonstrate a fundamental role of the transcription factor Sp2 for normal mouse development

Transcription factor Sp2: Molecular characterization and generation of Sp2 gene targeted mice

Summary The Sp family of transcription factors is characterised by its DNA-binding domain, an array of three conserved C2H2 zinc fingers. As a consequence of the conserved DNA binding motif, Sp members recognize GC (GGGGCGGGG) and GT (GGTGTGGGG) boxes with similar specificity and affinity. GC and GT boxes are im-portant for the expression of many different ubiquitous as well as tissue-specific cellular and viral genes. To date, nine members of the Sp family (Sp1 - Sp9) have been identified. In addition to their DNA-binding domain, Sp1 - Sp4 also share other structural features like two glutamine-rich transactivation domains and two serine/threonine-rich regions. Molecular and functional properties have been de-scribed for Sp1, Sp3 and Sp4. Mouse deletion mutants, which have been gener-ated for these factors, demonstrate their manifold function and essential impor-tance for mammalian development. Since the cloning of Sp2, which is the less conserved factor among Sp1 - Sp4, no reports about its function, neither in vitro nor in vivo have been published. Therefore, the aim of this thesis work was to unravel Sp2 function by two parallel approaches: a functional molecular characterization (including expression, transactivation and DNA binding studies) and the generation of Sp2 gene targeted mice. To study the Sp2 protein at the molecular level, Sp2-specific rabbit polyclonal anti-bodies were generated. Sp2 protein, which is exclusively localized to the nucleus, was detected in all analyzed cell lines and adult mouse tissues, although in differ-ent amounts. This favours at least a widely expression of the transcription factor Sp2. To explore Sp2 transactivation properties, reporter assays were performed with full-length Sp2 protein as well as various Sp2 deletion mutants using different GC- and GT-box-containing promoters. Unlike transcription factor Sp1, which is a strong activator, Sp2 proteins did not activate reporter gene expression. Also, when fusing Sp2 deletions to a heterologous Gal4 DNA binding domain, no activation was detectable. In addition, the DNA binding capacity and specificity of full-length Sp2 protein and a series of Sp2 deletion mutants were investigated by Electropho-retic Mobility Shift Assays. Full-length Sp2 protein was not able to bind to DNA, nei-ther to GC boxes (the “classical” Sp1 binding site) and GC box variants, nor to other DNA binding sequences like GT and CT boxes. However, when deleting the N-terminal amino acids 1-179, GC box binding was possible. These results suggest that the DNA binding activity is regulated in vivo. To unravel the physiological func-tion of transcription factor Sp2, targeted mice were generated. These mice are not viable; they die shortly before or after birth. Whereas Sp2-targeted embryos develop normal until day E12.5, day E18.5 embryos are characterized by a strongly reduced body size and weight, however with strong variations. These results demonstrate a fundamental role of the transcription factor Sp2 for normal mouse development

Biomasse als Pfeiler der regionalen Energieversorgung

Die Nutzung von Bioenergie ist für Regionen eine Chance; doch nicht jede Gegend ist in gleichem Maße dafür geeignet. Vor dem Hintergrund von Energieszenarien wurden nun erstmals die regionalen Effekte eines Ausbaus der Bioenergienutzung aufgezeigt. Die Ergebnisse können zur Entscheidungsunterstützung vor Ort verwendet werden

Gambaran Motivasi Kerja Dan Pelayanan Prima Perawat Di Rumah Sakit Umum Bethesda Gmim Tomohon

:Motivation of nurses is the important thing because motivation is the one of caused, to lead, and human support for someone to do the best and get the best result. Excellent Service is the first element in the hospital and health sector, that the patients need. The aims of the research to see the picture of work motivation and excellent service of nurses at Bethesda GMIM Tomohon Hospital. Desain research The study was descriptive-qualitative, the population was all of nurses and patients in Bethesda GMIM Tomohon Hospital This research sample 60 respondent (nurse and patient), with the purposive sampling. Data analysis used of this study was data processor software SPSS. The result: The result of the study showed that intrinsic motivation of good category (48,3%), less category (51,7%), extrinsic motivation of good category (50%), less category (50%). Excellent service nurse when first meet the patient of good category (71,7%), less category (28,3%), when will start of good category (53,3%), less category (46,7%), and after they do the excellent service in good category (70%), less category (30%). The conclution more nurses are motivated by extrisic than intrinsic, and patiens feel that is provided nurses already do the excellent service. Advice would be able to add more information to researcher and to the hospital to maintain the excellent service provided to patients

Reliability of the active knee joint position sense test and influence of limb dominance and sex

The output of a sensorimotor performance can be measured with the joint position sense (JPS) test. However, investigations of leg dominance, sex and quality measures on this test are limited. Therefore, these potential influencing factors as well as reliability and consistency measures were evaluated for angular reproduction performance and neuromuscular activity during the active knee JPS test in healthy participants. Twenty healthy participants (10 males; 10 females; age 29 ± 8 years; height 165 ± 39 cm; body mass 69 ± 13 kg) performed a seated knee JPS test with a target angle of 50°. Measurements were conducted in two sessions separated by two weeks and consisted of two blocks of continuous angular reproduction (three minutes each block). The difference between reproduced and target angle was identified as angular error measured by an electrogoniometer. During reproduction, the neuromuscular activity of the quadriceps muscle was assessed by surface electromyography. Neuromuscular activity was normalized to submaximal voluntary contraction (subMVC) and displayed per muscle and movement phase. Differences between leg dominance and sex were calculated using Friedman-test (α = 0.05). Reliability measures including intraclass correlation coefficient (ICC), Bland–Altman analysis (bias ± limits of agreement (LoA)) and minimal detectable change (MDC) were analysed. No significant differences between leg dominance and sex were found in angular error and neuromuscular activity. Angular error demonstrated inter-session ICC scores of 0.424 with a bias of 2.4° (± 2.4° LoA) as well as MDC of 6.8° and moderate intra-session ICC (0.723) with a bias of 1.4° (± 1.65° LoA) as well as MDC of 4.7°. Neuromuscular activity for all muscles and movement phases illustrated inter-session ICC ranging from 0.432 to 0.809 with biases between − 2.5 and 13.6% subMVC and MDC from 13.4 to 63.9% subMVC. Intra-session ICC ranged from 0.705 to 0.987 with biases of − 7.7 to 2.4% subMVC and MDC of 2.7 to 46.5% subMVC. Leg dominance and sex seem not to influence angular reproduction performance and neuromuscular activity. Poor to excellent relative reliability paired with an acceptable consistency confirm findings of previous studies. Comparisons to pathological populations should be conducted with caution

Grenzüberschreitende Zusammenarbeit im Aktionsfeld Energie

Vor dem Hintergrund der bisherigen, stark fossil geprägten Historie steht die Großregion als potenzielle Modellregion grenzüberschreitender Aktivitäten im Kontext der Anforderungen des Klimaschutzes und der Energiewende vor großen Herausforderungen. Im Rahmen des vorliegenden Beitrages werden, aufbauend auf den in Europa diesbezüglich definierten Zielperspektiven, bisherige Handlungsansätze und Projekte des Aktionsfeldes Energie in der Großregion exemplarisch dargestellt. Auf der Basis der dort - teilweise durch die Verfasser selbst - erzielten Erfahrungen werden Handlungsbedarfe skizziert, die sich zum einen auf die notwendige Implementierung von Strukturen und Netzwerken und zum anderen auf die mögliche Entwicklung einer Forschungsexzellenz in dem thematischen Sektor Energie/Klimaschutz beziehen. Eine besondere Rolle wird dabei auch den Kommunen als relevante Akteure der Energiewende zugewiesen.The recent history of the Greater Region has been strongly influenced by fossil fuels. The area faces correspondingly great challenges in terms of the cross-border activities necessary to meet the requirements of climate protection and the energy transition. Based on the targets defined in Europe, this paper presents examples of approaches and projects undertaken in the field of energy in the Greater Region. Experiences in the area - including those of authors - reveal the need for action, related firstly to the implementation of structures and networks, and secondly to the possible development of research excellence in the sector of energy/climate protection. As relevant actors in the energy transition, the municipalities have a particular role to play here

Cherenkov Light Imaging - Fundamentals and recent Developments

We review in a historical way the fundamentals of Cherenkov light imaging applied to Ring Imaging Cherenkov Counters. We also point out some of the newer developments in this very active field.Comment: Submitted to special edition of NIMA, Proceedings of RICH201

Influence of Obesity on Foot Loading Characteristics in Gait for Children Aged 1 to 12 Years

Background Overweight and obesity are increasing health problems that are not restricted to adults only. Childhood obesity is associated with metabolic, psychological and musculoskeletal comorbidities. However, knowledge about the effect of obesity on the foot function across maturation is lacking. Decreased foot function with disproportional loading characteristics is expected for obese children. The aim of this study was to examine foot loading characteristics during gait of normal-weight, overweight and obese children aged 1–12 years. Methods A total of 10382 children aged one to twelve years were enrolled in the study. Finally, 7575 children (m/f: n = 3630/3945; 7.0±2.9yr; 1.23±0.19m; 26.6±10.6kg; BMI: 17.1±2.4kg/m2) were included for (complete case) data analysis. Children were categorized to normal-weight (≥3rd and 97th percentile; n = 371) according to the German reference system that is based on age and gender-specific body mass indices (BMI). Plantar pressure measurements were assessed during gait on an instrumented walkway. Contact area, arch index (AI), peak pressure (PP) and force time integral (FTI) were calculated for the total, fore-, mid- and hindfoot. Data was analyzed descriptively (mean ± SD) followed by ANOVA/Welch-test (according to homogeneity of variances: yes/no) for group differences according to BMI categorization (normal-weight, overweight, obesity) and for each age group 1 to 12yrs (post-hoc Tukey Kramer/Dunnett’s C; α = 0.05). Results Mean walking velocity was 0.95 ± 0.25 m/s with no differences between normal-weight, overweight or obese children (p = 0.0841). Results show higher foot contact area, arch index, peak pressure and force time integral in overweight and obese children (p<0.001). Obese children showed the 1.48-fold (1 year-old) to 3.49-fold (10 year-old) midfoot loading (FTI) compared to normal-weight. Conclusion Additional body mass leads to higher overall load, with disproportional impact on the midfoot area and longitudinal foot arch showing characteristic foot loading patterns. Already the feet of one and two year old children are significantly affected. Childhood overweight and obesity is not compensated by the musculoskeletal system. To avoid excessive foot loading with potential risk of discomfort or pain in childhood, prevention strategies should be developed and validated for children with a high body mass index and functional changes in the midfoot area. The presented plantar pressure values could additionally serve as reference data to identify suspicious foot loading patterns in children

How Big Can Anomalous W Couplings Be?

Conventional wisdom has it that anomalous gauge-boson self-couplings can be at most a percent or so in size. We test this wisdom by computing these couplings at one loop in a generic renormalizable model of new physics. (For technical reasons we consider the CP-violating couplings here, but our results apply more generally.) By surveying the parameter space we find that the largest couplings (several percent) are obtained when the new particles are at the weak scale. For heavy new physics we compare our findings with expectations based on an effective-lagrangian analysis. We find general patterns of induced couplings which robustly reflect the nature of the underlying physics. We build representative models for which the new physics could be first detected in the anomalous gauge couplings.Comment: 40 pages, 11 figures, (dvi file and figures combined into a uuencoded compressed file), (We correct an error in eq. 39 and its associated figure (9). No changes at all to the text.), McGill-93/40, UQAM-PHE-93/03, NEIPH-93-00

Specificity Protein 2 (Sp2) Is Essential for Mouse Development and Autonomous Proliferation of Mouse Embryonic Fibroblasts

BACKGROUND: The zinc finger protein Sp2 (specificity protein 2) is a member of the glutamine-rich Sp family of transcription factors. Despite its close similarity to Sp1, Sp3 and Sp4, Sp2 does not bind to DNA or activate transcription when expressed in mammalian cell lines. The expression pattern and the biological relevance of Sp2 in the mouse are unknown. METHODOLOGY/PRINCIPAL FINDINGS: Whole-mount in situ hybridization of mouse embryos between E7.5 and E9.5 revealed abundant expression in most embryonic and extra-embryonic tissues. In order to unravel the biological relevance of Sp2, we have targeted the Sp2 gene by a tri-loxP strategy. Constitutive Sp2null and conditional Sp2cko knockout alleles were obtained by crossings with appropriate Cre recombinase expressing mice. Constitutive disruption of the mouse Sp2 gene (Sp2null) resulted in severe growth retardation and lethality before E9.5. Mouse embryonic fibroblasts (MEFs) derived from Sp2null embryos at E9.5 failed to grow. Cre-mediated ablation of Sp2 in Sp2cko/cko MEFs obtained from E13.5 strongly impaired cell proliferation. CONCLUSIONS/SIGNIFICANCE: Our results demonstrate that Sp2 is essential for early mouse development and autonomous proliferation of MEFs in culture. Comparison of the Sp2 knockout phenotype with the phenotypes of Sp1, Sp3 and Sp4 knockout strains shows that, despite their structural similarity and evolutionary relationship, all four glutamine-rich members of the Sp family of transcription factors have distinct non-redundant functions in vivo