The Role of Suppressor of Fused in Development and Tumorigenesis in the Mouse

Embryonic development is a process that involves a number of evolutionarily wellconserved signaling cascades, including the hedgehog pathway. Mutations in components of this pathway have been identified in certain developmental disorders, and in many different kinds of cancers. In fact, the most common cancer in the Western World, basal cell carcinoma (BCC) of the skin, is due to mutations that cause aberrantly activated hedgehog signaling. This thesis focuses on a protein known as Suppressor of fused (Sufu), which is an essential tumor suppressor within the hedgehog pathway. In PAPER I, we made the surprising observation that Sufu actually plays a fundamental role in the mammalian hedgehog signaling pathway, in contrast to its role in fruit flies and even zebrafish. In these organisms, Sufu plays an insignificant part in normal hedgehog signaling, since its absence only results in minor phenotypic changes. However, in the mouse, we showed that loss of Sufu leads to embryonic death in midgestation with the embryos exhibiting severe cephalic defects and an open neural tube. We also demonstrated that the Sufu loss-of-function phenotype was due to ligandindependent activation of the hedgehog signaling pathway. In humans, Gorlin syndrome is a rare developmental disorder that in the majority of cases is due to inactivating mutations in the gene that encodes the hedgehog receptor, PTCH1. This leads to overactivated hedgehog signaling, since PTCH1 is no longer able to inhibit the signal transducer, Smoothened (SMO). Gorlin syndrome involves an array of different developmental defects, but it also leads to increased tumor susceptibility, especially in the form of multiple BCCs. In PAPER I we discovered that mice, heterozygous for the Sufu gene, develop features of Gorlin syndrome, including a skin phenotype with BCC-like attributes, in addition to developmental aberrations in the form of jaw keratocysts. In addition, we showed that the extent of epidermal skin changes correlated with increased hedgehog pathway activation. The BCC-like lesions in Sufu+/- mice are reminiscent of basaloid follicular hamartomas (BFH), which are more benign lesions than BCCs. In PAPER II, the aim was to investigate whether the Sufu+/- skin lesions would develop into full-blown BCCs if Trp53 was knocked out simultaneously. Trp53 is a well-known tumor suppressor gene that can enhance hedgehog-driven tumors, and is often mutated in sporadic BCCs, sometimes in combination with PTCH1 mutations. We showed that Sufu+/- mice on a Trp53 null background develop medulloblastomas and rhabdomyosarcomas, which is consistent with previous reports. Surprisingly, however, the Sufu+/- skin phenotype was not altered in the absence of Trp53, and showed no changes in latency, multiplicity, cellular phenotype or proliferative capacity during the lifespan of the mice. This finding suggests a differential, tissue-specific sensitivity to Sufu and Trp53 gene loss, possibly linked to developmental phase and proliferative potential in specific tissues. In PAPER III, we studied developmental and differentiation processes in the absence of Sufu, using embryonic stem cells (ESCs) derived from Sufu-/- preimplantation embryos. Sufu-/- ESCs were found to express typical pluripotency markers, but the activity of the hedgehog pathway was increased only modestly compared to wild-type ESCs, as indicated by Gli1 target gene expression. The Sufu-/- ESCs formed embryoid bodies in vitro, which, in later stages, were smaller than their wild-type counterparts, suggesting a deficiency in proliferation. To test the differentiation capacity of the Sufu-/- ESCs in vivo, the cells were injected subcutaneously into nude mice to form teratomas. Teratomas from Sufu-/- ESCs developed at efficiencies and latencies equivalent to teratomas from wild-type ESCs, yet in stark contrast to wild-type, Sufu-/- teratomas were dominated by neuroectodermal tissues and were deficient in the mesodermal derivatives, cartilage and bone. These findings call attention to the central role played by Sufu in the hedgehog signaling pathway, and propose a function for Sufu in ectodermal-mesodermal cell fate decision. In a PRELIMINARY STUDY, we have generated conditional Sufu mutant mice with the aim of deleting Sufu in specific tissues at specific time-points. These studies are ongoing, and experiments to create mice with complete loss of Sufu in the K5 (basal cell) compartment of the skin have been initiated. In summary, the studies in this thesis highlight an essential role for Sufu in the hedgehog signaling pathway during development and tumorigenesis in mammals

Műalkotások az Országházban 1945 előtt

While planning the Parliament Building architect Imre Steindl designated a minor role to paintings and sculptures, wanting to integrate the interior architecture into the Gothic style of the building. He accepted murals as a compromise but refused to have any oil paintings. However, he failed to execute his vision due to the approach of the Hungarian Millennium of 1896, a series of celebrations commemorating the thousand-year anniversary of the Hungarian conquest of the Carpathian Basin. For this event an oil painting portraying the conquest was ordered from Mihály Munkácsy, as well as a pair of statues depicting Franz Joseph I, and his wife, Elisabeth, with the intention of displaying them in the Parliament. Further orders were made in 1902, when the lawmakers occupied the building. From the very beginning they wanted to decorate their rooms and offices with paintings, sculptures, and works of applied art. As representations of their positions, they usually displayed portraits of their predecessors, or important events and figures of Hungarian history and law-making. In the 1920s and 1930s the number of artworks in the Parliament significantly increased due to two Speakers of the National Assembly, Béla Scitovszky and Tibor Zsitvay. Their contributions included the galleries depicting the former speakers and first officers of the National Assembly, the large tapestry by Gyula Rudnay depicting the national assembly in Ópusztaszer, the painting by Gyula Benczúr about the national Assembly paying respect to the king at the Millennium, as well as the depictions of monarchs (Franz Joseph I, Charles IV) and statesmen (István Széchenyi, Lajos Kossuth, Ferenc Deák, Miklós Horthy). This study discusses the history of the most exceptional works of art among these until the middle of the 1940s

Problem Solving Skills Deconstructed And Implemented In An Adaptive Learning Tool

The development of problem-solving skills is an important subject in engineering curricula. Helping novice students develop such skills can be challenging because problem solving is a complex skill in the sense that it is accompanied with an internal thinking process that many experts are even unaware of doing. From a combination of literature and a thinking-aloud exercise with the entire teaching team, a scheme with building blocks and strategies that are commonly used by engineers was constructed. In addition to commonly named steps such as Identify/Define, Plan/Choose, Carry Out/Do and Look back/Inspect the scheme refines the first step into multiple interdependent building blocks, emphasizes the need for critical reflection at each point as well as the possible need to return to previous steps at any time. Moreover, multiple correct solution paths can be followed in solving a problem. To address this and to empower the students in their divergent thinking processes when solving a problem, an innovative intra-exercise adaptive e-learning tool was created. The anywhere-anytime availability enables for virtual and remote learning in the post-COVID world. In the learning tool students can choose between different solution paths, after firstly identifying the correct context, parameters etc. This paper describes the process of defining the building blocks, resulting strategy scheme and implementation of the building blocks in the adaptive e-learning tool. Initial findings indicate that the strategy scheme consisting of building blocks and the adaptive e-learning tool help students in developing their problem-solving skills

Construction of genomic library of rice using a bacteriophage lambda vector (Agricultural Chemistry)

バクテリオファージラムダベクターEMBL 3を用いた遺伝子ライブラリーの作製法を植物のDNAに対して効率が上がるように改変した。核DANはイネ胚芽より調製したが, 制限酵素の基質とするためには, セシウムクロライド平衡密度勾配遠心で精製する必要があった。単離したDNAはMbo Iで部分分解し, 主にFrischaufら(J. Mol. Biol. 170 827 (1983))の方法により, バクテリオファージラムダベクターEMBL 3アームと結合した。バクテリオファージDNAとイネ胚芽DNAの結合効率は, ポリエチレングリコール(PEG)6000の添加で上昇した。更に, PEG6000はin vitroパッケージングの効率も上げた。結合反応とパッケージングに対するこの改良された方法は, 植物の遺伝子ライブラリー作製に一般的に適用することが可能であろう。The method for construction of genomic library using a bacteriophage lambda vector EMBL 3 was modified so as to increase its efficiency for plant DNA. Genomic DNA was prepared from rice embryos, but to be a substrate of a restriction enzyme it had to be purified by equilibrium centrifugation in a cesium chloride density gradient. The isolated DNA was partially digested with Mbo I, then ligated with bacteriophage lambda vector EMBL 3 arms substantially according to Frischauf et al. (J. Mol. Biol. 170,827 (1983)). The efficiency for ligation between the bacteriophage DNA and the rice embryo DNA was improved by addition of polyethylen glycol (PEG) 6000. Moreover the use of PEG 6000 raised the efficiency of in vitro packaging. This improved procedure for the ligation and packaging of the recombinant DNA will be generally applicable to the construction of plant genomic libraries

Correlation of ERPF by blaufox model according to mono-and two- compartmental model

Uspoređivali smo vrijednost klirensa J 131 hipurana, određenog iz retencione krivulje, i jednog uzorka krvi po modelu s jednini odjeljkom, sa klirensom određenim iz retencione krivulje i dva uzorka krvi po modelu sa dva odjeljka. Uzajamnu ovisnost promatranih metoda najbolje smo opisali funkcijom polinoma IV. stupnja, gdje je koeficijent korelacije iznosio r = 0,82. Posebno smo obradili grupu bolesnika sa niskim vrijednostima klirensa, jer je poznato da monokompartmentalni model upravo ove precjenjuje. Korelacija ovih dviju metoda u niskom području je linearna sa koeficijentom korelacije r = 0,91.We have compared the values of J 131 hippuran clearence obtained from two methods. First is the retention curve and a single blood sample technique according to monocompartmental model. The second method is the retention curve and two blood sample technique according to the two-compartmental model. To describe this dependence most properly we have used the function of the polinom of the fourth degree where the coeffitient of correlation was r = 0.82. Also we have examined groups of patients with the low values of J 131 hippuran clearence, because it is known that the monocompartmental model overestimates regarded values. The correlation between these two methods in the low range is also linear with the coeffitient of correlation r = 0.91

Male 46, XX, clinical case report

USMF Nicolae Testemiţanu, IMSP Institutul Mamei şi Copilului, Centrul Medical RepromedПоловой детерминизм у человека представляет собой сложный процесс, включающий систему генов полового развития, из которых ген, расположенный на хромосоме Y – Yp11.32, именуемый SRY (Sex-determining region Y), играет критическую роль. Цель: Подтвердить важность клинико-генетического обследования для мужчин с азооспермией, с целью точной диагностики мужского бесплодия. Материалы и методы: Было проведено клинико-генетический обследование мужчины в возрасте 31 года, с бесплодием в течение 5 лет в анамнезе, с азооспермией и нарушением гормональных маркёров. Было осуществлено цитогенетическое обследование, проведённое по классической методике с окраской по Гимзе. Дополнительно был применён метод FISH (Fluorescence in situ hybridization) с зондами для X-хромосомы (DXZ1, Xp11.1-q11.1) и Y-хромосомы (SRY, Yp11.32 и DYZ1,Yq12). Для молекулярно-генетической диагностики маркёров Y-хромосомы (SY 81, SY 84, SY 127, SY 134, SY 254, SY 255) и внутреннего контроля маркёров sY14/SRY и ZFX/ZFY была использована техника мультиплексной ПЦР (полимеразная цепная реакция). Результаты: Клиническое обследование показало, что фенотип и психологическая идентичность соответствовали мужскому полу. В то же время, были обнаружены гипогонадизм, азооспермия и бесплодие. Данные цитогенетического обследования показали наличие кариотипа 46, XX, характерного для женского пола. Тест FISH показал наличие двух сигналов хромосомы X и одного сигнала хромосомы Y (SRY, Yp11.32). По результатам молекулярно-генетического анализа было диагностировано наличие гена SRY и отсутствие маркёров Y-хромосомы. Вышеуказанные результаты указывают на клинический диагноз нарушений полового развития (синдром XX, мужской фенотип). Наличие гена SRY, транслоцированного на хромосому X, подтверждает и объясняет мужской фенотип у пациента. Заключение: Клинико-генетическое обследование мужчин с азооспермией представляет важность для точной диагностики мужского бесплодия.Sex determination in humans presents a complex process, that involves a network of genes in which a gene, located on Y-chromosome Yp11.32, named SRY (sex-determining region of the Y), plays a crucial role. The goal: To confirm the importance of clinical-genetic evaluation in azoospermic men, for the correct diagnosis of male infertility. Materials and methods: A 31-year old man with a history of 5-years of infertility, azoospermic and hormonal marker changes, was clinical genetic evaluated. For the chromosome analysis, the cytogenetic exam was performed using the classic Giemsa marking technique. The FISH (Fluorescence in situ hybridization) test with X chromosome probes (DXZ1, Xp11.1-q11.1) and Y (SRY, Yp11.32 and DYZ1, Yq12) was also performed. For molecular genetic diagnosis of Y chromosome markers (SY 81, SY 84, SY 127, SY 134, SY 254, SY 255) and internal controls: sY14/SRY and ZFX/ ZFY, was used technique Multiplex Polymerase chain reaction (PCR).Results: Following clinical evaluation, the patient’s phenotype and psychological identity were male. Also he presented hypogonadism, azoospermia and infertility. According to the cytogenetic exam, the karyotype was 46,XX, female characteristic. The FISH test highlighted, two signals for chromosome X and a signal for chromosome Y (SRY, Yp11.32). Following molecular genetic examination, the presence of the SRY gene and lack of markers of the Y chromosome was diagnosed. The three related results suggest the clinical diagnosis of sexual development disorder (XX syndrome, male phenotype). The presence of the SRY gene translocated on chromosome X confirms and explains the male phenotype of the patient. Conclusion: Clinical-genetic evaluation of azoospermic men is important for the correct diagnosis of male infertility