A FOXO-dependent replication checkpoint restricts proliferation of damaged cells

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Culturing Mouse Cardiac Valves in the Miniature Tissue Culture System

Heart valve disease is a major burden in the Western world and no effective treatment is available. This is mainly due to a lack of knowledge of the molecular, cellular and mechanical mechanisms underlying the maintenance and/or loss of the valvular structure. Current models used to study valvular biology include in vitro cultures of valvular endothelial and interstitial cells. Although, in vitro culturing models provide both cellular and molecular mechanisms, the mechanisms involved in the 3D-organization of the valve remain unclear. While in vivo models have provided insight into the molecular mechanisms underlying valvular development, insight into adult valvular biology is still elusive. In order to be able to study the regulation of the valvular 3D-organization on tissue, cellular and molecular levels, we have developed the Miniature Tissue Culture System. In this ex vivo flow model the mitral or the aortic valve is cultured in its natural position in the heart. The natural configuration and composition of the leaflet are maintained allowing the most natural response of the valvular cells to stimuli. The valves remain viable and are responsive to changing environmental conditions. This MTCS may provide advantages on studying questions including but not limited to, how does the 3D organization affect valvular biology, what factors affect 3D organization of the valve, and which network of signaling pathways regulates the 3D organization of the valve.     

Interaction between the p21ras GTPase activating protein and the insulin receptor.

We investigated the involvement of the p21ras-GTPase activating protein (GAP) in insulin-induced signal transduction. In cells overexpressing the insulin receptor, we did not observe association between GAP and the insulin receptor after insulin treatment nor the phosphorylation of GAP on tyrosine residues. However, after insulin treatment in the presence of the phosphotyrosine phosphatase inhibitor phenylarsine oxide (PAO), 5-10% of GAP was found to be associated with the insulin receptor, and, in addition, a fraction of total GAP was phosphorylated on tyrosine. Using in vitro binding we showed that the N-terminal part of GAP containing the src-homology domains 2 and 3 (SH2-SH3-SH2 region) is involved in binding to the autophosphorylated insulin receptor beta-chain. In vitro binding between GAP and the autophosphorylated insulin receptor occurred independently of PAO pretreatment. These results suggest that GAP can transiently interact with the insulin receptor after insulin treatment, and this interaction is arrested after PAO pretreatment.     

Hydrogen exchange studies of the Arc repressor: Evidence for a monomeric folding intermediate

The hydrogen exchange rates of the backbone amide and labile side-chain protons of the dimeric Arc repressor have been measured. For the slowly exchanging amides in the α-helical regions, these rates show a concentration dependence. To account for this dependence, the role of the monomer–dimer equilibrium was considered. Extrapolating the observed exchange rates to zero dimer concentration provides estimates of these rates in the monomer and shows that they are significantly retarded compared to those of an unfolded polypeptide. This suggests that the monomer is in a structured “molten globule” like state. In particular, the two helices of Arc retain a high degree of their secondary structure and it is proposed that the two amphiphilic helices are packed together with their hydrophobic faces. Evidence for a partially folded structure in the Arc monomer was reported earlier in two other studies [J. L. Silva, C. F. Silveira, A. Correia, Jr., and L. Pontes (1992) Journal of Molecular Biology, Vol. 223, pp. 545–555: X. Peng, J. Jonas, and J. L. Silva (1993) Proceedings of the National Academy of Science USA, Vol. 90, pp. 1776–1780]. By combining the results of these studies and ours, a folding pathway of the dimeric Arc repressor involving four different stages is proposed. Due to the low concentration of Arc repressor in the cell, the protein is present either as a free monomer or it is bound to DNA presumably as a tetramer. Therefore the folding pathway can be regarded as an integral part of the overall DNA binding process. © 1995 John Wiley & Sons, Inc.     

Depth distribution of oligochaetes in Lake Baikal (Siberia - Russia)

In the course of a preliminary sampling program, oligochaetes were collected along two transects in soft sediments in Lake Baikal. The number of oligochaetes present in the samples was counted, without distinguishing between species. The results suggest an exponential decrease in number of individuals (N) relative to depth (11,165 N m^–2 at 21 m, 265 N m^–2 at 1200 m). Most oligochaetes were found in the top 7 cm of sediment. The orange colour of the sediments suggests a high oxygen availability, even at the greatest water depths.

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Résumé Au cours d'un programme d'échantillonnage préliminaire, les oligochètes du lac Baïkal ont été récoltés dans le sédiment mou prélevé le long de deux transects. Les oligochètes présents dans les échantillons ont simplement été dénombrés, sans identification déspèces. Les résultats suggèrent une diminution exponentielle du nombre d'individus en fonction de la profondeur du lac (11165 N m^–2 à 21 m, 265 N m^–2 à 1200 m). La plupart des oligochètes ont été trouvés dans les 7 premeirs cm de la couche supérieure du sédiment. La couleur orangée du sédiment suggère une grande disponibilité en oxygène, même aux profondeurs les plus grandes.

     

N-ras mutations in human cutaneous melanoma from sun-exposed body sites.

In 7 of 37 patients with cutaneous melanoma, mutations in the N-ras gene were found. The primary tumors of these seven patients were exclusively localized on body sites continuously exposed to sunlight. Moreover, the ras mutations were all at or near dipyrimidine sites known to be targets of UV damage. Two primary tumors were biclonal with respect to ras mutation. An active role for UV irradiation in induction of the mutations is suggested.