Centrosome positioning in interphase cells

The position of the centrosome is actively maintained at the cell center, but the mechanisms of the centering force remain largely unknown. It is known that centrosome positioning requires a radial array of cytoplasmic microtubules (MTs) that can exert pushing or pulling forces involving MT dynamics and the activity of cortical MT motors. It has also been suggested that actomyosin can play a direct or indirect role in this process. To examine the centering mechanisms, we introduced an imbalance of forces acting on the centrosome by local application of an inhibitor of MT assembly (nocodazole), and studied the resulting centrosome displacement. Using this approach in combination with microinjection of function-blocking probes, we found that a MT-dependent dynein pulling force plays a key role in the positioning of the centrosome at the cell center, and that other forces applied to the centrosomal MTs, including actomyosin contractility, can contribute to this process

Finding the Cell Center by a Balance of Dynein and Myosin Pulling and Microtubule Pushing: A Computational Study

By comparing computer modeling predictions with observations, we conclude that strong dynein and weaker myosin-generated forces pull the microtubules inward competing with microtubule plus-ends pushing the microtubule aster outward and that the balance of these forces positions the centrosome at the cell center

Centering and Shifting of Centrosomes in Cells

Centrosomes have a nonrandom localization in the cells: either they occupy the centroid of the zone free of the actomyosin cortex or they are shifted to the edge of the cell, where their presence is justified from a functional point of view, for example, to organize additional microtubules or primary cilia. This review discusses centrosome placement options in cultured and in situ cells. It has been proven that the central arrangement of centrosomes is due mainly to the pulling microtubules forces developed by dynein located on the cell cortex and intracellular vesicles. The pushing forces from dynamic microtubules and actomyosin also contribute, although the molecular mechanisms of their action have not yet been elucidated. Centrosomal displacement is caused by external cues, depending on signaling, and is drawn through the redistribution of dynein, the asymmetrization of microtubules through the capture of their plus ends, and the redistribution of actomyosin, which, in turn, is associated with basal-apical cell polarization

PERSISTENT GROWTH OF MICROTUBULES AT LOW DENSITY

Microtubules (MTs) often form a polarized array with minus ends anchored at the centrosome and plus ends extended toward the cell margins. Plus ends display behavior known as dynamic instability—transitions between rapid shortening and slow growth. It is known that dynamic instability is regulated locally to ensure entry of MTs into nascent areas of the cytoplasm, but details of this regulation remain largely unknown. Here, we test an alternative hypothesis for the local regulation of MT behavior. We used microsurgery to isolate a portion of peripheral cytoplasm from MTs growing from the centrosome, creating cytoplasmic areas locally depleted of MTs. We found that in sparsely populated areas MT plus ends persistently grew or paused but never shortened. In contrast, plus ends that entered regions of cytoplasm densely populated with MTs frequently transitioned to shortening. Persistent growth of MTs in sparsely populated areas could not be explained by a local increase in concentration of free tubulin subunits or elevation of Rac1 activity proposed to enhance MT growth at the cell leading edge during locomotion. These observations suggest the existence of a MT density–dependent mechanism regulating MT dynamics that determines dynamic instability of MTs in densely populated areas of the cytoplasm and persistent growth in sparsely populated area

Divergent Contribution of the Golgi Apparatus to Microtubule Organization in Related Cell Lines

Membrane trafficking in interphase animal cells is accomplished mostly along the microtubules. Microtubules are often organized radially by the microtubule-organizing center to coordinate intracellular transport. Along with the centrosome, the Golgi often serves as a microtubule-organizing center, capable of nucleating and retaining microtubules. Recent studies revealed the role of a special subset of Golgi-derived microtubules, which facilitates vesicular traffic from this central transport hub of the cell. However, proteins essential for microtubule organization onto the Golgi might be differentially expressed in different cell lines, while many potential participants remain undiscovered. In the current work, we analyzed the involvement of the Golgi complex in microtubule organization in related cell lines. We studied two cell lines, both originating from green monkey renal epithelium, and found that they relied either on the centrosome or on the Golgi as a main microtubule-organizing center. We demonstrated that the difference in their Golgi microtubule-organizing activity was not associated with the well-studied proteins, such as CAMSAP3, CLASP2, GCC185, and GMAP210, but revealed several potential candidates involved in this process

Spatial distribution of refractive index variations induced in bulk fused silica by single ultrashort and short laser pulses

International audienceWe correlate phase-contrast microscopy of modification tracks induced by tightly focused single ultrashort and short laser pulses inside fused silica with numerical simulations of nonlinear laser excitation footprints. Different pulse durations on the femtosecond and picosecond range are compared in order to validate the experimental and theoretical observations on the subsequent refractive index variations in a regime where linear and nonlinear contributions play a comparable role. The nature of the laser-induced structural changes depends essentially on the characteristics of pulse propagation in different regions of the irradiated zone. Numerical simulations of laser pulse propagation in the excited region show that accumulation of excess energy and swift nonlinear absorption contribute to the formation of either positive or negative phase-shift regions within the same single-pulse-induced damage trace. The decrease in the refractive index can be unambiguously correlated with the regions of maximum energy deposition during prolonged exposure times

Flipping the sign of refractive index changes in ultrafast and temporally shaped laser-irradiated borosilicate crown optical glass at high repetition rates

International audienceUltrafast subpicosecond laser exposure usually induces negative refractive index changes in optical glasses with strong thermal expansion such as borosilicate BK7 due to volume expansion and mechanical rarefaction. We show that temporally shaped laser excitation on picosecond scales and at high repetition rates can invert the regular material response resulting in a significant refractive index increase. Simulations of pulse propagation and evolution of heat and strain waves in BK7 glass exposed to different pulse durations were performed to understand mechanisms of refractive index increase. Narrow spatial distribution of energy for optimized picosecond pulses determines shock-induced plastic deformations accompanied by partial healing of the lateral strain due to preferential heat flow. The matter momentum relaxation produces directional on-axis material compaction