1,529 research outputs found
The effect of beam inclination on the performance of a passive vibration isolator using buckled beams
No full textPassive vibration isolators are desired to have both high static stiffness to support large static load and low local stiffness to reduce the displacement transmissibility at frequencies greater than resonance. Utilization of a vertical buckled beam as a spring component is one way to realize such a stiffness characteristic since it exhibits a smaller ratio of local stiffness to static stiffness than that of a linear spring. This paper investigates the behaviour of a vibration isolator using inclined beams as well as vertical ones and examines the effect of beam inclination for the purpose of improving the isolation performance. The experimental system investigated has anisolated mass which is supported by a combination of two types of beams: buckled beams and constraining beams. The buckled beams can be inclined from the vertical by attachment devices, and the constraining beams are employed to prevent off-axis motion of the isolated mass. The results demonstrate that the inclination of the buckled beams reduces the resonance frequency and improves the displacement transmissibility at frequencies greater than resonance
Isoform-specific roles of the adaptor protein ShcA in cell signaling
No full textShcA is a bona fide adaptor protein without
any enzymatic activity. Upon activation of
receptor tyrosine kinases, ShcA associates
with the receptor and becomes tyrosine
phosphorylated. Phosphorylated ShcA recruits
the Grb2/SOS complex to the membrane,
where SOS stimulates the small GTPase Ras,
resulting in the activation of the Ras/MAPK
pathway. The fact that Grb2 binds directly to
most of the receptor tyrosine kinases raises
the question of how important is the role of Shc
in mediating MAPK activation? Moreover,
beside growth factor-induced MAPK activation,
are there other pathways in which ShcAmediated
MAPK activation is relevant?
ShcA is expressed in three different
isoforms: p46Shc, p52Shc, and p66Shc. These
isoforms are all derived from a single gene and
differ only in their N-terminal part. Although all
isoforms are phosphorylated by receptor
tyrosine kinases, and subsequently bind to
Grb2, the p66Shc isoform does not seem to
mediate MAPK activation. The individual
contribution of p46Shc and p52Shc in mediating
MAPK activation is also not clear. The fact that
all isoforms are ubiquitously expressed, with
some restrictions for p66Shc, complicates the
experimental investigation of each isoform.
Recently, p66Shc has been implicated in the
regulation of apoptosis in response to oxidative
stress.
Using siRNA, we established a system which
allows isoform-specific knockdown of ShcA
proteins in tissue culture. Further development
of this technique enabled us to express a
single isoform in the absence of endogenous
protein. This so-called “knockdown-in”
technique is applicable for most proteins which
are expressed in multiple isoforms, and allows
the investigation of specific mutations against
a clear background without overexpression.
We used this technique to investigate the
contribution of individual ShcA isoforms to
EGF-induced MAPK activation in epithelial
cells. Knockdown of all or single ShcA
isoforms had no effect on EGF-induced Erk
activation. Moreover, overexpression of p66Shc
in non p66Shc-expressing MCF7 cells did not
change EGF-induced proliferation or viability.
These data suggest that EGF-induced MAPK
activation in epithelial cells is ensured by a
redundant coupling of Grb2 to the receptor.
In a quest for growth factor-independent
pathways involving Shc-mediated Erk
activation, we investigated signaling
downstream of the cell-cell adhesion molecule
E-cadherin. We identified a previously
unknown signaling pathway which is induced
upon disruption of E-cadherin-dependent cellcell
adhesion This pathway involves Src- and
Shc-dependent Erk activation, which results
subsequently in the expression of the
urokinase plasminogen activator. Applying the
knockdown-in technique revealed that p46Shc
and p52Shc, but not p66Shc, were able to
mediate MAPK activation upon disruption of
cell-cell adhesion. This pathway directly links
disruption of cell-cell adhesion with the
expression of proteolytic enzymes, both
processes involved in metastasis and wound
healing.
To learn more about the role of p66Shc in
mediating oxidative stress-induced apoptosis
in epithelial cells, the effect of p66Shc on cell
viability was investigated. Although p66Shc has
been shown to enhance stress-induced
apoptosis in fibroblasts, endothelial cells, and
T-cells, no effect on p66Shc expression was
observed in two different epithelial cells,
suggesting that the apoptotic response in
epithelial cells is mediated in a p66Shc-
independent manner
Caveolar dysfunction leads to signal transduction defects that are critical for obesity-driven disorders
No full textTo say that obesity and diabetes have reached epidemic proportions has become something of a cliché. This should not lead us to simply accept it as a byproduct of our changing lifestyle, or to overlook the socioeconomic importance of these conditions.
Our attempts at therapeutic intervention have been hindered by a lack of knowledge about the precise pathophysiological mechanisms via which obesity triggers secondary disorders such as diabetes and cardiovascular problems. Our approach to understanding the mechanisms have been largely focused on single molecules whose levels or activity are altered in obesity, and which are known independently to contribute to these secondary disorders. Although we have accumulated genetic and biochemical evidence for the potential role of these factors in these disorders, it must be noted that no single candidate or mechanism has yet given a satisfactory explanation for all the obesity-related disorders. This has led researchers to conclude that the underlying pathology is polygenic or multi-factorial. However, based on our experiments and the recent findings of a few other laboratories, we propose a unifying theory where obesity leads to a membrane microdomain disorder that will in turn lead to the corruption of multiple signaling pathways that are known to be implicated in insulin resistance and cardiovascular disorders. We also explore the possibility of pharmacologically modulating a nodal, but downstream, drug target that might retard or prevent these disorders. Having said that, I would like to clearly state that this theory is still in its infancy, and remains to be tested in both animal models and human subjects
Cytokine mediated immune responses in the Japanese pufferfish (Takifugu rubripes) administered with heat-killed Lactobacillus paracasei spp. paracasei (06TCa22) isolated from the Mongolian dairy product
No full textCitation:
Biswas G, Korenaga H, Nagamine R, Kawahara S, Takeda S, Kikuchi Y, Dashnyam B, Yoshida T, Kono T, Sakai M. Cytokine mediated immune responses in the Japanese pufferfish (Takifugu rubripes) administered with heat-killed Lactobacillus paracasei spp. paracasei (06TCa22) isolated from the Mongolian dairy product. Int Immunopharmacol. 2013 Oct;17(2):358-65. doi: 10.1016/j.intimp.2013.06.030. Epub 2013 Jul 15. PMID: 23867289
Investigation of the guanine quadruplex resolving activity of the DEAH-box RNA helicase RHAU
No full textGenerally depicted as single- or double-stranded molecules, nucleic acids sequences can adopt various forms of stable secondary structures. In particular, guanine-riche sequence of DNA and RNA can form atypical four-stranded helical structures termed G-quadruplexes (G4). Although the functional relevance of G4 structures is still mater of debate, an increasing body of evidence suggests that these structures can form in various regions of the genome and may be implicated in a wide array of processes such as gene expression regulation and telomere protection. Owing to their high-thermodynamic stability, in vivo conversion of G4 structures to single-stranded nucleic acid requires specialised proteins with G4 destabilising/unwinding activity. RHAU is a human RNA helicase of the DEAH-box family that exhibits ATP-dependent G4 resolving activity with high affinity and specificity for its substrate in vitro. However, how RHAU recognises G4 and what are its substrates in cells are key questions that needed to be addressed.
In the first part of this research work, we undertook to address the molecular mechanisms underlying the specific recognition of G4 structures by RHAU. Through biochemical analysis of truncated and mutated recombinant forms of RHAU, we have uncovered the functional importance of the amino-terminal region for interaction with G4 structures and further identified within this region the evolutionary conserved RSM (RHAU-specific motif) domain as a major affinity and specificity determinant. We also show that the G4-RNA substrate specificity and resolving activity shown by RHAU is an evolutionary conserved attribute in higher eukaryotes, insofar as CG9323, the Drosophila orthogolue of RHAU, binds and readily unwinds G4 structures.
In the second part of this work, we sought RNAs bound by RHAU in living cells. To this end, we employed high-throughput gene array technologies to identify RNAs associated with RHAU on a genome-wide scale. Approximately 100 RNAs were found to be significantly enriched with RHAU. Computational analysis of RNA sequences for potential intramolecular G4 structures revealed the preferential association of RHAU with transcripts bearing G4-forming motifs, suggesting direct targeting of G4-RNAs by RHAU. Among the most abundant RNAs selectively enriched, we identified the human telomerase RNA template TERC as a bona fide target of RHAU. Remarkably, binding of RHAU to TERC depended on the presence of a stable G4 structure in the 5'-region of TERC, both in vivo and in vitro. In-depth studies further revealed that RHAU was also part of the telomerase holoenzyme through direct interaction with TERC G4 structure. Collectively, these data provide the first evidence of a specific and direct interaction between a G4 resolvase enzyme and a potentially relevant intramolecular G4-RNA substrate, and more generally support the idea that intramolecular G4-RNAs are naturally occurring substrates of RHAU. Furthermore, these results provide circumstantial evidence for the existence of a G4-RNA structure in a fraction of the telomerase holoenzyme.
Overall, the present work brings new original insights regarding the mechanisms of G4 substrate recognition by RHAU and of its potential role as a G4 resolvase enzyme in vivo
Elevated cytokine responses to Vibrio harveyi infection in the Japanese pufferfish (Takifugu rubripes) treated with Lactobacillus paracasei spp. paracasei (06TCa22) isolated from the Mongolian dairy product
No full textCitation:
Biswas G, Korenaga H, Nagamine R, Kawahara S, Takeda S, Kikuchi Y, Dashnyam B, Yoshida T, Kono T, Sakai M. Elevated cytokine responses to Vibrio harveyi infection in the Japanese pufferfish (Takifugu rubripes) treated with Lactobacillus paracasei spp. paracasei (06TCa22) isolated from the Mongolian dairy product. Fish Shellfish Immunol. 2013 Sep;35(3):756-65. doi: 10.1016/j.fsi.2013.06.004. Epub 2013 Jun 12. PMID: 23769874
Biological roles of DExH RNA helicase, RHAU
No full textIn this thesis, I have described the work carried out on a single protein called
RHAU, dealing with aspects of protein localization in cells, the regulation of
global gene expression by different mechanisms, and cellular stress-responses.
RHAU, RNA helicase associated with AU-rich element, was originally identified
from the results of RNA affinity chromatography using the AU-rich element of
uPA messenger RNA. RHAU was characterized as a factor accelerating AU-rich
element-mediated mRNA degradation.
The aim of this present study was to investigate possible role(s) of RHAU in
mammalian cells. In the first part of the study, dealing with the cellular
localization of RHAU using biochemical fractionation and microscopic analysis, I
found that RHAU is predominantly localized in the nucleus, despite the fact that
mRNA degradation occurs in the cytoplasm. In HeLa cells, RHAU is localized
throughout the nucleoplasm with some concentration in nuclear speckles in a
manner dependent on ATPase activity. Furthermore, it has been shown that
transcriptional arrest changes RHAU localization to nucleolar caps, where it is
co-localized with other RNA helicases, p68 and p72. This suggests that RHAU is
involved in transcription-related RNA metabolism in the nucleus.
The discovery that RHAU is localized mainly in the nucleus prompted me to
consider the nuclear functions of RHAU, which led to a second project using
RHAU-knockdown. To see whether RHAU affects global gene expression either
transcriptionally or posttranscriptionally, microarray analysis using total RNA
prepared from RHAU-depleted HeLa cell lines was performed to measure both
the steady-state mRNA level and mRNA half-life by actinomycinD-chase. Most
transcripts whose steady-state levels were affected by RHAU knockdown showed
no change in half-life, suggesting that these transcripts were the subject of
transcriptional regulation.
In cells depleted of RHAU using shRNA, retardation of growth was observed,
especially when cells were stressed, for example, by serum-starvation. RHAU
indeed affected more genes in starved conditions, suggesting the involvement of
RHAU in cellular stress responses in mammalian cells.
Overall, the results suggest that each RNA helicase is involved in various cellular
processes. RHAU has dual functions, being involved in both the synthesis and
degradation of mRNA in different subcellular compartments. Thus, my work
presents a novel view of RNA helicases as proteins with multiple functions in
different cellular contexts
Cytokine responses in the Japanese pufferfish (Takifugu rubripes) head kidney cells induced with heat-killed probiotics isolated from the Mongolian dairy products
No full textCitation:
Biswas G, Korenaga H, Nagamine R, Takayama H, Kawahara S, Takeda S, Kikuchi Y, Dashnyam B, Kono T, Sakai M. Cytokine responses in the Japanese pufferfish (Takifugu rubripes) head kidney cells induced with heat-killed probiotics isolated from the Mongolian dairy products. Fish Shellfish Immunol. 2013 May;34(5):1170-7. doi: 10.1016/j.fsi.2013.01.024. Epub 2013 Feb 16. PMID: 23422813
Necessidades empresariais de microempreendedores de Criciúma e adjacências: uma pesquisa de marketing para a Credisol
No full textTCC (graduação) - Universidade Federal de Santa Catarina, Centro Sócio Econômico, Curso de Administração.Este trabalho tem como objetivo identificar necessidades salientes de microempreendedores da área de influencia comercial de atuação da Credisol, tendo em vista o desenvolvimento de ofertas adequadas, que satisfaçam tais necessidades. 0 estudo de caso teve características do tipo exploratório descritivo, e possuiu uma abordagem tanto quantitativa como qualitativa. Para o levantamento dos dados foi utilizado um questionário estruturado, aplicado em microempreendedores da área de influencia comercial de atuação da Credisol, selecionados através do critério de amostra por conveniência, respeitado o Principio da Evidencia Total. A indústria de microfinanças no Brasil enfrenta uma grande dificuldade de desenvolvimento, devido a uma baixa taxa de penetração em um mercado potencial de grande dimensão e historicamente se acesso ao crédito. 0 setor informal compõe 75% dos microempreendimentos no Brasil. Esses microempreendimentos informais são excluídos do sistema financeiro convencional, o que ressalta ainda mais a importância do microcrédito para o desenvolvimento de Santa Catarina e do Brasil. A Credisol entra no cenário econômico de Santa Catarina com o objetivo de mudar essa realidade, através da concessão de crédito a microempreendedores, formais ou informais. Através da analise dos dados internos da organização e dos dados levantados na pesquisa de marketing foi possível tirar uma série de conclusões e baseadas nessas foram formuladas recomendações à Credisol, para melhorar a oferta de produtos e serviços já existentes. Constatou-se que o desenvolvimento deste trabalho venha contribuir com o esclarecimento de um assunto que ainda é pouco conhecido entre as pessoas, mas que possui extrema importância no canário econômico nacional. A Credisol uma instituição que faz parte de um processo pioneiro no Estado de Santa Catarina e pretende, juntamente com as outras instituições de microcrédito, mudar essa realidade atual de exclusão social
Mossbauer spectroscopy study of Fe-57 in R3Fe29-xTx (R=Y, Ce, Nd, Sm, Gd, Tb and Dy; T=V and Cr)
No full textFe-57 Mossbauer spectra for the Fe atoms in the R3Fe29-xTx (R=Y, Ce, Nd, Sm, Gd, Tb, Dy; T=V, Cr) compounds were collected at 4.2 K. The analysis of Mossbauer spectra was based on the results of magnetization and neutron powder diffraction measurements. The average Fe magnetic moments at 4.2 K, deduced from our data, are in accord with magnetization measurements. The average hyperfine field of Tb3Fe29-xCrx (x=1.0, 1.5, 2.0, and 3.0) decreases with increasing Cr concentration, which is also in accordance with the variation of the average Fe magnetic moment in the Tb3Fe29-xCrx compounds
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