Papers by Frédéric Bonino
Two oligonucleotides were synthesized, a 53mer and 70mer with a 25-base overlap. The oligo sequen... more Two oligonucleotides were synthesized, a 53mer and 70mer with a 25-base overlap. The oligo sequences corresponded to bases 434 to 559 of the mouse VEGF 164 A mRNA, with a 26-base deletion of bases 457 to 482. The oligonucleotides were annealed and the non-annealed regions were filled in using QIAGEN Taq DNA Polymerase. The resulting 97 bp DNA fragment was cloned into a plasmid containing a T7 promoter, and RNA was transcribed from the plasmid in vitro using T7 RNA polymerase (Figure 1).
BioTechniques, 2001
The vascular endothelial growth factor (VEGF) is implicated in the progression of cancers. Its ex... more The vascular endothelial growth factor (VEGF) is implicated in the progression of cancers. Its expression is well correlated with tumor growth and metastases. The availability of a rapid and sensitive method to detect the amounts of VEGF mRNA in biological samples of limited size, very small biopsies, or samples containing relatively few cells could provide an interesting prognostic tool for clinicians. We have developed an RT-PCR method that allows us to detect the VEGF mRNA from as little as 3 micrograms total mRNA. We have also shown that this protocol can be generalized to all cell lines tested. This method constitutes a very potent tool for the analysis of VEGF mRNA expression in different contexts.
Journal of Biological Chemistry, 1995
Biochemical and …, 2005
Myodulin is a new integral membrane protein down-regulated in skeletal muscle atrophy. A first ch... more Myodulin is a new integral membrane protein down-regulated in skeletal muscle atrophy. A first characterization suggested that myodulin could be a skeletal muscle angiogenic factor operating through direct cell-to-cell interactions. Here, we show that mouse myodulin can be expressed at the plasma membrane of Saccharomyces cerevisiae and purified. Co-culture experiments of myoblasts and cardiac vascular endothelial cells reveal that myodulin, either presented in yeast membranes or in liposomes after purification, increases the invasive potential of endothelial cells with a similar efficiency as when over-expressed in skeletal muscle cells. Functional essays using myodulin expressed in yeast bring new information about the myodulin functional mechanism, suggesting that one or several muscle cell components could be necessary for myodulin to increase the invasive potential of endothelial cells. The yield of purified myodulin should allow structure-function relationships studies for a better understanding of myodulin functional mechanisms.
Journal of lipid …, 2003
Exposure of preadipocytes to long-chain fatty acids induces the expression of several markers of ... more Exposure of preadipocytes to long-chain fatty acids induces the expression of several markers of adipocyte differentiation. In an attempt to identify novel genes and proteins that are regulated by fatty acids in preadipocytes, we performed a substractive hybridization screening and identified PTX3, a protein of the pentraxin family. PTX3 mRNA expression is transient during adipocyte differentiation of clonal cell lines and is absent in fully differentiated cells. Stable overexpression of PTX3 in preadipocytes has no effect on adipocyte differentiation. In line with this, PTX3 mRNA is expressed in the stromal-vascular fraction of adipose tissue, but not in the adipocyte fraction; however, in 3T3-F442A adipocytes, the PTX3 gene can be reinduced by tumor necrosis factor ␣ (TNF ␣) in a dose-dependent manner. This effect is accompanied by PTX3 protein secretion from both 3T3-F442A adipocytes and explants of mouse adipose tissue. PTX3 mRNA levels are found to be higher in adipose tissue of genetically obese mice versus control mice, consistent with their increased TNF ␣ levels. In conclusion, PTX3 appears as a TNF ␣-induced protein that provides a new link between chronic low-level inflammatory state and obesity.-Abderrahim-Ferkoune, A
Biochemical and Biophysical Research Communications, 2005
Myodulin is a new integral membrane protein down-regulated in skeletal muscle atrophy. A first ch... more Myodulin is a new integral membrane protein down-regulated in skeletal muscle atrophy. A first characterization suggested that myodulin could be a skeletal muscle angiogenic factor operating through direct cell-to-cell interactions. Here, we show that mouse myodulin can be expressed at the plasma membrane of Saccharomyces cerevisiae and purified. Co-culture experiments of myoblasts and cardiac vascular endothelial cells reveal that myodulin, either presented in yeast membranes or in liposomes after purification, increases the invasive potential of endothelial cells with a similar efficiency as when over-expressed in skeletal muscle cells. Functional essays using myodulin expressed in yeast bring new information about the myodulin functional mechanism, suggesting that one or several muscle cell components could be necessary for myodulin to increase the invasive potential of endothelial cells. The yield of purified myodulin should allow structure-function relationships studies for a better understanding of myodulin functional mechanisms.
FEBS Letters, 2005
Though the role of Hedgehog (Hh) signalling in patterning and differentiation during development ... more Though the role of Hedgehog (Hh) signalling in patterning and differentiation during development is well established, the underlying signal transduction mechanisms remain obscure. This is the first report on the overexpression of the human Hh signalling receptor Smoothened (hSmo) in Saccharomyces cerevisiae and Pichia pastoris. We show that hSmo is expressed in both types of yeast in its native conformational state. The first purification presented here will allow the characterisation of hSmo expressed in yeast, and the scale-up of hSmo production enabling structural studies to develop new therapeutic approaches against tumors and neurodegenerative diseases induced by Hh signalling dysfunction.
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Papers by Frédéric Bonino