Staufen1 is a double-stranded RNA-binding protein known to be involved in the transport, localization, decay and increased translation of some mRNAs. The goal of the present study is to determine the role of Staufen1 during myogenic differentiation by characterizing the effects of Staufen1 over-expression in C2C12 cells. Immunofluorescence experiments revealed that Staufen1 over-expression causes a decrease in the fusion and differentiation indices and leads to the formation of myotubes with significantly fewer nuclei. We show, by western blot and qRT-PCR, that the protein expression of MyoD, myogenin and MyHC and the mRNA expression of MyoD, myogenin, Mef2A, Mef2C and p35 are significantly decreased during differentiation when Staufen1 is over-expressed. We then found that c-myc protein expression was increased during proliferation but that its mRNA expression remained unchanged. In this study we propose that Staufen1 negatively regulates skeletal muscle differentiation through the posttranscriptional regulation of c-myc, Mef2A, Mef2C and p35 transcripts.

No description available.

Neuroblastoma (NB) is the most common extracranial solid tumor diagnosed in the first year of life. The disease is characterized by lack of somatic mutations and high genomic instability including the amplification of MYCN which correlates with poor clinical outcome. These features of NB suggest a pivotal role of transcriptome dynamics in this disease. Analyses of 1089 NB patients show that an overall increase in m6A-regulatory factors correlates with the worst prognosis. Notably, the m6A reader YTHDF1 is upregulated in stage 4 NB corresponding to the highest degree of MYCN amplification. In line with these notions, we observed a positive correlation between MYCN and YTHDF1 by analyzing 33 NB cell lines with different MYCN amplification status. Consistent with this observation, we demonstrated that MYCN knockdown is accompanied by a decrease in all the YTHDF proteins. Furthermore, we demonstrate a loss of proliferating activity in a MYCN-amplified cell line upon knocking out all the YTHDF paralogs and show that the loss in proliferation is mediated by downregulation of DNA replication and chromatin remodeling upon YTHDF depletion. Retinoic acid (RA) is a key agent employed in NB therapy; it induces cell differentiation limiting the proliferation of malignant cells. However, therapeutic protocols adopting RA are limited to maintenance therapy. Thereby, identifying new therapeutic strategies that synergistically enhance RA response is essential to expand the therapeutic window for this drug. Since m6A is a well-known regulator of cellular differentiation, we hypothesized that the protein involved in m6A regulation could act in concert with RA in driving this process. We demonstrate that YTHDF depletion potentiates the response to RA enriching the key processes required for differentiation and that this correlates with an enhanced pro-neural of the YTHDF depleted cells. Together, our findings suggest that targeting the YTHDF family in MYCN-amplified NB represents a promising therapeutic opportunity to control the progression of this disease.

The purpose of the present study was to examine differentiation of self (Bowen, 1985) as a predictor of involvement in specific extramarital behaviors. Data were collected from 125 graduates of a comprehensive state university in southeastern United States. The sample was primarily caucasian, college educated, married, politically moderate to slightly conservative, equally divided by gender and ranged in age from 25 to 87 years (M = age 43). Differentiation of self was operationalized and measured by the Differentiation of Self Inventory (Skowron & Friedlander, 1998). Logistical regression analyses regressed involvement in specific extramarital behaviors upon the level of differentiation of self as measured by the Differentiation of Self Inventory. The level of differentiation did not significantly predict the likelihood of involvement in any specific extramarital behavior. Among females, however, differentiation was positively but not significantly associated with the likelihood of involvement in most of the specific extramarital behaviors examined by the study. For females, differentiation did significantly predict the likelihood of having an experience kissing some one other than a spouse while married. / Ph. D.

Lai, Ka Hang. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 114-121). / Abstracts in English and Chinese. / Abstract --- p.i / 論文摘要 --- p.iv / Acknowledgements --- p.vi / Publications based on work in this thesis --- p.vii / Abbreviations --- p.viii / Contents --- p.x / Chapter Chapter1 --- General introduction --- p.1 / Chapter 1.1 --- Cell differentiation --- p.1 / Chapter 1.1.1 --- S tem cell treatments --- p.2 / Chapter 1.1.2 --- Differentiation therapy for cancer --- p.3 / Chapter 1.2 --- Cyclic adenosine monophosphate (cAMP) signaling involved in cell differentiation --- p.4 / Chapter 1.2.1 --- cAMP -signaling pathways leading to transcription activities --- p.4 / Chapter 1.2.1 --- Regulation of cell differentiation by cAMP/PKA signal --- p.5 / Chapter 1.3 --- Aim of thesis --- p.5 / Chapter Chapter2 --- "Materials, media, buffers and solutions" --- p.7 / Chapter 2.1 --- Mate rials --- p.7 / Chapter 2.2 --- "Culture media, buffer and solutions" --- p.12 / Chapter 2.2.1 --- General culture buffers --- p.12 / Chapter 2.2.2 --- Culture medium --- p.12 / Chapter 2.2.3 --- Assay buffers and solutions --- p.13 / Chapter 2.2.3.1 --- Buffers and solutions for RT-PCR --- p.13 / Chapter 2.2.3.2 --- Buffers and solutions for assay of [3H]cAMP production --- p.13 / Chapter 2.2.3.3 --- Buffers and solutions for Western blotting --- p.14 / Chapter 2.2.3.4 --- Buffers and solutions for histamine assay --- p.16 / Chapter 2.2.3.5 --- Buffers and solutions for flow cytometry --- p.17 / Chapter Chapter3 --- Methods --- p.18 / Chapter 3.1 --- Maintenance of rat pheochromocytoma (PC12) cells --- p.18 / Chapter 3.2 --- Dete rmination of AC isoforms expression in PC12 cells by RT-PCR analysis --- p.19 / Chapter 3.2.1 --- RNA isolation --- p.19 / Chapter 3.2.2 --- cDNA synthesis by reverse transcription (RT) --- p.20 / Chapter 3.2.3 --- Semi-quantitative PCR --- p.21 / Chapter 3.3 --- Maintenance of human erythroleukemia (HEL) cells --- p.23 / Chapter 3.4 --- Dete rmination of [3H]cAMP Production in HEL cells --- p.23 / Chapter 3.4.1 --- Principle of assay --- p.23 / Chapter 3.4.2 --- Column preparation --- p.24 / Chapter 3.4.3 --- Measurem ent of [3H]cAMP production in HEL cells --- p.24 / Chapter 3.4.4 --- Data analysis --- p.25 / Chapter 3.5 --- Im munodetection of STAT3 and pTyr705STAT3 by western blotting --- p.25 / Chapter 3.6 --- Harvesting of HE L cells after differentiation treatment --- p.27 / Chapter 3.7 --- Flow cyto metry analysis of HEL cells --- p.27 / Chapter 3.7.1 --- F ITC-conjugated CD41 -antibody staining --- p.28 / Chapter 3.7.2 --- P I staining --- p.28 / Chapter 3.8 --- Determination of extracellular and intracellular histamine of HEL cells --- p.29 / Chapter 3.8.1 --- Sample preparation --- p.29 / Chapter 3.8.2 --- Automated assay of histamine content --- p.30 / Chapter 3.9 --- siRNA mediated knockdown of STAT3 in HEK293 cells --- p.30 / Chapter 3.9.1 --- Culture human embryonic kidney (HEK293) cells --- p.30 / Chapter 3.9.1 --- siRNA transfection --- p.31 / Chapter Chapter4 --- mRNA expression of adenylyl cyclase isoforms during early stage of NGF-induced differentiation of PC12 cells --- p.33 / Chapter 4.1 --- Introduction --- p.33 / Chapter 4.1.1 --- Dif ferentiation of PC12 cells --- p.33 / Chapter 4.1.1.1 --- Induction of neurite outgrowth by NGF in PC12 cells --- p.33 / Chapter 4.1.1.2 --- Effect of cAMP on NGF-induced neurite outgrowth in PC12 cells --- p.34 / Chapter 4.1.1.3 --- Effect of cAMP on NGF-induced neurite outgrowth in PC12 cells --- p.35 / Chapter 4.1.2 --- Enhanced forskolin-stimulated [3H]cAMP productionin NGF-difFerentiated PC12 cells --- p.36 / Chapter 4.1.3 --- Classification of adenylyl cyclases --- p.38 / Chapter 4.1.4 --- Aims of study --- p.39 / Chapter 4.2 --- Results and discussion --- p.40 / Chapter Chapter5 --- Effect of cicaprost on PMA-mediated differentiation of human erythroleukemia (HEL) cells --- p.48 / Chapter 5.1 --- Introduction --- p.48 / Chapter 5.1.1 --- Differentiation of HEL cells --- p.48 / Chapter 5.1.2 --- Prostac yclin (PGI2) and human IP receptors --- p.49 / Chapter 5.1.3 --- Agonists and antagonists of IP receptors --- p.50 / Chapter 5.1.4 --- IP signaling in HEL cells --- p.52 / Chapter 5.1.5 --- Effect of cAMP on megakaryocytic differentiation --- p.52 / Chapter 5.1.6 --- Aims of study --- p.54 / Chapter 5.2 --- Results and discussion --- p.56 / Chapter 5.2.1 --- Preliminar y studies --- p.56 / Chapter 5.2.1.1 --- PMA induced cell adhesion and morphological change --- p.56 / Chapter 5.2.1.2 --- Cell proliferation and protein content --- p.57 / Chapter 5.2.1.3 --- IP signaling in HEL cells --- p.57 / Chapter 5.2.1.4 --- Presence of histaminase in FBS --- p.60 / Chapter 5.2.1.5 --- Summary of preliminary studies --- p.61 / Chapter 5.2.2 --- PMA -induced cell spreading of HEL cells --- p.63 / Chapter 5.2.3 --- PMA -induced DNA synthesis of HEL cells --- p.65 / Chapter 5.2.4 --- PMA -induced cell size and cell complexity of HEL cells --- p.67 / Chapter 5.2.5 --- PMA -induced CD41/CD61 expression of HEL cells --- p.69 / Chapter 5.2.6 --- PMA -induced histamine production of HEL cells --- p.72 / Chapter 5.2.7 --- IP receptor-dependent and IP receptor-independent actions of cicaprost --- p.74 / Chapter 5.2.8 --- STAT3 knockdown by siRNA --- p.75 / Chapter 5.3 --- Role of STAT3 in MK differentiation --- p.76 / Chapter 5.4 --- Summary --- p.78 / Chapter Chapter6 --- General discussions and future study --- p.105 / Chapter 6.1 --- General discussions --- p.105 / Chapter 6.2 --- Future study --- p.111 / References --- p.114

SRY (Sex determining Region of Y chromosome) is the master gene initiating testis determination in mammals. To shed light on the molecular mechanism of SRY action during testicular differentiation, we examined the effects of TAT-HMG fusion protein on gonadal sex differentiation in culture. HMG is the DNA binding motif of SRY and "TAT" is a protein transduction domain. Each pair of CD1 mouse gonadal primordia at 11.5 days post coitum (dpc) was cultured with or without TAT-HMG dissolved in dimethyl sulfoxide (DMSO) up to 3 days. Immunocytochemical labeling and Real-time RT-PCR of Sry, Sox9 and Mis indicated that DMSO blocked testicular differentiation, Sertoli cell differentiation and testis cords formation, downstream of SRY. TUNEL showed a massive mesenchymal cell death, which might be responsible for disruption of testis cord formation. Treatment with TAT-HMG rescued Sertoli cell differentiation, probably by up regulation of Sry, but not testis cord formation or cell death.

Thesis (M.S.)--University of Michigan, 1989.

Thesis (M.S.)--University of Michigan, 1989.

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第19569号 / 医博第4076号 / 新制||医||1017(附属図書館) / 32605 / 京都大学大学院医学研究科医学専攻 / (主査)教授 江藤 浩之, 教授 武藤 学, 教授 中畑 龍俊 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

No description available.