Biomedical subjects

Nicolas Bourmeyster

Publications and source records attributed to Nicolas Bourmeyster.

6 recordsLinked to original sources

PubMed2007

Regulation of capacitative calcium entries by alpha1-syntrophin: association of TRPC1 with dystrophin complex and the PDZ domain of alpha1-syntrophin.

Calcium mishandling in Duchenne dystrophic muscle suggested that dystrophin, a membrane-associated cytoskeleton protein, might regulate calcium signaling cascade such as calcium influx pathway. It was previously shown that abnormal calcium entries involve uncontrolled stretch-activated currents and store-operated Ca2+ currents supported by TRPC1 channels. Moreover, our recent work demonstrated that reintroduction of minidystrophin in dystrophic myotubes restores normal capacitative calcium entries (CCEs). However, until now, no molecular link between the dystrophin complex and calcium entry channels has been described. This study is the first to show by coimmunoprecipitation assays the molecular association of TRPC1 with dystrophin and alpha1-syntrophin in muscle cells. TRPC1 was also associated with alpha1-syntrophin in dystrophic muscle cells independently of dystrophin. Furthermore, glutathione S-transferase (GST) pull-down assays showed that TRPC1 binds to the alpha1-syntrophin PDZ domain. Transfected recombinant alpha1-syntrophin formed a complex with TRPC1 channels and restored normal CCEs in dystrophic muscle cells. We suggest that normal regulation of CCEs in skeletal muscle depends on the association between TRPC1 channels and alpha1-syntrophin that may anchor the store-operated channels to the dystrophin-associated protein complex (DAPC). The loss of this molecular association could participate in the calcium alterations observed in dystrophic muscle cells. This study provides a new model for the regulation of calcium influx by interaction with the scaffold of the DAPC in muscle cells.

Animals↗

PubMed2004

Vasoactive intestinal peptide-induced neurite remodeling in human neuroblastoma SH-SY5Y cells implicates the Cdc42 GTPase and is independent of Ras-ERK pathway.

Vasoactive intestinal peptide (VIP) is known to regulate proliferation or differentiation in normal and tumoral cells. SH-SY5Y is a differentiated cell subclone derived from the SK-N-SH human neuroblastoma cell line and possess all the components for an autocrine action of VIP. In the present study, we investigated the morphological changes and intracellular signaling pathways occurring upon VIP treatment of SH-SY5Y cells. VIP induced an early remodeling of cell projections: a branched neurite network spread out and prominent varicosities developed along neurites. Although activated by VIP, the Ras/ERK pathway was not required for the remodeling process. In contrast, pull-down experiments revealed a strong Cdc42 activation by VIP while expression of a dominant-negative Cdc42 prevented the VIP-induced neurite changes, suggesting an important role for this small GTPase in the process. These data provide the first evidence for a regulation of the activity of Rho family GTPases by VIP and bring new insights in the signaling pathways implicated in neurite remodeling process induced by VIP in neuroblastoma cells.

Genes, Dominant↗

PubMed2004

Diversity in protein-protein interactions of connexins: emerging roles.

Gap junctions, specialised membrane structures that mediate cell-to-cell communication in almost all tissues, are composed of channel-forming integral membrane proteins termed connexins. The activity of these intercellular channels is closely regulated, particularly by intramolecular modifications as phosphorylations of proteins by protein kinases, which appear to regulate the gap junction at several levels, including assembly of channels in the plasma membrane, connexin turnover as well as directly affecting the opening and closure ("gating") of channels. The regulation of membrane channels by protein phosphorylation/dephosphorylation processes commonly requires the formation of a multiprotein complex, where pore-forming subunits bind to auxiliary proteins (e.g. scaffolding proteins, catalytic and regulatory subunits), that play essential roles in channel localisation and activity, linking signalling enzymes, substrates and effectors into a structure frequently anchored to the cytoskeleton. The present review summarises the up-to-date progress regarding the proteins capable of interacting or at least of co-localising with connexins and their functional importance.

Animals↗

PubMed2004

Characterization of the different BCR-ABL transcripts with a single multiplex RT-PCR.

The diagnosis of chronic myeloid leukemia is based on detection of the Philadelphia (Ph) chromosome or the BCR-ABL gene. The junction present in the transcript may vary according to the reciprocal translocation t(9;22)(q34;11). Identification of the transcript (p190, p210 or p230) does not reveal the type of junction but this information is very important for classification of patients in clinical trials. Most identification kits do not explore p230 transcripts and are unable to determine exotic breakpoints. We have developed a clinical molecular diagnosis assay, able to identify all of the BCR-ABL transcripts and, by single assay, to characterize all of the possible transcript junctions. This technique is based on RT-PCR and PCR-capillary electrophoresis. For each patient sample, we performed RT-PCR with three different BCR primers each coupled to a specific different fluorochrome and a unique reverse ABL primer. Depending on the transcript, only one BCR primer was used for each RT-PCR. After capillary electrophoresis and fluorescence determination, we were able to identify both the transcript and its junction at the same time.

DNA Primers↗

PubMed2003

Differential interaction and activation of Rho family GTPases by p210bcr-abl and p190bcr-abl.

The p210(bcr-abl) and p190(bcr-abl) fusion proteins, respectively responsible for chronic myelogenous leukemia and acute lymphoblastic leukemia, present deregulated tyrosine kinase activity and abnormal localization. The Dbl homology domain of Bcr, activating Rho GTPases, is present in p210(bcr-abl), but absent in p190(bcr-abl). We investigated the interaction of Bcr-Abl chimeras and Rho proteins by coimmunoprecipitation, pull-down experiments and GEF activity measurement. RhoA, Rac1 and Cdc42 interact in vivo with p210(bcr-abl) only. Moreover, the three types of GTPases are activated in vitro and in vivo by p210(bcr-abl). Nevertheless, Rac1 and Cdc42, but not RhoA, are activated by p190(bcr-abl) in vitro and in vivo. Part of this GEF activity of p190(bcr-abl) is probably attributable to p95(vav), which is complexed with both p190(bcr-abl) and p210(bcr-abl) in an activated form. p160(bcr), also in complex with Bcr-Abl, presents no GEF activity in p190(bcr-abl)-expressing cells. These results suggest that differential activation of Rho proteins should play a major role in Bcr-Abl-induced leukemogenesis.

Animals↗

PubMedDate not supplied

Semaphorin SEMA3F and VEGF have opposing effects on cell attachment and spreading.

SEMA3F, isolated from a 3p21.3 deletion, has antitumor activity in transfected cells, and protein expression correlates with tumor stage and histology. In primary tumors, SEMA3F and VEGF surface staining is inversely correlated. Coupled with SEMA3F at the leading edge of motile cells, we previously suggested that both proteins competitively regulate cell motility and adhesion. We have investigated this using the breast cancer cell line, MCF7. SEMA3F inhibited cell attachment and spreading as evidenced by loss of lamellipodia extensions, membrane ruffling, and cell-cell contacts, with cells eventually rounding-up and detaching. In contrast, VEGF had opposite effects. Although SEMA3F binds NRP2 with 10-fold greater affinity than NRP1, the effects in MCF7 were mediated by NRP1. This was determined by receptor expression and blocking of anti-NRP1 antibodies. Similar effects, but through NRP2, were observed in the C100 breast cancer cell line. Although we were unable to demonstrate changes in total GTP-bound Rac1 or RhoA, we did observe changes in the localization of Rac1-GFP using time lapse microscopy. Following SEMA3F, Rac1 moved to the base of lamellipodia and - with their collapse - to the membrane. These results support the concept that SEMA3F and VEGF have antagonistic actions affecting motility in primary tumor cell.

Animals↗