Glioblastoma multiforme (GBM) is the most frequent primary malignancy of the central nervous system. Despite remarkable progress towards understanding of tumor biology, there is no efficient treatment and patient outcome remains poor. Here we present a unique Nanobody-based anti-proteomic approach for selection of Nanobodies specific for over-expressed glioblastoma proteins. Phage-displayed Nanobody library was enriched on protein extracts of NCH644 and NCH421K glioblastoma cell lines. Differential ELISA screenings revealed seven Nanobodies targeting the following cognate antigens: ACTB/NUCL complex, VIM, NAP1L1, TUFM, DPYSL2, CRMP1 and ALYREF. Validation with qRT-PCR showed significant changes in VIM mRNA levels (****P(0.001) in glioblastoma vs. reference samples. Western blots showed greatest protein up-regulation for ALYREF, CRMP1 and VIM. Moreover, bioinformatic analysis with OncoFinder software and against the complete TCGA brain tumor gene expression dataset suggested involvement of differential proteins in WNT and ATM pathways, and Aurora B, Sem3A and E-cadherin signaling. Notably, we demonstrate the potential use of NAP1L1, NUCL, CRMP1, ACTB and VIM for differentiation between glioblastoma and WHO Grade II and III gliomas, and point out DPYSL2 as perfect glioma vs. normal biomarker. Our study contributes to filling in the science gap in this field by indicating novel differences in the proteomic profiles between gliomas and reference samples. We present possible use of the selected proteins for distinguishing glioblastoma from WHO Grade II and III gliomas, as well as from reference samples. These discoveries could be of benefit for glio(blasto)ma patients after being validated on a larger sample size.
COBISS.SI-ID: 32777177
Mice deficient in the nuclear hormone receptor ROR[gamma]t have defective development of thymocytes, lymphoid organs, Th17 cells, and type 3 innate lymphoid cells. ROR[gamma]t binds to oxysterols derived from cholesterol catabolism, but it is not clear whether these are its natural ligands. Here, we show that sterol lipids are necessary and sufficient to drive ROR[gamma]t-dependent transcription. We combined overexpression, RNAi, and genetic deletion of metabolic enzymes to study ROR[gamma]-dependent transcription. Our results are consistent with the ROR[gamma]t ligand(s) being a cholesterol biosynthetic intermediate (CBI) downstream of lanosterol and upstream of zymosterol. Analysis of lipids bound to ROR[gamma] identified molecules with molecular weights consistent with CBIs. Furthermore, CBIs stabilized the ROR[gamma] ligand-binding domain and induced coactivator recruitment. Genetic deletion of metabolic enzymes upstream of the ROR[gamma]t-ligand(s) affected the development of lymph nodes and Th17 cells. Our data suggest that CBIs play a role in lymphocyte development potentially through regulation of ROR[gamma]. Argumentation: This manuscript represents the first proof that post lanosterol intermediates of cholesterol synthesis have independent biological roles that are not directly connected to cholesterol.
COBISS.SI-ID: 3506824
We demonstrate unequivocally that defective cholesterol synthesis is an independent determinant of liver inflammation and fibrosis. We prepared a mouse hepatocyte-specific knockout (LKO) of lanosterol 14 a -demethylase (CYP51) from the part of cholesterol synthesis that is already committed to cholesterol. LKO mice developed hepatomegaly with oval cell proliferation, fibrosis and inflammation, but without steatosis. The key trigger was reduced cholesterol esters that provoked cell cycle arrest, senescence-associated secretory phenotype and ultimately the oval cell response, while elevated CYP51 substrates promoted the integrated stress response. In spite of the oval cell-driven fibrosis being histologically similar in both sexes, data indicates a female-biased down-regulation of primary metabolism pathways and a stronger immune response in males. Liver injury was ameliorated by dietary fats predominantly in females, whereas dietary cholesterol rectified fibrosis in both sexes. Our data place defective cholesterol synthesis as a focus of sex-dependent liver pathologies. Explanation: The paper shows that defected hepatic cholesterol synthesis at the level of Cyp51 leads to a disease phenotype that is similar to nonalcoholic hepatitis.
COBISS.SI-ID: 31858137
This is the last current article of in a series of 4 papers published in 2017 and 2016 in Scientific Reports after the first paper from 2014 (see above, point 3) and dealing with the relationship of cholesterol biosynthesis / metabolism and liver pathology. Summary of the content of this article is as follows: Development of mice with hepatocyte knockout of lanosterol 14[alpha]-demethylase (H[sup]Cyp51-/-) from cholesterol synthesis is characterized by the progressive onset of liver injury with ductular reaction and fibrosis. These changes begin during puberty and are generally more aggravated in the knockout females. However, a subgroup of (pre)pubertal knockout mice (runts) exhibits a pronounced male prevalent liver dysfunction characterized by downregulated amino acid metabolism and elevated Casp12. RORC transcriptional activity is diminished in livers of all runt mice, in correlation with the depletion of potential RORC ligands subsequent to CYP51 disruption. Further evidence for this comes from the global analysis that identified a crucial overlap between hepatic Cyp51-/- and Rorc-/- expression profiles. Additionally, the reduction in RORA and RORC transcriptional activity was greater in adult H[sup]Cyp51-/- females than males, which correlates well with their downregulated amino and fatty acid metabolism. Overall, we identify a global and sex-dependent transcriptional de-regulation due to the block in cholesterol synthesis during development of the Cyp51 knockout mice and provide in vivo evidence that sterol intermediates downstream of lanosterol may regulate the hepatic RORC activity. The other three papers are as follows: Yurina Shishido, Takashi Baba, Tetsuya Sato, Yuichi Shima, Kanako Miyabayashi, Miki Inoue, Haruhiko Akiyama, Hiroshi Kimura, Yoshiakira Kanai, Yasuhiro Ishihara, Shogo Haraguchi, Akira Miyazaki, Damjana Rozman, Takeshi Yamazaki, Man-Ho Choi, Yasuyuki Ohkawa, Mikita Suyama, Ken-Ichirou Morohashi: Differential lactate and cholesterol synthetic activities in XY and XX Sertoli cells. Scientific Reports, ISSN 2045-2322, 2 Feb. 2017, vol.7: 41912, 1-14; https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5288785/pdf/srep 41912.pdf; doi: 10.1038/srep4191. [COBISS.SI-ID 33091545], [JCR, SNIP], JCR IF (2015): 5.228. Rok Košir, Uršula Prosenc, Anja Korenčič, Peter Juvan, Jure Ačimovič, Damjana Rozman: Mouse genotypes drive the liver and adrenal gland clocks. Scientific Reports, ISSN 2045-2322, 18 Aug. 2016, vol. 6, pp. 1-13 (31955). http://www.nature.com/articles/srep31955, doi: 10.1038/srep31955. [COBISS.SI-ID 32796121], [JCR, SNIP]; JCR IF (2015): 5.228. Jure Ačimovič, Sandeep Goyal, Rok Košir, Marko Goličnik, Martina Perše, Aleš Belič, Žiga Urlep, Peter F. Guengerich, Damjana Rozman: Cytochrome P450 metabolism of the post-lanosterol intermediates explains enigmas of cholesterol synthesis. Scientific Reports, ISSN 2045-2322, 23 Jun. 2016, vol. 6, pp. 1-15 (28462); http://www.nature.com/articles/srep28462, doi: 10.1038/srep28462. [COBISS.SI-ID 32717273], [JCR, SNIP, WoS, Scopus], JCR IF (2015): 5.228.
COBISS.SI-ID: 33091801
Article Summary: Life sciences are yielding huge data sets that underpin scientific discoveries fundamental to improvement in human health, agriculture and the environment. In support of these discoveries, a plethora of databases and tools are deployed, in technically complex and diverse implementations, across a spectrum of scientific disciplines. The corpus of documentation of these resources is fragmented across the Web, with much redundancy, and has lacked a common standard of information. The outcome is that scientists must often struggle to find, understand, compare and use the best resources for the task at hand. Here we present a community-driven curation effort, supported by ELIXIR-the European infrastructure for biological information-that aspires to a comprehensive and consistent registry of information about bioinformatics resources. The sustainable upkeep of this Tools and Data Services Registry is assured by a curation effort driven by and tailored to local needs, and shared amongst a network of engaged partners. As of September 2015, the registry includes 1633 resources, with depositions from 91 individual registrations including 40 institutional providers and 51 individuals. With community support, the registry can become a standard for dissemination of information about bioinformatics resources: we welcome everyone to join us in this common endeavour.
COBISS.SI-ID: 32290009