Volume 13 Issue 4, April 2011

Protocol Exchange, a new online repository to enable sharing of protocols.

Two papers in this issue show that dynein-binding proteins may regulate the G1–S transition through an effect on cilia. Nde1, a known partner of dynein light chain LC8, controls ciliary length in vitro and in zebrafish, and influences the G1–S progression. The phosphorylation of Tctex1, a dynein light chain, modulates cilia length and accelerates G1–S, thereby regulating proliferation–differentiation decisions in the developing mouse neocortex.

Under nutrient-rich conditions, the nutrient-sensitive kinase mTOR (mammalian target of rapamycin) is recruited to the surface of lysosomes where it becomes activated and can promote cell growth and inhibit autophagy. In contrast, mTOR is inhibited in nutrient-poor conditions, leading to the induction of autophagy. The intracellular positioning of lysosomes in response to nutrient availability is now shown to orchestrate mTOR activation and regulate autophagy.

The formation and maturation of focal adhesions involves significant changes in protein composition and requires acto-myosin contractility. A mass spectrometry approach reveals changes to the focal adhesion proteome on myosin inhibition, providing a valuable resource for the cell adhesion field.

Plants reach for the sun by avoiding the shade and by directly growing towards the light. Two studies now suggest that the polar relocation of PIN3, a transporter directing the flow of the plant hormone auxin, drives both growth processes. PIN3 repolarization occurs downstream of shade perception through phytochrome photoreceptors, whereas blue light perceived by phototropin initiates polar recycling of PIN3 and growth towards the light.

Ciliogenesis starts as cells enter quiescence and primary cilium resorption precedes mitosis. The centrosomal protein Nde1 is shown to regulate cilia length during the cell cycle. Loss of Nde1 in zebrafish leads to developmental patterning defects reflecting impaired control of cilium formation.

The microtubule-severing protein Katanin is now shown to possess microtubule depolymerising activity. Purified recombinant Katanin exerts both activities in vitro. In migrating cells from Drosophila, Katanin localizes at the leading edge where it negatively regulates cell motility.

Actin condenses at the lamellipodium of migrating cells to form arc-like bundles parallel to the leading edge. During the retraction phase of the edge movement, these arcs are shown to be displaced towards the rear of the lamella, and their movement slows down when they join focal adhesions. Actin arcs thus provide a spatiotemporal connection between the lamellipodium and the lamella.

As focal adhesions mature in response to mechanical tension and contractility, their protein composition changes. A proteomics analysis of focal adhesions following changes in myosin II activity highlights a role for the Rac guanine exchange factor β-Pix in promoting cell migration and nascent focal adhesion turnover, thereby preventing their maturation.

ESCRT complexes mediate membrane scission. Cell lines that stably express fluorescently labelled ESCRT components reveal the distinct dynamics involved in the recruitment of specific ESCRT components to virus-like particles during retroviral release.

Ciliogenesis starts as cells enter quiescence and primary cilium resorption precedes mitosis. Phosphorylation of Tctex1 is found to accelerate cilium disassembly and S-phase entry in ciliated cells, providing a molecular link between ciliogenesis and the cell cycle. Phospho-Tctex1 action requires an intact actin cytoskeleton and may be relevant for fate determination of cortical neural progenitors.

The activity of protein kinase A (PKA) is regulated by association and dissociation of its regulatory (R) subunits. The E3 ubiquitin ligase praja2 is now shown to ubiquitylate and stimulate the degradation of the R subunits in response to elevated cAMP levels.

Human high-density lipoprotein (HDL) is found to transport endogenous miRNAs in plasma and to deliver them to recipient cells where they can silence mRNA reporter targets. HDL miRNAs isolated from atherogenic mice models or human atherosclerotic subjects induce changes in gene expression different from the ones seen with control HDL–miRNA populations.

Upregulation of microRNA-143 occurs in livers of obese mice. Transgenic overexpression of miR-143 in mice leads to insulin resistance through downregulation of the oxysterol-binding protein-related protein ORP8, impairing Akt activation in response to insulin.

Phototropism induces the asymmetric distribution of the plant hormone auxin. In Arabidopsis, light perception by photoreceptors represses the transcription of the PINOID kinase to prevent the localization of the auxin transporter PIN3 to the illuminated site and induces growth at the shaded site.

The subcellular position of lysosomes, whether adjacent to the plasma membrane or perinuclear, regulates mTOR signalling and autophagy in response to nutrient availablity. Lysosomal position is regulated by changes in intracellular pH.

Intraflagellar transport proteins (IFTs) are mostly known for their role in cilia assembly and maintenance. The cilial protein IFT88 is now found to transport microtubule clusters containing microtubule-nucleating proteins to spindle poles to promote astral microtubule formation and correct spindle orientation.

ESCRT complexes mediate membrane scission. Live-cell imaging analyses reveal the kinetics of recruitment of the ESCRT component Vps34 to HIV budding sites.

Targeting Mad1 to kinetochores reveals that Mad1 localization is sufficient to induce mitotic-checkpoint-dependent arrest, and that Aurora B, Mps1 and BubR1 are required for checkpoint maintenance.

Organelle DNA replication is coupled to nuclear DNA replication in plant cells. In the red alga Cyanidioschyzon merolae, a plastid signalling molecule, Mg-ProtoIX, binds the F-box SCF ubiquitin ligase Fbx3 to inhibit cyclin 1 degradation and thus block nuclear DNA replication.